@article {3743, title = {A dimer between monomers and hexamers-Oligomeric variations in glucosamine-6-phosphate deaminase family.}, journal = {PLoS One}, volume = {18}, year = {2023}, month = {2023}, pages = {e0271654}, abstract = {

In bacteria that live in hosts whose terminal sugar is a sialic acid, Glucosamine-6-phosphate deaminase (NagB) catalyzes the last step in converting sialic acid into Fructose-6-phosphate. These bacteria then use the Fructose-6-phosphate as an energy source. The enzyme NagB exists as a hexamer in Gram-negative bacteria and is allosterically regulated. In Gram-positive bacteria, it exists as a monomer and lacks allosteric regulation. Our identification of a dimeric Gram-negative bacterial NagB motivated us to characterize the structural basis of two closely related oligomeric forms. We report here the crystal structures of NagB from two Gram-negative pathogens, Haemophilus influenzae (Hi) and Pasturella multocida (Pm). The Hi-NagB is active as a hexamer, while Pm-NagB is active as a dimer. Both Hi-NagB and Pm-NagB contain the C-terminal helix implicated as essential for hexamer formation. The hexamer is described as a dimer of trimers. In the Pm-NagB dimer, the dimeric interface is conserved. The conservation of the dimer interface suggests that the three possible oligomeric forms of NagB are a monomer, a dimer, and a trimer of dimers. Computational modeling and MD simulations indicate that the residues at the trimeric interface have less stabilizing energy of oligomer formation than those in the dimer interface. We propose that Pm-NagB is the evolutionary link between the monomer and the hexamer forms.

}, keywords = {Aldose-Ketose Isomerases, Bacterial Proteins, Haemophilus influenzae, N-Acetylneuraminic Acid, Pasteurella multocida, Polymers}, issn = {1932-6203}, doi = {10.1371/journal.pone.0271654}, author = {Srinivasachari, Sathya and Tiwari, Vikas R and Kharbanda, Tripti and Sowdamini, Ramanathan and Subramanian, Ramaswamy} } @article {3717, title = {Systematic review of articular cartilage derived chondroprogenitors for cartilage repair in animal models.}, journal = {J Orthop}, volume = {35}, year = {2023}, month = {2023 Jan}, pages = {43-53}, abstract = {

PURPOSE OF RESEARCH: The potential for cartilage repair using articular cartilage derived chondroprogenitors has recently gained popularity due to promising results from in-vitro and in-vivo studies. Translation of results from in-vitro to a clinical setting requires a sufficient number of animal studies displaying significant positive outcomes. Thus, this systematic review comprehensively discusses the available literature (January 2000-March 2022) on animal models employing chondroprogenitors for cartilage regeneration, highlighting the results and limitations associated with their use.As per Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines, a web-based search of PubMed and SCOPUS databases was performed for the following terminologies: "chondroprogenitors", "cartilage-progenitors", and "chondrogenic-progenitors", which yielded 528 studies. A total of 12 studies met the standardized inclusion criteria, which included chondroprogenitors derived from hyaline cartilage isolated using fibronectin adhesion assay (FAA) or migratory assay from explant cultures, further analyzing the role of chondroprogenitors using in-vivo animal models.

PRINCIPAL RESULTS: Analysis revealed that FAA chondroprogenitors demonstrated the ability to attenuate osteoarthritis, repair chondral defects and form stable cartilage in animal models. They displayed better outcomes than bone marrow-derived mesenchymal stem cells but were comparable to chondrocytes. Migratory chondroprogenitors also demonstrated superiority to BM-MSCs in terms of higher chondrogenesis and lower hypertrophy, although a direct comparison to FAA-CPs and other cell types is warranted.

MAJOR CONCLUSIONS: Chondroprogenitors exhibit superior properties for chondrogenic repair; however, limited data on animal studies necessitates further studies to optimize their use before clinical translation for neo-cartilage formation.

}, issn = {0972-978X}, doi = {10.1016/j.jor.2022.10.012}, author = {Vinod, Elizabeth and Padmaja, Kawin and Ramasamy, Boopalan and Sathishkumar, Solomon} } @article {2881, title = {Age-stratified adeno-associated virus serotype 3 neutralizing and total antibody prevalence in hemophilia A patients from India.}, journal = {J Med Virol}, volume = {94}, year = {2022}, month = {2022 Sep}, pages = {4542-4547}, abstract = {

Gene therapy using an adeno-associated virus (AAV) vector offers a new treatment option for individuals with monogenetic disorders. The major bottleneck is the presence of pre-existing anti-AAV antibodies, which impacts its use. Even very low titers of neutralizing antibodies (NAb) to capsids from natural AAV infections have been reported to inhibit the transduction of intravenously administered AAV in animal models and are associated with limited efficacy in human trials. Assessing the level of pre-existing NAb is important for determining the primary eligibility of patients for AAV vector-based gene therapy clinical trials. Techniques used to screen AAV-antibodies include AAV capsid enzyme-linked immunosorbent assay\ (ELISA) and transduction inhibition assay (TIA) for detecting total capsid-binding (TAb) and Nab, respectively. In this study, we screened 521 individuals with hemophilia A from India for TAb and NAb using ELISA and TIA, respectively. The prevalence of TAb and NAb in hemophilia A patients from India were 96\% and 77.5\%, respectively. There was a significant increase in anti-AAV3 NAb prevalence with age in the hemophilia A patient group from India. There was a trend in anti-AAV3 TAb positivity between the pediatric age group (94.4\%) and the adult age group (97.4\%).

}, keywords = {Adult, Animals, Antibodies, Neutralizing, Antibodies, Viral, Child, Dependovirus, Genetic Vectors, Hemophilia A, Humans, Prevalence, Serogroup}, issn = {1096-9071}, doi = {10.1002/jmv.27859}, author = {Daniel, Hubert D-J and Kumar, Sanjay and Kannangai, Rajesh and J, Farzana and Joel, Joseph N and Abraham, Aby and Lakshmi, Kavitha M and Agbandje-McKenna, Mavis and Coleman, Kirsten E and Srivastava, Arun and Srivastava, Alok and Abraham, Asha M} } @article {2885, title = {An Amphiphilic Double-Brush Polymer Hydrogel for Sustained Release of Small Molecules and Biologics: Insulin Delivering-Hydrogel to Control Hyperglycemia}, year = {2022}, doi = { https://doi.org/10.1002/cnma.202200184}, author = {Dhayani, Ashish and Bej, Sujoy and Mudnakudu-Nagaraju, Kiran K and Chakraborty, Saheli and Srinath, Preetham and Kumar, Ashok H and PS, Ann Maria and Kristi, Anand and Ramakrishnan, S and Vemula PK} } @article {2504, title = {Assessment of the inherent chondrogenic potential of human articular cartilage-derived chondroprogenitors in pellet culture using a novel whole pellet processing approach.}, journal = {J Orthop}, volume = {31}, year = {2022}, month = {2022 May-Jun}, pages = {45-51}, abstract = {

Purpose: Cartilage-derived chondroprogenitors have been reported to possess the biological potential for cartilage repair. However, its inherent chondrogenic potential in pellet culture needs evaluation. In-vitro cartilage regeneration models based on pellet cultures have been employed to evaluate the chondrogenic potential of stem cells. Evaluation of the degree of differentiation routinely involves paraffin embedding, sectioning, and immunohistochemical staining of the pellet. However, since chondrogenic differentiation is commonly non-uniform, processing random sections could lead to inaccurate conclusions. The study aimed at assessing the inherent lineage bias of chondroprogenitors with and without chondrogenic induction, using a novel whole pellet processing technique.

Methods: Human chondroprogenitors (n=3) were evaluated for MSC markers and processed in pellet cultures either with stromal medium (uninduced) or chondrogenic differentiation medium (induced) for 28 days. The whole pellets and the conventional paraffin-embedded sectioned pellets were subjected to Collagen type II immunostaining and assessed using confocal laser microscopy. The staining intensities of the whole pellet were compared to the paraffin sections and revalidated using qRT-PCR for COL2A1 expression.

Results: Uninduced and induced pellets displayed Collagen type II in all the layers with comparable fluorescence intensities. COL2A1 expression in both pellets was comparable to confocal results. The study demonstrated that uninduced chondroprogenitors in pellet culture possess promising inherent chondrogenic potential. Confocal imaging of whole pellets displayed different degrees of chondrogenic differentiation in the entire pellet, thus its probable in-vivo behavior.

Conclusion: The novel approach presented in this study could serve as an efficient in-vitro alternative for understanding translational application for cartilage repair.

}, issn = {0972-978X}, doi = {10.1016/j.jor.2022.03.007}, author = {Johnson, Noel Naveen and Amirtham, Soosai Manickam and Sandya Rani, B and Sathishkumar, Solomon and Rebekah, Grace and Vinod, Elizabeth} } @article {2466, title = {The CCR5 Gene Edited CD34+CD90+ Hematopoietic Stem Cell Population Serves as an Optimal Graft Source for HIV Gene Therapy}, journal = {Front. Immunol}, year = {2022}, abstract = {

Transplantation of allogenic hematopoietic stem and progenitor cells (HSPCs) with C-C chemokine receptor type 5 (CCR5) Δ32 genotype generates HIV-1 resistant immune cells. CCR5 gene edited autologous HSPCs can be a potential alternative to hematopoietic stem cell transplantation (HSCT) from HLA-matched CCR5 null donor. However, the clinical application of gene edited autologous HSPCs is critically limited by the quality of the graft, as HIV also infects the HSPCs. In this study, by using mobilized HSPCs from healthy donors, we show that the CD34+CD90+ hematopoietic stem cells (HSCs) express 7-fold lower CD4/CCR5 HIV receptors, higher levels of SAMHD1 anti-viral restriction factor, and possess lower susceptibility to HIV infection than the CD34+CD90- hematopoietic progenitor cells. Further, the treatment with small molecule cocktail of Resveratrol, UM729 and SR1(RUS) improved the in vivo engraftment potential of CD34+CD90+ HSCs. To demonstrate that CD34+CD90+ HSC population as an ideal graft for HIV gene therapy, we sort purified CD34+CD90+ HSCs, treated with RUS and then gene edited the CCR5 with single sgRNA. On transplantation, 100,000 CD34+CD90+ HSCs were sufficient for long-term repopulation of the entire bone marrow of NBSGW mice. Importantly, the gene editing efficiency of ~90\% in the infused product was maintained in vivo, facilitating the generation of CCR5 null immune cells, resistant to HIV infection. Altogether, CCR5 gene editing of CD34+CD90+ HSCs provide an ideal gene manipulation strategy for autologous HSCT based gene therapy for HIV infection.

}, doi = {https://doi.org/10.3389/fimmu.2022.792684}, author = {Karthik V. Karuppusamy and John Paul Demosthenes and Vigneshwaran Venkatesan and Abisha Crystal Christopher and Prathibha Babu and Manojkumar K. Azhagiri and Annlin Jacob and Veena Vadhini Ramalingam and Sumathi Rangaraj and Mohankumar Kumarasamypet Murugesan and Srujan Marepally and George Varghese and Alok Srivastava and Rajesh Kannangai and Saravanabhavan Thangavel} } @article {2542, title = {DDX24 is required for muscle fiber organization and the suppression of wound-induced Wnt activity necessary for pole re-establishment during planarian regeneration.}, journal = {Dev Biol}, volume = {488}, year = {2022}, month = {2022 May 04}, pages = {11-29}, abstract = {

Planarians have a remarkable ability to undergo whole-body regeneration. Successful regeneration outcome is determined by processes like polarity establishment at the wound site, which is followed by pole (organizer) specification. Interestingly, these determinants are almost exclusively expressed by muscles in these animals. However, the molecular toolkit that enables the functional versatility of planarian muscles remains poorly understood. Here we report that SMED_DDX24, a D-E-A-D Box RNA helicase, is necessary for planarian survival and regeneration. We found that DDX24 is enriched in muscles and its knockdown disrupts muscle fiber organization. This leads to defective pole specification, which in turn results in misregulation of many positional control genes specifically during regeneration. ddx24 RNAi also upregulates wound-induced Wnt signalling. Suppressing this ectopic Wnt activity rescues the knockdown phenotype by enabling better anterior pole regeneration. To summarize, our work highlights the role of an RNA helicase in muscle fiber organization, and modulating amputation-induced wnt levels, both of which seem critical for pole re-organization, thereby regulating whole-body regeneration.

}, issn = {1095-564X}, doi = {10.1016/j.ydbio.2022.04.011}, author = {Sarkar, Souradeep R and Dubey, Vinay Kumar and Jahagirdar, Anusha and Lakshmanan, Vairavan and Haroon, Mohamed Mohamed and Sowndarya, Sai and Sowdhamini, Ramanathan and Palakodeti, Dasaradhi} } @article {3715, title = {Efficient and error-free correction of sickle mutation in human erythroid cells using prime editor-2.}, journal = {Front Genome Ed}, volume = {4}, year = {2022}, month = {2022}, pages = {1085111}, abstract = {

Sickle cell anaemia (SCA) is one of the common autosomal recessive monogenic disorders, caused by a transverse point mutation (GAG \> GTG) at the sixth codon of the beta-globin gene, which results in haemolytic anaemia due to the fragile RBCs. Recent progress in genome editing has gained attention for the therapeutic cure for SCA. Direct correction of SCA mutation by homology-directed repair relies on a double-strand break (DSB) at the target site and carries the risk of generating beta-thalassaemic mutations if the editing is not error-free. On the other hand, base editors cannot correct the pathogenic SCA mutation resulting from A \> T base transversion. Prime editor (PE), the recently described CRISPR/Cas 9 based gene editing tool that enables precise gene manipulations without DSB and unintended nucleotide changes, is a viable approach for the treatment of SCA. However, the major limitation with the use of prime editing is the lower efficiency especially in human erythroid cell lines and primary cells. To overcome these limitations, we developed a modular lenti-viral based prime editor system and demonstrated its use for the precise modelling of SCA mutation and its subsequent correction in human erythroid cell lines. We achieved highly efficient installation of SCA mutation (up to 72\%) and its subsequent correction in human erythroid cells. For the first time, we demonstrated the functional restoration of adult haemoglobin without any unintended nucleotide changes or indel formations using the PE2 system. We also validated that the off-target effects mediated by the PE2 system is very minimal even with very efficient on-target conversion, making it a safe therapeutic option. Taken together, the modular lenti-viral prime editor system developed in this study not only expands the range of cell lines targetable by prime editor but also improves the efficiency considerably, enabling the use of prime editor for myriad molecular, genetic, and translational studies.

}, issn = {2673-3439}, doi = {10.3389/fgeed.2022.1085111}, author = {George, Anila and Ravi, Nithin Sam and Prasad, Kirti and Panigrahi, Lokesh and Koikkara, Sanya and Rajendiran, Vignesh and Devaraju, Nivedhitha and Paul, Joshua and Pai, Aswin Anand and Nakamura, Yukio and Kurita, Ryo and Balasubramanian, Poonkuzhali and Thangavel, Saravanabhavan and Marepally, Srujan and Velayudhan, Shaji R and Srivastava, Alok and Mohankumar, Kumarasamypet M} } @article {3341, title = {Erythroid lineage-specific lentiviral RNAi vectors suitable for molecular functional studies and therapeutic applications.}, journal = {Sci Rep}, volume = {12}, year = {2022}, month = {2022 08 18}, pages = {14033}, abstract = {

Numerous genes exert multifaceted roles in hematopoiesis. Therefore, we generated novel\ lineage-specific RNA interference\ (RNAi) lentiviral\ vectors, H23B-Ery-Lin-shRNA\ and H234B-Ery-Lin-shRNA, to probe the functions of these genes in erythroid cells\ without affecting other hematopoietic lineages. The lineage specificity of these vectors was confirmed by\ transducing multiple hematopoietic cells to express a fluorescent protein. Unlike the previously reported erythroid lineage RNAi vector, our vectors were designed for cloning the short hairpin RNAs (shRNAs) for\ any gene, and they also provide superior knockdown of the target gene expression with a\ single shRNA integration per cell. High-level lineage-specific downregulation of BCL11A and ZBTB7A,\ two\ well-characterized transcriptional repressors of HBG in adult erythroid cells, was achieved with substantial induction of fetal hemoglobin with a single-copy lentiviral vector integration. Transduction of primary healthy donor CD34 cells with these vectors resulted in\ \>80\% reduction in the target\ protein levels and up to 40\% elevation in the γ-chain levels in the differentiated erythroid cells. Xenotransplantation of the human CD34 cells transduced with H23B-Ery-Lin-shBCL11A\ LV in immunocompromised mice showed ~ 60\% reduction in BCL11A protein expression with ~\ 40\% elevation of γ-chain levels in the erythroid cells derived from the transduced CD34 cells. Overall, the novel erythroid lineage-specific lentiviral RNAi vectors described in this study provide a\ high-level knockdown of target gene expression in the erythroid cells, making them suitable for their use in gene therapy for hemoglobinopathies. Additionally, the design of these vectors also makes them ideal for high-throughput RNAi screening for studying normal and pathological erythropoiesis.

}, keywords = {Animals, Cell Line, Tumor, Cell Lineage, DNA-Binding Proteins, Genetic Vectors, Humans, Lentivirus, Mice, RNA Interference, RNA, Small Interfering, Transcription Factors, Transduction, Genetic}, issn = {2045-2322}, doi = {10.1038/s41598-022-13783-0}, author = {Bagchi, Abhirup and Devaraju, Nivedhitha and Chambayil, Karthik and Rajendiran, Vignesh and Venkatesan, Vigneshwaran and Sayed, Nilofer and Pai, Aswin Anand and Nath, Aneesha and David, Ernest and Nakamura, Yukio and Balasubramanian, Poonkuzhali and Srivastava, Alok and Thangavel, Saravanabhavan and Mohankumar, Kumarasamypet M and Velayudhan, Shaji R} } @article {2465, title = {Identification of novel HPFH-like mutations by CRISPR base editing that elevate the expression of fetal hemoglobin.}, journal = {Elife}, volume = {11}, year = {2022}, month = {2022 02 11}, abstract = {

Naturally occurring point mutations in the promoter switch hemoglobin synthesis from defective adult beta-globin to fetal gamma-globin in sickle cell patients with hereditary persistence of fetal hemoglobin (HPFH) and ameliorate the clinical severity. Inspired by this natural phenomenon, we tiled the highly homologous proximal promoters using adenine and cytosine base editors that avoid the generation of large deletions and identified novel regulatory regions including a cluster at the -123 region. Base editing at -123 and -124 bp of promoter induced fetal hemoglobin (HbF) to a higher level than disruption of well-known BCL11A binding site in erythroblasts derived from human CD34+ hematopoietic stem and progenitor cells (HSPC). We further demonstrated in vitro that the introduction of -123T \> C and -124T \> C HPFH-like mutations drives gamma-globin expression by creating a de novo binding site for KLF1. Overall, our findings shed light on so far unknown regulatory elements within the promoter and identified additional targets for therapeutic upregulation of fetal hemoglobin.

}, keywords = {Adenine, Anemia, Sickle Cell, beta-Globins, beta-Thalassemia, Cell Line, Clustered Regularly Interspaced Short Palindromic Repeats, CRISPR-Cas Systems, Cytosine, Fetal Hemoglobin, gamma-Globins, Gene Editing, Hematopoietic Stem Cells, Humans, Point Mutation, Promoter Regions, Genetic}, issn = {2050-084X}, doi = {10.7554/eLife.65421}, author = {Ravi, Nithin Sam and Wienert, Beeke and Wyman, Stacia K and Bell, Henry William and George, Anila and Mahalingam, Gokulnath and Vu, Jonathan T and Prasad, Kirti and Bandlamudi, Bhanu Prasad and Devaraju, Nivedhitha and Rajendiran, Vignesh and Syedbasha, Nazar and Pai, Aswin Anand and Nakamura, Yukio and Kurita, Ryo and Narayanasamy, Muthuraman and Balasubramanian, Poonkuzhali and Thangavel, Saravanabhavan and Marepally, Srujan and Velayudhan, Shaji R and Srivastava, Alok and DeWitt, Mark A and Crossley, Merlin and Corn, Jacob E and Mohankumar, Kumarasamypet M} } @article {2400, title = {Inflammation-specific targeted carriers for local drug delivery to inflammatory bowel disease.}, journal = {Biomaterials}, volume = {281}, year = {2022}, month = {2022 Jan 05}, pages = {121364}, abstract = {

Delivering drugs directly to the inflamed intestinal sites to treat inflammatory bowel disease (IBD), particularly Crohn{\textquoteright}s and ulcerative colitis, is highly challenging. Recent advances in colitis therapy medications are expanding opportunities for improving local on-site drug availability by minimising the associated systemic side-effects. Drug delivery with targeted carrier systems has shown the potential to increase site-specificity, stability, and therapeutic efficacy. Herein, we report the development of a strong anionic charged inflammation targeted nanocarriers (IT-NCs) loaded with an immunosuppressant model drug. This system showed preferential adhesion on a charge-modified surface in vitro, and in both dextran sulfate sodium (DSS) and TNBS colitis mice in vivo models. IT-NCs showed improved colitis phenotype therapeutic efficacy in both animal models compared to free drug. Furthermore, ex vivo study of colon tissue biopsies from patients with colitis revealed that IT-NCs adhered preferentially to inflamed biopsies compared to normal. Together, our results suggest that IT-NCs have promising therapeutic potential as delivery carriers{\textquoteright} in colitis management.

}, issn = {1878-5905}, doi = {10.1016/j.biomaterials.2022.121364}, author = {Kotla, Niranjan G and Singh, Rajbir and Baby, Becca V and Rasala, Swetha and Rasool, Jawad and Hynes, Sean O and Martin, Darrell and Egan, Laurence J and Vemula, Praveen K and Jala, Venkatakrishna R and Rochev, Yury and Pandit, Abhay} } @article {3642, title = {Intermittent scavenging of storage lesion from stored red blood cells by electrospun nanofibrous sheets enhances their quality and shelf-life.}, journal = {Nat Commun}, volume = {13}, year = {2022}, month = {2022 Dec 01}, pages = {7394}, abstract = {

Transfusion of healthy red blood cells (RBCs) is a lifesaving process. However, upon storing RBCs, a wide range of damage-associate molecular patterns (DAMPs), such as cell-free DNA, nucleosomes, free-hemoglobin, and poly-unsaturated-fatty-acids are generated. DAMPs can further damage RBCs; thus, the quality of stored RBCs declines during the storage and limits their shelf-life. Since these DAMPs consist of either positive or negative charged species, we developed taurine and acridine containing electrospun-nanofibrous-sheets (Tau-AcrNFS), featuring anionic, cationic charges and an DNA intercalating group on their surfaces. We show that Tau-AcrNFS are efficient in scavenging DAMPs from stored human and mice RBCs ex vivo. We find that intermittent scavenging of DAMPs by Tau-AcrNFS during the storage reduces the loss of RBC membrane integrity and reduces discocytes-to-spheroechinocytes transformation in stored-old-RBCs. We perform RBC-transfusion studies in mice to reveal that intermittent removal of DAMPs enhances the quality of stored-old-RBCs equivalent to freshly collected RBCs, and increases their shelf-life by ~22\%. Such prophylactic technology may lead to the development of novel blood bags or medical device, and may therefore impact healthcare by reducing transfusion-related adverse effects.

}, keywords = {Acridines, Animals, Drug-Related Side Effects and Adverse Reactions, Erythrocytes, Humans, Mice, Nanofibers, Research Personnel}, issn = {2041-1723}, doi = {10.1038/s41467-022-35269-3}, author = {Pandey, Subhashini and Mahato, Manohar and Srinath, Preethem and Bhutani, Utkarsh and Goap, Tanu Jain and Ravipati, Priusha and Vemula, Praveen Kumar} } @article {2423, title = {Lysate-based pipeline to characterize microtubule-associated proteins uncovers unique microtubule behaviours.}, journal = {Nat Cell Biol}, volume = {24}, year = {2022}, month = {2022 Feb}, pages = {253-267}, abstract = {

The microtubule cytoskeleton forms complex macromolecular assemblies with a range of microtubule-associated proteins (MAPs) that have fundamental roles in cell architecture, division and motility. Determining how an individual MAP modulates microtubule behaviour is an important step in understanding the physiological roles of various microtubule assemblies. To characterize how MAPs control microtubule properties and functions, we developed an approach allowing for medium-throughput analyses of MAPs in cell-free conditions using lysates of mammalian cells. Our pipeline allows for quantitative as well as ultrastructural analyses of microtubule-MAP assemblies. Analysing 45 bona fide and potential mammalian MAPs, we uncovered previously unknown activities that lead to distinct and unique microtubule behaviours such as microtubule coiling or hook formation, or liquid-liquid phase separation along the microtubule lattice that initiates microtubule branching. We have thus established a powerful tool for a thorough characterization of a wide range of MAPs and MAP variants, thus opening avenues for the determination of mechanisms underlying their physiological roles and pathological implications.

}, issn = {1476-4679}, doi = {10.1038/s41556-021-00825-4}, author = {Jijumon, A S and Bodakuntla, Satish and Genova, Mariya and Bangera, Mamata and Sackett, Violet and Besse, Laetitia and Maksut, Fatlinda and Henriot, Veronique and Magiera, Maria M and Sirajuddin, Minhajuddin and Janke, Carsten} } @article {3342, title = {Methionine uptake via the SLC43A2 transporter is essential for regulatory T-cell survival.}, journal = {Life Sci Alliance}, volume = {5}, year = {2022}, month = {2022 Sep 09}, abstract = {

Cell death, survival, or growth decisions in T-cell subsets depend on interplay between cytokine-dependent and metabolic processes. The metabolic requirements of T-regulatory cells (Tregs) for their survival and how these are satisfied remain unclear. Herein, we identified a necessary requirement of methionine uptake and usage for Tregs survival upon IL-2 deprivation. Activated Tregs have high methionine uptake and usage to S-adenosyl methionine, and this uptake is essential for Tregs survival in conditions of IL-2 deprivation. We identify a solute carrier protein SLC43A2 transporter, regulated in a Notch1-dependent manner that is necessary for this methionine uptake and Tregs viability. Collectively, we uncover a specifically regulated mechanism of methionine import in Tregs that is required for cells to adapt to cytokine withdrawal. We highlight the need for methionine availability and metabolism in contextually regulating cell death in this immunosuppressive population of T cells.

}, keywords = {Interleukin-2, Methionine, Racemethionine, Solute Carrier Proteins, T-Lymphocytes, Regulatory}, issn = {2575-1077}, doi = {10.26508/lsa.202201663}, author = {Saini, Neetu and Naaz, Afsana and Metur, Shree Padma and Gahlot, Pinki and Walvekar, Adhish and Dutta, Anupam and Davathamizhan, Umamaheswari and Sarin, Apurva and Laxman, Sunil} } @article {2883, title = {Microbial metabolite restricts 5-fluorouracil-resistant colonic tumor progression by sensitizing drug transporters via regulation of FOXO3-FOXM1 axis.}, journal = {Theranostics}, volume = {12}, year = {2022}, month = {2022}, pages = {5574-5595}, abstract = {

The survival rate of colorectal cancer patients is adversely affected by the selection of tumors resistant to conventional anti-cancer drugs such as 5-fluorouracil (5FU). Although there is mounting evidence that commensal gut microbiota is essential for effective colon cancer treatment, the detailed molecular mechanisms and the role of gut microbial metabolites remain elusive. The goal of this study is to decipher the impact and mechanisms of gut microbial metabolite, urolithin A (UroA) and its structural analogue, UAS03 on reversal of 5FU-resistant (5FUR) colon cancers. We have utilized the SW480 and HCT-116 parental (5FU-sensitive) and 5FUR colon cancer cells to examine the chemosensitization effects of UroA or UAS03 by using both and models. The effects of mono (UroA/UAS03/5FU) and combinatorial therapy (UroA/UAS03 + 5FU) on cell proliferation, apoptosis, cell migration and invasion, regulation of epithelial mesenchymal transition (EMT) mediators, expression and activities of drug transporters, and their regulatory transcription factors were examined using molecular, cellular, immunological and flowcytometric methods. Further, the anti-tumor effects of mono/combination therapy (UroA or UAS03 or 5FU or UroA/UAS03 + 5FU) were examined using pre-clinical models of 5FUR-tumor xenografts in NRGS mice and azoxymethane (AOM)-dextran sodium sulfate (DSS)-induced colon tumors. Our data showed that UroA or UAS03 in combination with 5FU significantly inhibited cell viability, proliferation, invasiveness as well as induced apoptosis of the 5FUR colon cancer cells compared to mono treatments. Mechanistically, UroA or UAS03 chemosensitized the 5FUR cancer cells by downregulating the expression and activities of drug transporters (MDR1, BCRP, MRP2 and MRP7) leading to a decrease in the efflux of 5FU. Further, our data suggested the UroA or UAS03 chemosensitized 5FUR cancer cells to 5FU treatment through regulating FOXO3-FOXM1 axis. Oral treatment with UroA or UAS03 in combination with low dose i.p. 5FU significantly reduced the growth of 5FUR-tumor xenografts in NRGS mice. Further, combination therapy significantly abrogated colonic tumors in AOM-DSS-induced colon tumors in mice. In summary, gut microbial metabolite UroA and its structural analogue UAS03 chemosensitized the 5FUR colon cancers for effective 5FU chemotherapy. This study provided the novel characteristics of gut microbial metabolites to have significant translational implications in drug-resistant cancer therapeutics.

}, keywords = {Animals, Antimetabolites, Antineoplastic, ATP Binding Cassette Transporter, Subfamily G, Member 2, Azoxymethane, Cell Line, Tumor, Colonic Neoplasms, Coumarins, Drug Resistance, Neoplasm, Fluorouracil, Forkhead Box Protein M1, Forkhead Box Protein O3, Gastrointestinal Microbiome, Humans, Mice, Neoplasm Proteins}, issn = {1838-7640}, doi = {10.7150/thno.70754}, author = {Ghosh, Sweta and Singh, Rajbir and Vanwinkle, Zachary Matthew and Guo, Haixun and Vemula, Praveen Kumar and Goel, Ajay and Haribabu, Bodduluri and Jala, Venkatakrishna Rao} } @article {2399, title = {Microscale engineering of hollow microneedle tips: design, manufacturing, optimization and validation.}, journal = {Drug Deliv Transl Res}, volume = {12}, year = {2022}, month = {2022 Feb}, pages = {350-367}, abstract = {

Transdermal and intradermal drug delivery utilizing microneedles is an emerging front in painless therapeutics. Drug delivery using hollow microneedles is the most preferred method for delivering generic transdermal drugs in the clinical setup. The needle tip must be extremely short as the drug is administered to sub-millimeter depths. Also, they need to be sharp enough to pierce through the skin with minimal skin flexing. There are multiple challenges in engineering a tip profile that is short and sharp at the same time. Stainless steel (SS) hypodermic needles with the lancet tip profile are ubiquitous in subcutaneous and intramuscular injections. They have long bevel lengths that make them inappropriate as microneedles. Thus, designing a unique tip profile and developing the manufacturing technology for microneedle applications are necessary. This article presents the design and optimization of microneedle tip profiles through analytical models. Further, manufacturing strategies for reliably obtaining designed profiles are discussed. The article concludes with experimental validation of improved piercing performance of the optimized tip profile compared to other tip profiles. The article discusses about tip geometries of stainless steel needles for microneedle applications, where depth of delivery is less than 1 mm. Through series of analyses, the optimum needle tip geometry evolved from single plane bevel (SPB) to hex plane bevel (HPB) progressively improving piercing performance.

}, issn = {2190-3948}, doi = {10.1007/s13346-021-01062-w}, author = {Badnikar, Kedar and Jayadevi, Shreyas Nataraja and Pahal, Suman and Vemula, Praveen Kumar and Nayak, Mangalore Manjunatha and Subramanyam, Dinesh Narasimhaiah} } @article {3338, title = {Modulation of biliverdin dynamics and spectral properties by Sandercyanin.}, journal = {RSC Adv}, volume = {12}, year = {2022}, month = {2022 Jul 06}, pages = {20296-20304}, abstract = {

Biliverdin IX-alpha (BV), a tetrapyrrole, is found ubiquitously in most living organisms. It functions as a metabolite, pigment, and signaling compound. While BV is known to bind to diverse protein families such as heme-metabolizing enzymes and phytochromes, not many BV-bound lipocalins (ubiquitous, small lipid-binding proteins) have been studied. The molecular basis of binding and conformational selectivity of BV in lipocalins remains unexplained. Sandercyanin (SFP)-BV complex is a blue lipocalin protein present in the mucus of the Canadian walleye (). In this study, we present the structures and binding modes of BV to SFP. Using a combination of designed site-directed mutations, X-ray crystallography, UV/VIS, and resonance Raman spectroscopy, we have identified multiple conformations of BV that are stabilized in the binding pocket of SFP. In complex with the protein, these conformers generate varied spectroscopic signatures both in their absorption and fluorescence spectra. We show that despite no covalent anchor, structural heterogeneity of the chromophore is primarily driven by the D-ring pyrrole of BV. Our work shows how conformational promiscuity of BV is correlated to the rearrangement of amino acids in the protein matrix leading to modulation of spectral properties.

}, issn = {2046-2069}, doi = {10.1039/d2ra02880h}, author = {Ghosh, Swagatha and Mondal, Sayan and Yadav, Keerti and Aggarwal, Shantanu and Schaefer, Wayne F and Narayana, Chandrabhas and Subramanian, Ramaswamy} } @article {3640, title = {Nanobody derived using a peptide epitope from the spike protein receptor-binding motif inhibits entry of SARS-CoV-2 variants.}, journal = {J Biol Chem}, volume = {299}, year = {2022}, month = {2022 Nov 22}, pages = {102732}, abstract = {

The emergence of new escape mutants of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has escalated its penetration among the human population and has reinstated its status as a global pandemic. Therefore, developing effective antiviral therapy against emerging SARS-CoV variants and other viruses in a short period becomes essential. Blocking SARS-CoV-2 entry into human host cells by disrupting the spike glycoprotein-angiotensin-converting enzyme 2 interaction has already been exploited for vaccine development and monoclonal antibody therapy. Unlike the previous reports, our study used a nine-amino acid peptide from the receptor-binding motif of the spike protein as an epitope. We report the identification of an efficacious nanobody N1.2 that blocks the entry of pseudovirus-containing SARS-CoV-2 spike as the surface glycoprotein. Moreover, using mCherry fluorescence-based reporter assay, we observe a more potent neutralizing effect against both the hCoV19 (Wuhan/WIV04/2019) and the Omicron (BA.1) pseudotyped spike virus with a bivalent version of the N1.2 nanobody. In summary, our study presents a rapid and efficient methodology to use peptide sequences from a protein-receptor interaction interface as epitopes for screening nanobodies against potential pathogenic targets. We propose that this approach can also be widely extended to target other viruses and pathogens in the future.

}, issn = {1083-351X}, doi = {10.1016/j.jbc.2022.102732}, author = {Mendon, Nivya and Ganie, Rayees A and Kesarwani, Shubham and Dileep, Drisya and Sasi, Sarika and Lama, Prakash and Chandra, Anchal and Sirajuddin, Minhajuddin} } @article {2422, title = {Novel Mutations in β- Gene in Indian Patients With Dilated Cardiomyopathy.}, journal = {CJC Open}, volume = {4}, year = {2022}, month = {2022 Jan}, pages = {1-11}, abstract = {

Background: Heart failure is a hallmark of severe hypertrophic cardiomyopathy and dilated cardiomyopathy (DCM). Several mutations in the gene lead to hypertrophic cardiomyopathy. Recently, causative mutations in the gene have also been detected in DCM from different populations.

Methods: Here, we sequenced the gene in 137 Indian DCM patients and 167 ethnically matched healthy controls to detect the frequency of mutations and their association.

Results: Our study revealed 27 variations, of which 7 mutations (8.0\%) were detected exclusively in Indian DCM patients for the first time. These included 4 missense mutations-Arg723His, Phe510Leu, His358Leu, and Ser384Tyr (2.9\%); a frameshift mutation-Asn676_T-del (1.5\%); and 2 splice-site mutations (IVS17+2T) T\>G and (IVS19-1G) G\>A (3.6\%). Remarkably, all 4 missense mutations altered evolutionarily conserved amino acids. All 4 missense mutations were predicted to be pathogenic by 2 bioinformatics tools-polymorphism phenotyping v2 (PolyPhen-2) and sorting intolerant from tolerant (SIFT). In addition, the 4 homology models of β-MYH7-p.Leu358, p.Tyr384, p.Leu510, and p.His723-displayed root-mean-square deviations of \~{}2.55 {\r A}, \~{}1.24 {\r A}, \~{}3.36 {\r A}, and \~{}3.86 {\r A}, respectively.

Conclusions: In the present study, we detected numerous novel, unique, and rare mutations in the gene exclusively in Indian DCM patients (8.0\%). Here, we demonstrated how each mutant (missense) uniquely disrupts a critical network of non-bonding interactions at the mutation site (molecular level) and may contribute to development of dilated cardiomyopathy (DCM). Therefore, our findings may provide insight into the understanding of the molecular bases of disease and into diagnosis along with promoting novel therapeutic strategies (through personalized medicine).

}, issn = {2589-790X}, doi = {10.1016/j.cjco.2021.07.020}, author = {Rani, Deepa Selvi and Vijaya Kumar, Archana and Nallari, Pratibha and Sampathkumar, Katakam and Dhandapany, Perundurai S and Narasimhan, Calambur and Rathinavel, Andiappan and Thangaraj, Kumarasamy} } @article {3716, title = {Omicron infection increases IgG binding to spike protein of predecessor variants.}, journal = {J Med Virol}, year = {2022}, month = {2022 Dec 22}, abstract = {

BACKGROUND: SARS-CoV-2 transmission in India in 2020-2022 was driven predominantly by Wild (Wuhan-Hu-1and D614G), Delta, and Omicron variants. The aim of this study was to examine the effect of infections on the humoral immune response and cross-reactivity to spike proteins of Wuhan-Hu-1, Delta, C.1.2., and Omicron.

OBJECTIVES: Residual archival sera (N=81) received between January 2020 and March 2022 were included. Infection status was inferred by a positive SARS-CoV-2 RT-PCR and/or serology (anti-N and anti-S antibodies) and sequencing of contemporaneous samples (N=18) to infer lineage. We estimated the levels and cross-reactivity of infection-induced sera including Wild, Delta, Omicron as well as vaccine breakthrough infections (Delta and Omicron).

RESULTS: We found ~2-fold increase in spike-specific IgG antibody binding in post-Omicron infection compared to the pre-Omicron period, whilst the change in pre- and post-Delta infections were similar. Further investigation of Omicron-specific humoral responses revealed primary Omicron infection as an inducer of cross-reactive antibodies against predecessor variants, in spite of weaker degree of humoral response compared to Wuhan-Hu-1 and Delta infection. Intriguingly, Omicron vaccine-breakthrough infections when compared with primary infections, exhibited increased humoral responses against RBD (7.7-fold) and Trimeric S (Trimeric form of spike protein) (34.6-fold) in addition to increased binding of IgGs towards previously circulating variants (4.2 - 6.5-fold). Despite Delta breakthrough infections showing a higher level of humoral response against RBD (2.9-fold) and Trimeric S (5.7-fold) compared to primary Delta sera, a demonstrably reduced binding (36-49\%) was observed to Omicron spike protein.

CONCLUSIONS: Omicron vaccine breakthrough infection results in increased intensity of humoral response and wider breadth of IgG binding to spike proteins of antigenically-distinct, predecessor variants. This article is protected by copyright. All rights reserved.

}, issn = {1096-9071}, doi = {10.1002/jmv.28419}, author = {Mahalingam, Gokulnath and Periyasami, Yogapriya and Arjunan, Porkizhi and Subaschandrabose, Rajesh Kumar and Mathivanan, Tamil Venthan and Mathew, Roshlin Susan and Ramya Devi, Kt and Premkumar, Prasanna Samuel and Muliyil, Jayaprakash and Srivastava, Alok and Moorthy, Mahesh and Marepally, Srujan} } @article {2884, title = {Preferential Expansion of Human CD34CD133CD90 Hematopoietic Stem Cells Enhances Gene-Modified Cell Frequency for Gene Therapy.}, journal = {Hum Gene Ther}, volume = {33}, year = {2022}, month = {2022 02}, pages = {188-201}, abstract = {

CD34CD133CD90 hematopoietic stem cells (HSCs) are responsible for long-term multilineage hematopoiesis, and the high frequency of gene-modified HSCs is crucial for the success of hematopoietic stem and progenitor cell (HSPC) gene therapy. However, the culture and gene manipulation steps of HSPC graft preparation significantly reduce the frequency of HSCs, thus necessitating large doses of HSPCs and reagents for the manipulation. In this study, we identified a combination of small molecules, Resveratrol, UM729, and SR1 that preferentially expands CD34CD133CD90 HSCs over other subpopulations of adult HSPCs in culture. The preferential expansion enriches the HSCs in culture, enhances the adhesion, and results in a sixfold increase in the long-term engraftment in NSG mice. Further, the culture-enriched HSCs are more responsive to gene modification by lentiviral transduction and gene editing, increasing the frequency of gene-modified HSCs up to 10-fold . The yield of gene-modified HSCs obtained by the culture enrichment is similar to the sort-purification of HSCs and superior to Cyclosporin-H treatment. Our study addresses a critical challenge of low frequency of gene modified HSCs in HSPC graft by developing and demonstrating a facile HSPC culture condition that increases the frequency of gene-modified cells . This strategy will improve the outcome of HSPC gene therapy and also simplify the gene manipulation process.

}, keywords = {Animals, Antigens, CD34, Fetal Blood, Genetic Therapy, Hematopoietic Stem Cell Transplantation, Hematopoietic Stem Cells, Humans, Mice, Mice, Inbred NOD, Mice, SCID}, issn = {1557-7422}, doi = {10.1089/hum.2021.089}, author = {Christopher, Abisha Crystal and Venkatesan, Vigneshwaran and Karuppusamy, Karthik V and Srinivasan, Saranya and Babu, Prathibha and Azhagiri, Manoj Kumar K and Chambayil, Karthik and Bagchi, Abhirup and Rajendiran, Vignesh and Ravi, Nithin Sam and Kumar, Sanjay and Marepally, Srujan Kumar and Mohankumar, Kumarasamypet Murugesan and Srivastava, Alok and Velayudhan, Shaji R and Thangavel, Saravanabhavan} } @article {3347, title = {S. mediterranea ETS-1 regulates the function of cathepsin-positive cells and the epidermal lineage landscape via basement membrane remodeling.}, journal = {J Cell Sci}, volume = {135}, year = {2022}, month = {2022 10 15}, abstract = {

Extracellular matrix (ECM) is an important component of stem cell niche. Remodeling of ECM mediated by ECM regulators, such as matrix metalloproteinases (MMPs) plays a vital role in stem cell function. However, the mechanisms that modulate the function of ECM regulators in the stem cell niche are understudied. Here, we explored the role of the transcription factor (TF) ETS-1, which is expressed in the cathepsin-positive cell population, in regulating the expression of the ECM regulator, mt-mmpA, thereby modulating basement membrane thickness. In planarians, the basement membrane around the gut/inner parenchyma is thought to act as a niche for pluripotent stem cells. It has been shown that the early epidermal progenitors migrate outwards from this region and progressively differentiate to maintain the terminal epidermis. Our data shows that thickening of the basement membrane in the absence of ets-1 results in defective migration of stem cell progeny. Furthermore, the absence of ets-1 leads to a defective epidermal progenitor landscape, despite its lack of expression in those cell types. Together, our results demonstrate the active role of ECM remodeling in regulating tissue homeostasis and regeneration in the planarian Schmidtea mediterranea. This article has an associated First Person interview with one of the co-first authors of the paper.

}, keywords = {Animals, Basement Membrane, Cathepsins, Cell Differentiation, Epidermis, Humans, Matrix Metalloproteinases, Mediterranea, Planarians, Transcription Factors}, issn = {1477-9137}, doi = {10.1242/jcs.259900}, author = {Dubey, Vinay Kumar and Sarkar, Souradeep R and Lakshmanan, Vairavan and Dalmeida, Rimple and Gulyani, Akash and Palakodeti, Dasaradhi} } @article {3645, title = {Single step fabrication of hollow microneedles and an experimental package for controlled drug delivery.}, journal = {Int J Pharm}, volume = {632}, year = {2022}, month = {2022 Dec 24}, pages = {122546}, abstract = {

Hollow microneedle arrays (HMNs) are an excellent choice for managing chronic diseases requiring the administration of multiple drug doses over a prolonged duration. However, HMNs have gained partial success due to limitations in their manufacturing capabilities, and cumbersome processes. In the present study, polymeric HMNs were fabricated using a novel single-step drop-casting process without needing cleanroom facilities, and sophisticated instrumentation. When drop casted on the pyramidal tip stainless steel needles, the optimized polymer solution allowed the reproducible formation of desired height HMMs on a detachable acrylic base. To enable broader applications, the base with HMNs was integrated into an experimental package built to deliver a dose of\ \~{}\ 5\ {\textmu}L per 30{\textdegree} clockwise rotation of the actuator, allowing multiple metered drug dose administrations. The fabricated HMNs were optically imaged, and tested for mechanical integrity and stability. The working and functional utility of the HMNs package in delivering metered drug doses was demonstrated by delivering vitamin B12 (ex vivo) and insulin (in vivo), respectively. The optimized process can be used for the large-scale manufacturing of HMNs and the experimental package shows the potential to be further developed into a wearable device.

}, issn = {1873-3476}, doi = {10.1016/j.ijpharm.2022.122546}, author = {Ghate, Vivek and Renjith, Anu and Badnikar, Kedar and Pahal, Suman and Jayadevi, Shreyas N and Nayak, Manjunatha M and Vemula, Praveen K and Subramanyam, Dinesh N} } @article {3344, title = {Sirtuin1 meditated modification of Notch1 intracellular domain regulates nucleolar localization and activation of distinct signaling cascades.}, journal = {Front Cell Dev Biol}, volume = {10}, year = {2022}, month = {2022}, pages = {988816}, abstract = {

Notch signaling is involved in cell fate decisions in the development and maintenance of tissue homeostasis. Spatial regulation of the Notch1 intracellular domain (NIC1), has been shown to underpin signaling outcomes mediated by this receptor. We recently reported a putative Nucleolar Localization Sequence (NoLS) in NIC1. Here we investigate if the putative NoLS identified in NIC1 regulates localization in the nucleolus and anti-apoptotic activity. Confocal imaging of live cells expressing NIC1 or forms modified by deletion or site-directed mutagenesis established that the putative NoLS in NIC1 is required for nucleolar localization and regulated by the deacetylase Sirtuin1. Subsequent analysis of anti-apoptotic activity revealed signaling cascades linked to nucleolar localization. For this, etoposide and 4-Nitroquinoline 1-oxide, an inhibitor of topoisomerase-II and a UV mimetic drug respectively, were used as prototypic triggers of genomic damage in a mammalian cell line. While NIC1 blocked apoptosis regardless of its localization to the nucleoplasm or nucleolus, modifications of NIC1 which promoted localization to the nucleolus triggered a dependence on the nucleolar proteins fibrillarin and nucleolin for anti-apoptotic activity. Further, cells co-expressing NIC1 and Sirtuin1 (but not its catalytically inactive form), confirmed both spatial regulation and the switch to dependence on the nucleolar proteins. Finally, site-directed mutagenesis showed that the NoLS lysine residues are targets of Sirtuin1 activity. NIC1 mediated transcription is not similarly regulated. Thus, NIC1 localization to the nucleolus is regulated by Sirtuin1 modification of the lysine residues in NoLS and triggers a distinct signaling cascade involving nucleolar intermediates for anti-apoptotic activity.

}, issn = {2296-634X}, doi = {10.3389/fcell.2022.988816}, author = {Saini, Neetu and Bheeshmachar, Geetha and Sarin, Apurva} } @article {3345, title = {Snail maintains the stem/progenitor state of skin epithelial cells and carcinomas through the autocrine effect of matricellular protein Mindin.}, journal = {Cell Rep}, volume = {40}, year = {2022}, month = {2022 09 20}, pages = {111390}, abstract = {

Preservation of a small population of cancer stem cells (CSCs) within a heterogeneous carcinoma serves as a paradigm to understand how select cells in a tissue maintain their undifferentiated status. In both embryogenesis and cancer, Snail has been correlated with stemness, but the molecular underpinning of this phenomenon remains largely ill-defined. In models of cutaneous squamous cell carcinoma (cSCC), we discovered a non-epithelial-mesenchymal transition function for the transcription factor Snail in maintaining the stemness of epidermal keratinocytes. Snail-expressing cells secrete the matricellular protein Mindin, which functions in an autocrine fashion to activate a Src-STAT3 pathway to reinforce their stem/progenitor phenotype. This pathway is activated by the engagement of Mindin with the leukocyte-specific integrin, CD11b (ITGAM), which is also unexpectedly expressed by epidermal keratinocytes. Interestingly, disruption of this signaling module in human cSCC attenuates tumorigenesis, suggesting that targeting Mindin would be a promising therapeutic approach to hinder cancer recurrence.

}, keywords = {Carcinoma, Squamous Cell, Cell Line, Tumor, Epithelial Cells, Extracellular Matrix Proteins, Humans, Integrins, Neoplasm Proteins, Neoplasm Recurrence, Local, Neoplastic Stem Cells, Skin Neoplasms, Snail Family Transcription Factors}, issn = {2211-1247}, doi = {10.1016/j.celrep.2022.111390}, author = {Badarinath, Krithika and Dam, Binita and Kataria, Sunny and Zirmire, Ravindra K and Dey, Rakesh and Kansagara, Gaurav and Ajnabi, Johan and Hegde, Akshay and Singh, Randhir and Masudi, Tafheem and Sambath, Janani and Sachithanandan, Sasikala P and Kumar, Prashant and Gulyani, Akash and He, You-Wen and Krishna, Sudhir and Jamora, Colin} } @article {2882, title = {Supplementation of articular cartilage-derived chondroprogenitors with bone morphogenic protein-9 enhances chondrogenesis without affecting hypertrophy.}, journal = {Biotechnol Lett}, year = {2022}, month = {2022 Aug 03}, abstract = {

INTRODUCTION: Chondroprogenitors (CPCs) have emerged as a promising cellular therapy for cartilage-related pathologies due to their inherent primed chondrogenic potential. Studies report that the addition of growth factors such as parathyroid hormone (PTH) and Bone Morphogenic Protein (BMP) enhance the chondroinducive potential in chondrocytes and mesenchymal stem cells. This study evaluated if supplementation of the standard culture medium for cell expansion with 1-34 PTH and BMP-9 would enhance the chondrogenic potential of CPCs and reduce their hypertrophic tendency.

METHODS: Human chondrocytes were isolated from patients undergoing total knee replacement for osteoarthritis (n = 3). Following fibronectin adhesion assay, passage 1 CPCs were divided and further expanded under three culture conditions (a) control, i.e., cells continued under standard culture conditions, (b) 1-34 PTH group, additional intermittent 6\ h exposure with 1-34 PTH and (c) BMP-9 group, additional BMP-9 during culture expansion. All the groups were evaluated for population-doubling, cell cycle analysis, surface marker and gene expression for chondrogenesis, hypertrophy, multilineage differentiation and GAG (glycosaminoglycan)/DNA following chondrogenic differentiation.

RESULTS: Concerning growth kinetics, the BMP-9 group exhibited a significantly lower S-phase and population-doubling when compared to the other two groups. Qualitative analysis for chondrogenic potential (Alcian blue, Safranin O staining and Toluidine blue for GAG) revealed that the BMP-9 group exhibited the highest uptake. The BMP-9 group also showed significantly higher COL2A1 expression than the control group, with no change in the hypertrophy marker expression.

CONCLUSION: BMP-9 can potentially be used as an additive for CPCs expansion, to enhance their chondrogenic potential without affecting their low hypertrophic tendency. The mitigating effects of 1-34PTH on hypertrophy would benefit further investigation when used in combination with BMP-9 to enhance chondrogenesis whilst reducing hypertrophy.

}, issn = {1573-6776}, doi = {10.1007/s10529-022-03280-9}, author = {Padmaja, Kawin and Amirtham, Soosai Manickam and Rebekah, Grace and Sathishkumar, Solomon and Vinod, Elizabeth} } @article {2463, title = {Whole genome sequencing delineates regulatory, copy number, and cryptic splice variants in early onset cardiomyopathy.}, journal = {NPJ Genom Med}, volume = {7}, year = {2022}, month = {2022 Mar 14}, pages = {18}, abstract = {

Cardiomyopathy (CMP) is a heritable disorder. Over 50\% of cases are gene-elusive on clinical gene panel testing. The contribution of variants in non-coding DNA elements that result in cryptic splicing and regulate gene expression has not been explored. We analyzed whole-genome sequencing (WGS) data in a discovery cohort of 209 pediatric CMP patients and 1953 independent replication genomes and exomes. We searched for protein-coding variants, and non-coding variants predicted to affect the function or expression of genes. Thirty-nine percent of cases harbored pathogenic coding variants in known CMP genes, and 5\% harbored high-risk loss-of-function (LoF) variants in additional candidate CMP genes. Fifteen percent harbored high-risk regulatory variants in promoters and enhancers of CMP genes (odds ratio 2.25, p = 6.70 {\texttimes} 10 versus controls). Genes involved in α-dystroglycan glycosylation (FKTN, DTNA) and desmosomal signaling (DSC2, DSG2) were most highly enriched for regulatory variants (odds ratio 6.7-58.1). Functional effects were confirmed in patient myocardium and reporter assays in human cardiomyocytes, and in zebrafish CRISPR knockouts. We provide strong evidence for the genomic contribution of functionally active variants in new genes and in regulatory elements of known CMP genes to early onset CMP.

}, issn = {2056-7944}, doi = {10.1038/s41525-022-00288-y}, author = {Lesurf, Robert and Said, Abdelrahman and Akinrinade, Oyediran and Breckpot, Jeroen and Delfosse, Kathleen and Liu, Ting and Yao, Roderick and Persad, Gabrielle and McKenna, Fintan and Noche, Ramil R and Oliveros, Winona and Mattioli, Kaia and Shah, Shreya and Miron, Anastasia and Yang, Qian and Meng, Guoliang and Yue, Michelle Chan Seng and Sung, Wilson W L and Thiruvahindrapuram, Bhooma and Lougheed, Jane and Oechslin, Erwin and Mondal, Tapas and Bergin, Lynn and Smythe, John and Jayappa, Shashank and Rao, Vinay J and Shenthar, Jayaprakash and Dhandapany, Perundurai S and Semsarian, Christopher and Weintraub, Robert G and Bagnall, Richard D and Ingles, Jodie and Mel{\'e}, Marta and Maass, Philipp G and Ellis, James and Scherer, Stephen W and Mital, Seema} } @article {3643, title = {Zingerone-encapsulated Solid Lipid Nanoparticles as Oral Drug-delivery Systems to Potentially Target Inflammatory Diseases}, journal = {ChemNanoMat}, volume = {8}, year = {2022}, month = {November}, doi = {10.1002/cnma.202200388}, url = {https://doi.org/10.1002\%2Fcnma.202200388}, author = {Omprakash Sunnap and Singaravadivel Subramanian and Praveen Kumar Vemula and Sekar Karuppannan} } @article {2211, title = {Adiponectin receptor 1 variants contribute to hypertrophic cardiomyopathy that can be reversed by rapamycin.}, journal = {Sci Adv}, volume = {7}, year = {2021}, month = {2021 Jan}, abstract = {

Hypertrophic cardiomyopathy (HCM) is a heterogeneous genetic heart muscle disease characterized by hypertrophy with preserved or increased ejection fraction in the absence of secondary causes. However, recent studies have demonstrated that a substantial proportion of individuals with HCM also have comorbid diabetes mellitus (~10\%). Whether genetic variants may contribute a combined phenotype of HCM and diabetes mellitus is not known. Here, using next-generation sequencing methods, we identified novel and ultrarare variants in adiponectin receptor 1 () as risk factors for HCM. Biochemical studies showed that variants dysregulate glucose and lipid metabolism and cause cardiac hypertrophy through the p38/mammalian target of rapamycin and/or extracellular signal-regulated kinase pathways. A transgenic mouse model expressing an variant displayed cardiomyopathy that recapitulated the cellular findings, and these features were rescued by rapamycin. Our results provide the first evidence that variants can cause HCM and provide new insights into regulation.

}, issn = {2375-2548}, doi = {10.1126/sciadv.abb3991}, author = {Dhandapany, Perundurai S and Kang, Soojeong and Kashyap, Deepak K and Rajagopal, Raksha and Sundaresan, Nagalingam R and Singh, Rajvir and Thangaraj, Kumarasamy and Jayaprakash, Shilpa and Manjunath, Cholenahally N and Shenthar, Jayaprakash and Lebeche, Djamel} } @article {2328, title = {APOE4 Affects Basal and NMDAR-Mediated Protein Synthesis in Neurons by Perturbing Calcium Homeostasis.}, journal = {J Neurosci}, volume = {41}, year = {2021}, month = {2021 Oct 20}, pages = {8686-8709}, abstract = {

Apolipoprotein E (APOE), one of the primary lipoproteins in the brain has three isoforms in humans, APOE2, APOE3, and APOE4. APOE4 is the most well-established risk factor increasing the predisposition for Alzheimer{\textquoteright}s disease (AD). The presence of the APOE4 allele alone is shown to cause synaptic defects in neurons and recent studies have identified multiple pathways directly influenced by APOE4. However, the mechanisms underlying APOE4-induced synaptic dysfunction remain elusive. Here, we report that the acute exposure of primary cortical neurons or synaptoneurosomes to APOE4 leads to a significant decrease in global protein synthesis. Primary cortical neurons were derived from male and female embryos of Sprague Dawley (SD) rats or C57BL/6J mice. Synaptoneurosomes were prepared from P30 male SD rats. APOE4 treatment also abrogates the NMDA-mediated translation response indicating an alteration of synaptic signaling. Importantly, we demonstrate that both APOE3 and APOE4 generate a distinct translation response which is closely linked to their respective calcium signature. Acute exposure of neurons to APOE3 causes a short burst of calcium through NMDA receptors (NMDARs) leading to an initial decrease in protein synthesis which quickly recovers. Contrarily, APOE4 leads to a sustained increase in calcium levels by activating both NMDARs and L-type voltage-gated calcium channels (L-VGCCs), thereby causing sustained translation inhibition through eukaryotic translation elongation factor 2 (eEF2) phosphorylation, which in turn disrupts the NMDAR response. Thus, we show that APOE4 affects basal and activity-mediated protein synthesis responses in neurons by affecting calcium homeostasis. Defective protein synthesis has been shown as an early defect in familial Alzheimer{\textquoteright}s disease (AD). However, this has not been studied in the context of sporadic AD, which constitutes the majority of cases. In our study, we show that Apolipoprotein E4 (APOE4), the predominant risk factor for AD, inhibits global protein synthesis in neurons. APOE4 also affects NMDA activity-mediated protein synthesis response, thus inhibiting synaptic translation. We also show that the defective protein synthesis mediated by APOE4 is closely linked to the perturbation of calcium homeostasis caused by APOE4 in neurons. Thus, we propose the dysregulation of protein synthesis as one of the possible molecular mechanisms to explain APOE4-mediated synaptic and cognitive defects. Hence, the study not only suggests an explanation for the APOE4-mediated predisposition to AD, it also bridges the gap in understanding APOE4-mediated pathology.

}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.0435-21.2021}, author = {Ramakrishna, Sarayu and Jhaveri, Vishwaja and Konings, Sabine C and Nawalpuri, Bharti and Chakraborty, Sumita and Holst, Bj{\o}rn and Schmid, Benjamin and Gouras, Gunnar K and Freude, Kristine K and Muddashetty, Ravi S} } @article {2362, title = {Astrocytic reactivity triggered by defective autophagy and metabolic failure causes neurotoxicity in frontotemporal dementia type 3.}, journal = {Stem Cell Reports}, volume = {16}, year = {2021}, month = {2021 Nov 09}, pages = {2736-2751}, abstract = {

Frontotemporal dementia type 3 (FTD3), caused by a point mutation in the charged multivesicular body protein 2B (CHMP2B), affects mitochondrial ultrastructure and the endolysosomal pathway in neurons. To dissect the astrocyte-specific impact of mutant CHMP2B expression, we generated astrocytes from human induced pluripotent stem cells (hiPSCs) and confirmed our findings in CHMP2B mutant mice. Our data provide mechanistic insights into how defective autophagy causes perturbed mitochondrial dynamics with impaired glycolysis, increased reactive oxygen species, and elongated mitochondrial morphology, indicating increased mitochondrial fusion in FTD3 astrocytes. This shift in astrocyte homeostasis triggers a reactive astrocyte phenotype and increased release of toxic cytokines, which accumulate in nuclear factor kappa b (NF-κB) pathway activation with increased production of CHF, LCN2, and C3 causing neurodegeneration.

}, issn = {2213-6711}, doi = {10.1016/j.stemcr.2021.09.013}, author = {Chandrasekaran, Abinaya and Dittlau, Katarina Stoklund and Corsi, Giulia I and Haukedal, Henriette and Doncheva, Nadezhda T and Ramakrishna, Sarayu and Ambardar, Sheetal and Salcedo, Claudia and Schmidt, Sissel I and Zhang, Yu and Cirera, Susanna and Pihl, Maria and Schmid, Benjamin and Nielsen, Troels Tolstrup and Nielsen, J{\o}rgen E and Kolko, Miriam and Kobol{\'a}k, Julianna and Dinny{\'e}s, Andr{\'a}s and Hyttel, Poul and Palakodeti, Dasaradhi and Gorodkin, Jan and Muddashetty, Ravi S and Meyer, Morten and Aldana, Blanca I and Freude, Kristine K} } @article {2293, title = {Contribution of NMDA Receptors to Synaptic Function in Rat Hippocampal Interneurons.}, journal = {eNeuro}, volume = {8}, year = {2021}, month = {2021 Jul-Aug}, abstract = {

The ability of neurons to produce behaviorally relevant activity in the absence of pathology relies on the fine balance of synaptic inhibition to excitation. In the hippocampal CA1 microcircuit, this balance is maintained by a diverse population of inhibitory interneurons that receive largely similar glutamatergic afferents as their target pyramidal cells, with EPSCs generated by both AMPA receptors (AMPARs) and NMDA receptors (NMDARs). In this study, we take advantage of a recently generated GluN2A-null rat model to assess the contribution of GluN2A subunits to glutamatergic synaptic currents in three subclasses of interneuron found in the CA1 region of the hippocampus. For both parvalbumin-positive and somatostatin-positive interneurons, the GluN2A subunit is expressed at glutamatergic synapses and contributes to the EPSC. In contrast, in cholecystokinin (CCK)-positive interneurons, the contribution of GluN2A to the EPSC is negligible. Furthermore, synaptic potentiation at glutamatergic synapses on CCK-positive interneurons does not require the activation of GluN2A-containing NMDARs but does rely on the activation of NMDARs containing GluN2B and GluN2D subunits.

}, issn = {2373-2822}, doi = {10.1523/ENEURO.0552-20.2021}, author = {Booker, Sam A and Sumera, Anna and Kind, Peter C and Wyllie, David J A} } @article {2326, title = {Correction of amygdalar dysfunction in a rat model of fragile X syndrome.}, journal = {Cell Rep}, volume = {37}, year = {2021}, month = {2021 Oct 12}, pages = {109805}, abstract = {

Fragile X syndrome (FXS), a commonly inherited form of autism and intellectual disability, is associated with emotional symptoms that implicate dysfunction of the amygdala. However, current understanding of the pathogenesis of the disease is based primarily on studies in the hippocampus and neocortex, where FXS defects have been corrected by inhibiting group I metabotropic glutamate receptors (mGluRs). Here, we observe that activation, rather than inhibition, of mGluRs in the basolateral amygdala reverses impairments in a rat model of FXS. FXS rats exhibit deficient recall of auditory conditioned fear, which is accompanied by a range of in\ vitro and in\ vivo deficits in synaptic transmission and plasticity. We find presynaptic mGluR5 in the amygdala, activation of which reverses deficient synaptic transmission and plasticity, thereby restoring normal fear learning in FXS rats. This highlights the importance of modifying the prevailing mGluR-based framework for therapeutic strategies to include circuit-specific differences in FXS pathophysiology.

}, issn = {2211-1247}, doi = {10.1016/j.celrep.2021.109805}, author = {Fernandes, Giselle and Mishra, Pradeep K and Nawaz, Mohammad Sarfaraz and Donlin-Asp, Paul G and Rahman, Mohammed Mostafizur and Hazra, Anupam and Kedia, Sonal and Kayenaat, Aiman and Songara, Dheeraj and Wyllie, David J A and Schuman, Erin M and Kind, Peter C and Chattarji, Sumantra} } @article {2292, title = {Cross-diagnostic evaluation of minor physical anomalies in psychiatric disorders.}, journal = {J Psychiatr Res}, volume = {142}, year = {2021}, month = {2021 Jul 20}, pages = {54-62}, abstract = {

BACKGROUND: Minor physical anomalies (MPA) are markers of impaired neurodevelopment during the prenatal stage. Assessing MPA across psychiatric disorders may help understand their shared nature. In addition, MPA in family members would indicate a shared liability and endophenotype potential. We examined familial aggregation of MPA and their role as transdiagnostic and disorder-specific markers of 5 major psychiatric/neuropsychiatric conditions (schizophrenia, bipolar disorder, substance dependence, obsessive-compulsive disorder, and Alzheimer{\textquoteright}s dementia).

METHODS: Modified Waldrop{\textquoteright}s MPA scale was applied on 1321 individuals from 439 transdiagnostic multiplex families and 125 healthy population controls (HC). Stage of fetal development (morphogenetic/phenogenetic)- and anatomical location (craniofacial/peripheral)-based sub-scores were calculated. Familiality and endophenotypic potential of MPA were analyzed with serial negative binomial mixed-effect regression. Cross-diagnostic differences and the effect of family history density (FHD) of each diagnosis on MPA were assessed. Mixed-effects Cox models estimated the influence of MPA on age-at-onset of illness (AAO).

RESULTS: MPA were found to be heritable in families with psychiatric disorders, with a familiality of 0.52. MPA were higher in psychotic disorders after controlling for effects of sex and intrafamilial correlation. Morphogenetic variant MPA was noted to be lower in dementia in comparison to HC. FHD of schizophrenia and bipolar disorder predicted higher, and that of dementia and substance dependence predicted lower MPA. MPA brought forward the AAO [HR:1.07 (1.03-1.11)], and this was more apparent in psychotic disorders.

CONCLUSION: MPA are transmissible in families, are specifically related to the risk of developing psychoses, and predict an earlier age at onset. Neurodevelopmentally informed classification of MPA has the potential to enhance the etiopathogenic and translational understanding of psychiatric disorders.

}, issn = {1879-1379}, doi = {10.1016/j.jpsychires.2021.07.028}, author = {Sreeraj, Vanteemar S and Puzhakkal, Joan C and Holla, Bharath and Nadella, Ravi Kumar and Sheth, Sweta and Balachander, Srinivas and Ithal, Dhruva and Ali, Furkhan and Viswanath, Biju and Muralidharan, Kesavan and Venkatasubramanian, Ganesan and John, John P and Benegal, Vivek and Murthy, Pratima and Varghese, Mathew and Reddy, Yc Janardhan and Jain, Sanjeev} } @article {2273, title = {Discovery of a body-wide photosensory array that matures in an adult-like animal and mediates eye-brain-independent movement and arousal.}, journal = {Proc Natl Acad Sci U S A}, volume = {118}, year = {2021}, month = {2021 May 18}, abstract = {

The ability to respond to light has profoundly shaped life. Animals with eyes overwhelmingly rely on their visual circuits for mediating light-induced coordinated movements. Building on previously reported behaviors, we report the discovery of an organized, eye-independent (extraocular), body-wide photosensory framework that allows even a head-removed animal to move like an intact animal. Despite possessing sensitive cerebral eyes and a centralized brain that controls most behaviors, head-removed planarians show acute, coordinated ultraviolet-A (UV-A) aversive phototaxis. We find this eye-brain-independent phototaxis is mediated by two noncanonical rhabdomeric opsins, the first known function for this newly classified opsin-clade. We uncover a unique array of dual-opsin-expressing photoreceptor cells that line the periphery of animal body, are proximal to a body-wide nerve net, and mediate UV-A phototaxis by engaging multiple modes of locomotion. Unlike embryonically developing cerebral eyes that are functional when animals hatch, the body-wide photosensory array matures postembryonically in "adult-like animals." Notably, apart from head-removed phototaxis, the body-wide, extraocular sensory organization also impacts physiology of intact animals. Low-dose UV-A, but not visible light (ocular-stimulus), is able to arouse intact worms that have naturally cycled to an inactive/rest-like state. This wavelength selective, low-light arousal of resting animals is noncanonical-opsin dependent but eye independent. Our discovery of an autonomous, multifunctional, late-maturing, organized body-wide photosensory system establishes a paradigm in sensory biology and evolution of light sensing.

}, issn = {1091-6490}, doi = {10.1073/pnas.2021426118}, author = {Shettigar, Nishan and Chakravarthy, Anirudh and Umashankar, Suchitta and Lakshmanan, Vairavan and Palakodeti, Dasaradhi and Gulyani, Akash} } @article {2329, title = {Distinct temporal expression of the GW182 paralog TNRC6A in neurons regulates dendritic arborization.}, journal = {J Cell Sci}, volume = {134}, year = {2021}, month = {2021 Aug 15}, abstract = {

Precise development of the dendritic architecture is a critical determinant of mature neuronal circuitry. MicroRNA (miRNA)-mediated regulation of protein synthesis plays a crucial role in dendritic morphogenesis, but the role of miRNA-induced silencing complex (miRISC) protein components in this process is less studied. Here, we show an important role of a key miRISC protein, the GW182 paralog TNRC6A, in the regulation of dendritic growth. We identified a distinct brain region-specific spatiotemporal expression pattern of GW182 during rat postnatal development. We found that the window of peak GW182 expression coincides with the period of extensive dendritic growth, both in the hippocampus and cerebellum. Perturbation of GW182 function during a specific temporal window resulted in reduced dendritic growth of cultured hippocampal neurons. Mechanistically, we show that GW182 modulates dendritic growth by regulating global somatodendritic translation and actin cytoskeletal dynamics of developing neurons. Furthermore, we found that GW182 affects dendritic architecture by regulating the expression of actin modulator LIMK1. Taken together, our data reveal a previously undescribed neurodevelopmental expression pattern of GW182 and its role in dendritic morphogenesis, which involves both translational control and actin cytoskeletal rearrangement. This article has an associated First Person interview with the first author of the paper.

}, issn = {1477-9137}, doi = {10.1242/jcs.258465}, author = {Nawalpuri, Bharti and Sharma, Arpita and Chattarji, Sumantra and Muddashetty, Ravi S} } @article {2327, title = {Engineered RNA biosensors enable ultrasensitive SARS-CoV-2 detection in a simple color and luminescence assay.}, journal = {Life Sci Alliance}, volume = {4}, year = {2021}, month = {2021 12}, abstract = {

The continued resurgence of the COVID-19 pandemic with multiple variants underlines the need for diagnostics that are adaptable to the virus. We have developed toehold RNA-based sensors across the SARS-CoV-2 genome for direct and ultrasensitive detection of the virus and its prominent variants. Here, isothermal amplification of a fragment of SARS-CoV-2 RNA coupled with activation of our biosensors leads to a conformational switch in the sensor. This leads to translation of a reporter protein, for example, LacZ or nano-lantern that is easily detected using color/luminescence. By optimizing RNA amplification and biosensor design, we have generated a highly sensitive diagnostic assay that is capable of detecting as low as 100 copies of viral RNA with development of bright color. This is easily visualized by the human eye and quantifiable using spectrophotometry. Finally, this PHAsed NASBA-Translation Optical Method (PHANTOM) using our engineered RNA biosensors efficiently detects viral RNA in patient samples. This work presents a powerful and universally accessible strategy for detecting COVID-19 and variants. This strategy is adaptable to further viral evolution and brings RNA bioengineering center-stage.

}, keywords = {Biosensing Techniques, COVID-19, Humans, Luminescence, Nucleic Acid Amplification Techniques, RNA, RNA, Viral, SARS-CoV-2}, issn = {2575-1077}, doi = {10.26508/lsa.202101213}, author = {Chakravarthy, Anirudh and Nandakumar, Anirudh and George, Geen and Ranganathan, Shyamsundar and Umashankar, Suchitta and Shettigar, Nishan and Palakodeti, Dasaradhi and Gulyani, Akash and Ramesh, Arati} } @article {2323, title = {Genomic characterization and epidemiology of an emerging SARS-CoV-2 variant in Delhi, India.}, journal = {Science}, year = {2021}, month = {2021 Oct 14}, pages = {eabj9932}, abstract = {

Delhi, the national capital of India, has experienced multiple SARS-CoV-2 outbreaks in 2020 and reached population seropositivity of over 50\% by 2021. During April 2021, the city became overwhelmed by COVID-19 cases and fatalities, as a new variant B.1.617.2 (Delta) replaced B.1.1.7 (Alpha). A Bayesian model explains the growth advantage of Delta through a combination of increased transmissibility and reduced sensitivity to immune responses generated against earlier variants (median estimates; {\texttimes}1.5-fold, 20\% reduction). Seropositivity of an employee and family cohort increased from 42\% to 87.5\% between March and July 2021, with 27\% reinfections, as judged by increased antibody concentration after a previous decline. The likely high transmissibility and partial evasion of immunity by the Delta variant contributed to an overwhelming surge in Delhi.

}, issn = {1095-9203}, doi = {10.1126/science.abj9932}, author = {Dhar, Mahesh S and Marwal, Robin and Vs, Radhakrishnan and Ponnusamy, Kalaiarasan and Jolly, Bani and Bhoyar, Rahul C and Sardana, Viren and Naushin, Salwa and Rophina, Mercy and Mellan, Thomas A and Mishra, Swapnil and Whittaker, Charles and Fatihi, Saman and Datta, Meena and Singh, Priyanka and Sharma, Uma and Ujjainiya, Rajat and Bhatheja, Nitin and Divakar, Mohit Kumar and Singh, Manoj K and Imran, Mohamed and Senthivel, Vigneshwar and Maurya, Ranjeet and Jha, Neha and Mehta, Priyanka and A, Vivekanand and Sharma, Pooja and Vr, Arvinden and Chaudhary, Urmila and Soni, Namita and Thukral, Lipi and Flaxman, Seth and Bhatt, Samir and Pandey, Rajesh and Dash, Debasis and Faruq, Mohammed and Lall, Hemlata and Gogia, Hema and Madan, Preeti and Kulkarni, Sanket and Chauhan, Himanshu and Sengupta, Shantanu and Kabra, Sandhya and Gupta, Ravindra K and Singh, Sujeet K and Agrawal, Anurag and Rakshit, Partha and Nandicoori, Vinay and Tallapaka, Karthik Bharadwaj and Sowpati, Divya Tej and Thangaraj, K and Bashyam, Murali Dharan and Dalal, Ashwin and Sivasubbu, Sridhar and Scaria, Vinod and Parida, Ajay and Raghav, Sunil K and Prasad, Punit and Sarin, Apurva and Mayor, Satyajit and Ramakrishnan, Uma and Palakodeti, Dasaradhi and Seshasayee, Aswin Sai Narain and Bhat, Manoj and Shouche, Yogesh and Pillai, Ajay and Dikid, Tanzin and Das, Saumitra and Maitra, Arindam and Chinnaswamy, Sreedhar and Biswas, Nidhan Kumar and Desai, Anita Sudhir and Pattabiraman, Chitra and Manjunatha, M V and Mani, Reeta S and Arunachal Udupi, Gautam and Abraham, Priya and Atul, Potdar Varsha and Cherian, Sarah S} } @article {2311, title = {Glycomic and glycotranscriptomic profiling of mucin-type O-glycans in planarian Schmidtea mediterranea.}, journal = {Glycobiology}, year = {2021}, month = {2021 Sep 06}, abstract = {

O-Glycans on cell surfaces play important roles in cell-cell, cell-matrix, and receptor-ligand interaction. Therefore, glycan-based interactions are important for tissue regeneration and homeostasis. Free-living flatworm Schmidtea mediterranea, because of its robust regenerative potential, is of great interest in the field of stem cell biology and tissue regeneration. Nevertheless, information on the composition and structure of O-glycans in planaria is unknown. Using mass spectrometry and in silico approaches, we characterized the glycome and the related transcriptome of mucin-type O-glycans of planarian S. mediterranea. Mucin-type O-glycans were composed of multiple isomeric, methylated, and unusually extended mono- and di-substituted O-GalNAc structures. Extensions made of hexoses and 3-O methyl hexoses were the glycoforms observed. From glycotranscriptomic analysis, sixty genes belonging to five distinct enzyme classes were identified to be involved in mucin-type O-glycan biosynthesis. These genes shared homology with those in other invertebrate systems. While a majority of the genes involved in mucin-type O-glycan biosynthesis was highly expressed during organogenesis and in differentiated cells, a few select genes in each enzyme class were specifically enriched during early embryogenesis. Our results indicate a unique temporal and spatial role for mucin-type O-glycans during embryogenesis and organogenesis and in adulthood. In summary, this is the first report on O-glycans in planaria. This study expands the structural and biosynthetic possibilities in cellular glycosylation in the invertebrate glycome and provides a framework towards understanding the biological role of mucin-type O-glycans in tissue regeneration using planarians.

}, issn = {1460-2423}, doi = {10.1093/glycob/cwab097}, author = {Subramanian, Sabarinath Peruvemba and Lakshmanan, Vairavan and Palakodeti, Dasaradhi and Subramanian, Ramaswamy} } @article {2294, title = {Hair Follicle Grafting Therapy Promotes Re-Emergence of Critical Skin Components in Chronic Nonhealing Wounds}, journal = {JID Innovations}, volume = {1}, year = {2021}, abstract = {

An exploding public health crisis is the exponential growth in the incidence of chronic nonhealing ulcers associated with diseases such as diabetes. Various modalities have been developed to stimulate wound closure that is otherwise recalcitrant to standard clinical treatments. However, these approaches primarily focus on the process of re-epithelialization and are often deficient in regenerating the full spectrum of structures necessary for normal skin function. Autologous hair follicle grafting is a recent therapy to stimulate the closure of such nonhealing wounds, and we observed effects beyond the epidermis to other important components of the dermis. We found that hair follicle grafting facilitated the reappearance of various undifferentiated and differentiated layers of the epidermis with the restoration of epidermal junctions. In addition, other important structures that are critical for cutaneous health and function such as the blood and lymph vasculature, nerve fibers, and sweat gland structures were restored in postgrafted wounds. Interestingly, both immune cells and inflammatory signals were substantially decreased, indicating a reduction in the chronic inflammation that is a hallmark of nonhealing wounds. Our observation that punch wounds created on the postgrafted area likewise healed suggests that this is a self-sustaining long-term therapy for patients with chronic wounds.

}, author = {Saha, D}, editor = {Thannimangalath, S} } @article {2214, title = {iRGD conjugated nimbolide liposomes protect against endotoxin induced acute respiratory distress syndrome.}, journal = {Nanomedicine}, volume = {33}, year = {2021}, month = {2021 Jan 05}, pages = {102351}, abstract = {

Acute respiratory distress syndrome (ARDS) is a deadly respiratory illness associated with refractory hypoxemia and pulmonary edema. The recent pandemic outbreak of COVID-19 is associated with severe pneumonia and inflammatory cytokine storm in the lungs. The anti-inflammatory phytomedicine nimbolide (NIM) may not be feasible for clinical translation due to poor pharmacokinetic properties and lack of suitable delivery systems. To overcome these barriers, we have developed nimbolide liposomes conjugated with iRGD peptide (iRGD-NIMLip) for targeting lung inflammation. It was observed that iRGD-NIMLip treatment significantly inhibited oxidative stress and cytokine storm compared to nimbolide free-drug (f-NIM), nimbolide liposomes (NIMLip), and exhibited superior activity compared to dexamethasone (DEX). iRGD-NIMLip abrogated the LPS induced p65 NF-κB, Akt, MAPK, Integrin β3 and β5, STAT3, and DNMT1 expression. Collectively, our results demonstrate that iRGD-NIMLip could be a promising novel drug delivery system to target severe pathological consequences observed in ARDS and COVID-19 associated cytokine storm.

}, issn = {1549-9642}, doi = {10.1016/j.nano.2020.102351}, author = {Pooladanda, Venkatesh and Thatikonda, Sowjanya and Sunnapu, Omprakash and Tiwary, Shristy and Vemula, Praveen Kumar and Talluri, M V N Kumar and Godugu, Chandraiah} } @article {2269, title = {Kog1/Raptor mediates metabolic rewiring during nutrient limitation by controlling SNF1/AMPK activity.}, journal = {Sci Adv}, volume = {7}, year = {2021}, month = {2021 Apr}, abstract = {

In changing environments, cells modulate resource budgeting through distinct metabolic routes to control growth. Accordingly, the TORC1 and SNF1/AMPK pathways operate contrastingly in nutrient replete or limited environments to maintain homeostasis. The functions of TORC1 under glucose and amino acid limitation are relatively unknown. We identified a modified form of the yeast TORC1 component Kog1/Raptor, which exhibits delayed growth exclusively during glucose and amino acid limitations. Using this, we found a necessary function for Kog1 in these conditions where TORC1 kinase activity is undetectable. Metabolic flux and transcriptome analysis revealed that Kog1 controls SNF1-dependent carbon flux apportioning between glutamate/amino acid biosynthesis and gluconeogenesis. Kog1 regulates SNF1/AMPK activity and outputs and mediates a rapamycin-independent activation of the SNF1 targets Mig1 and Cat8. This enables effective glucose derepression, gluconeogenesis activation, and carbon allocation through different pathways. Therefore, Kog1 centrally regulates metabolic homeostasis and carbon utilization during nutrient limitation by managing SNF1 activity.

}, issn = {2375-2548}, doi = {10.1126/sciadv.abe5544}, author = {Rashida, Zeenat and Srinivasan, Rajalakshmi and Cyanam, Meghana and Laxman, Sunil} } @article {2212, title = {Microneedles for Extended Transdermal Therapeutics: A Route to Advanced Healthcare.}, journal = {Eur J Pharm Biopharm}, volume = {159}, year = {2021}, month = {2021 Feb}, pages = {151-169}, abstract = {

Sustained release of drugs over a pre-determined period is required to maintain an effective therapeutic dose for variety of drug delivery applications. Transdermal devices such as polymeric microneedle patches and other microneedle-based devices have been utilized for sustained release of their payload. Swift clearing of drugs can be prevented either by designing a slow-degrading polymeric matrix or by providing physiochemical triggers to different microneedle-based devices for on-demand release. These long-acting transdermal devices prevent the burst release of drugs. This review highlights the recent advances of microneedle-based devices for sustained release of vaccines, hormones, and antiretrovirals with their prospective safe clinical translation.

}, issn = {1873-3441}, doi = {10.1016/j.ejpb.2020.12.020}, author = {Pahal, Suman and Badnikar, Kedar and Ghate, Vivek and Bhutani, Utkarsh and Nayak, Mangalore Manjunatha and Subramanyam, Dinesh Narasimhaiah and Vemula, Praveen Kumar} } @article {2274, title = {Mitochondrial state determines functionally divergent stem cell population in planaria.}, journal = {Stem Cell Reports}, volume = {16}, year = {2021}, month = {2021 May 11}, pages = {1302-1316}, abstract = {

Mitochondrial state changes were shown to be critical for stem cell function. However, variation in the mitochondrial content in stem cells and the implication, if any, on differentiation is poorly understood. Here, using cellular and molecular studies, we show that the planarian pluripotent stem cells (PSCs) have low mitochondrial mass compared with their progenitors. Transplantation experiments provided functional validation that neoblasts with low mitochondrial mass are the true PSCs. Further, the mitochondrial mass correlated with OxPhos and inhibiting the transition to OxPhos dependent metabolism in cultured cells resulted in higher PSCs. In summary, we show that low mitochondrial mass is a hallmark of PSCs in planaria and provide a mechanism to isolate live, functionally active, PSCs from different cell cycle stages (G0/G1 and S, G2/M). Our study demonstrates that the change in mitochondrial metabolism, a feature of PSCs is conserved in planaria and highlights its role in organismal regeneration.

}, issn = {2213-6711}, doi = {10.1016/j.stemcr.2021.03.022}, author = {Mohamed Haroon, Mohamed and Lakshmanan, Vairavan and Sarkar, Souradeep R and Lei, Kai and Vemula, Praveen Kumar and Palakodeti, Dasaradhi} } @article {2244, title = {Oxylipin biosynthesis reinforces cellular senescence and allows detection of senolysis.}, journal = {Cell Metab}, year = {2021}, month = {2021 Mar 31}, abstract = {

Cellular senescence is a stress or damage response that causes a permanent proliferative arrest and secretion of numerous factors with potent biological activities. This senescence-associated secretory phenotype (SASP) has been characterized largely for secreted proteins that participate in embryogenesis, wound healing, inflammation, and many age-related pathologies. By contrast, lipid components of the SASP are understudied. We show that senescent cells activate the biosynthesis of several oxylipins that promote segments of the SASP and reinforce the proliferative arrest. Notably, senescent cells synthesize and accumulate an unstudied intracellular prostaglandin, 1a,1b-dihomo-15-deoxy-delta-12,14-prostaglandin J2. Released 15-deoxy-delta-12,14-prostaglandin J2 is a biomarker of senolysis in culture and in\ vivo. This and other prostaglandin D2-related lipids promote the senescence arrest and SASP by activating RAS signaling. These data identify an important aspect of cellular senescence and a method to detect senolysis.

}, issn = {1932-7420}, doi = {10.1016/j.cmet.2021.03.008}, author = {Wiley, Christopher D and Sharma, Rishi and Davis, Sonnet S and Lopez-Dominguez, Jose Alberto and Mitchell, Kylie P and Wiley, Samantha and Alimirah, Fatouma and Kim, Dong Eun and Payne, Therese and Rosko, Andrew and Aimontche, Eliezer and Deshpande, Sharvari M and Neri, Francesco and Kuehnemann, Chisaka and Demaria, Marco and Ramanathan, Arvind and Campisi, Judith} } @article {2401, title = {Pharmacological intervention in young adolescents rescues synaptic physiology and behavioural deficits in Syngap1 mice.}, journal = {Exp Brain Res}, year = {2021}, month = {2021 Nov 05}, abstract = {

Haploinsufficiency in SYNGAP1 is implicated in intellectual disability (ID) and autism spectrum disorder (ASD) and affects the maturation of dendritic spines. The abnormal spine development has been suggested to cause a disbalance of excitatory and inhibitory (E/I) neurotransmission at distinct developmental periods. In addition, E/I imbalances in Syngap1 mice might be due to abnormalities in K-Cl co-transporter function (NKCC1, KCC2), in a maner similar to the murine models of Fragile-X and Rett syndromes. To study whether an altered intracellular chloride ion concentration represents an underlying mechanism of modified function of GABAergic synapses in Dentate Gyrus Granule Cells of Syngap1 recordings were performed at different developmental stages of the mice. We observed depolarised neurons at P14-15 as illustrated by decreased Cl reversal potential in Syngap1 mice. The KCC2 expression was decreased compared to Wild-type (WT) mice at P14-15. The GSK-3β inhibitor, 6-bromoindirubin-3{\textquoteright}-oxime (6BIO) that crosses the blood-brain barrier, was tested to restore the function of GABAergic synapses. We discovered that the intraperitoneal administration of 6BIO during the critical period or young adolescents [P30 to P80 (4-week to 10-week)] normalised an altered E/I balance, the deficits of synaptic plasticity, and behavioural performance like social novelty, anxiety, and memory of the Syngap1 mice. In summary, altered GABAergic function in Syngap1 mice is due to reduced KCC2 expression leading to an increase in the intracellular chloride concentration that can be counteracted by the 6BIO, which restored cognitive, emotional, and social symptoms by pharmacological intervention, particularly in adulthood.

}, issn = {1432-1106}, doi = {10.1007/s00221-021-06254-x}, author = {Verma, Vijaya and Kumar, M J Vijay and Sharma, Kavita and Rajaram, Sridhar and Muddashetty, Ravi and Manjithaya, Ravi and Behnisch, Thomas and Clement, James P} } @article {2376, title = {Proteome plasticity in response to persistent environmental change.}, journal = {Mol Cell}, volume = {81}, year = {2021}, month = {2021 08 19}, pages = {3294-3309.e12}, abstract = {

Temperature is a variable component of the environment, and all organisms must deal with or adapt to temperature change. Acute temperature change activates cellular stress responses, resulting in refolding or removal of damaged proteins. However, how organisms adapt to long-term temperature change remains largely unexplored. Here we report that budding yeast responds to long-term high temperature challenge by switching from chaperone induction to reduction of temperature-sensitive proteins and re-localizing a portion of its proteome. Surprisingly, we also find that many proteins adopt an alternative conformation. Using Fet3p as an example, we find that the temperature-dependent conformational difference is accompanied by distinct thermostability, subcellular localization, and, importantly, cellular functions. We postulate that, in addition to the known mechanisms of adaptation, conformational plasticity allows some polypeptides to acquire new biophysical properties and functions when environmental change endures.

}, keywords = {Acclimatization, Adaptation, Physiological, Animals, Environmental Exposure, Gene Expression Regulation, Fungal, Hot Temperature, Proteome, Saccharomycetales, Stress, Physiological, Transcriptome}, issn = {1097-4164}, doi = {10.1016/j.molcel.2021.06.028}, author = {Domnauer, Matthew and Zheng, Fan and Li, Liying and Zhang, Yanxiao and Chang, Catherine E and Unruh, Jay R and Conkright-Fincham, Juliana and McCroskey, Scott and Florens, Laurence and Zhang, Ying and Seidel, Christopher and Fong, Benjamin and Schilling, Birgit and Sharma, Rishi and Ramanathan, Arvind and Si, Kausik and Zhou, Chuankai} } @article {2202, title = {Psychiatric symptoms and syndromes transcending diagnostic boundaries in Indian multiplex families: The cohort of ADBS study.}, journal = {Psychiatry Res}, volume = {296}, year = {2021}, month = {2021 Feb}, pages = {113647}, abstract = {

Syndromes of schizophrenia, bipolar disorder, obsessive-compulsive disorder, substance use disorders and Alzheimer{\textquoteright}s dementia are highly heritable. About 10-20\% of subjects have another affected first degree relative (FDR), and thus represent a {\textquoteright}greater{\textquoteright} genetic susceptibility. We screened 3583 families to identify 481 families with multiple affected members, assessed 1406 individuals in person, and collected information systematically about other relatives. Within the selected families, a third of all FDRs were affected with serious mental illness. Although similar diagnoses aggregated within families, 62\% of the families also had members with other syndromes. Moreover, 15\% of affected individuals met criteria for co-occurrence of two or more syndromes, across their lifetime. Using dimensional assessments, we detected a range of symptom clusters in both affected and unaffected individuals, and across diagnostic categories. Our findings suggest that in multiplex families, there is considerable heterogeneity of clinical syndromes, as well as sub-threshold symptoms. These families would help provide an opportunity for further research using both genetic analyses and biomarkers.

}, issn = {1872-7123}, doi = {10.1016/j.psychres.2020.113647}, author = {Sreeraj, Vanteemar S and Holla, Bharath and Ithal, Dhruva and Nadella, Ravi Kumar and Mahadevan, Jayant and Balachander, Srinivas and Ali, Furkhan and Sheth, Sweta and Narayanaswamy, Janardhanan C and Venkatasubramanian, Ganesan and John, John P and Varghese, Mathew and Benegal, Vivek and Jain, Sanjeev and Reddy, Yc Janardhan and Viswanath, Biju} } @article {2372, title = {Ribosomal protein S6 kinase beta-1 gene variants cause hypertrophic cardiomyopathy.}, journal = {J Med Genet}, year = {2021}, month = {2021 Dec 16}, abstract = {

BACKGROUND: Hypertrophic cardiomyopathy (HCM) is a genetic heart muscle disease with preserved or increased ejection fraction in the absence of secondary causes. Mutations in the sarcomeric protein-encoding genes predominantly cause HCM. However, relatively little is known about the genetic impact of signalling proteins on HCM.

METHODS AND RESULTS: Here, using exome and targeted sequencing methods, we analysed two independent cohorts comprising 401 Indian patients with HCM and 3521 Indian controls. We identified novel variants in ribosomal protein S6 kinase beta-1 () gene in two unrelated Indian families as a potential candidate gene for HCM. The two unrelated HCM families had the same heterozygous missense S6K1 variant (p.G47W). In a replication association study, we identified two S6K1 heterozygotes variants (p.Q49K and p.Y62H) in the UK Biobank cardiomyopathy cohort (n=190) compared with matched controls (n=16 479). These variants are neither detected in region-specific controls nor in the human population genome data. Additionally, we observed an S6K1 variant (p.P445S) in an Arab patient with HCM. Functional consequences were evaluated using representative S6K1 mutated proteins compared with wild type in cellular models. The mutated proteins activated the S6K1 and hyperphosphorylated the rpS6 and ERK1/2 signalling cascades, suggesting a gain-of-function effect.

CONCLUSIONS: Our study demonstrates for the first time that the variants in the gene are associated with HCM, and early detection of the variant carriers can help to identify family members at risk and subsequent preventive measures. Further screening in patients with HCM with different ethnic populations will establish the specificity and frequency of gene variants.

}, issn = {1468-6244}, doi = {10.1136/jmedgenet-2021-107866}, author = {Jain, Pratul Kumar and Jayappa, Shashank and Sairam, Thiagarajan and Mittal, Anupam and Paul, Sayan and Rao, Vinay J and Chittora, Harshil and Kashyap, Deepak K and Palakodeti, Dasaradhi and Thangaraj, Kumarasamy and Shenthar, Jayaprakash and Koranchery, Rakesh and Rajendran, Ranjith and Alireza, Haghighi and Mohanan, Kurukkanparampil Sreedharan and Rathinavel, Andiappan and Dhandapany, Perundurai S} } @article {2291, title = {The same stress has divergent effects on social versus asocial manifestations of anxiety-like behavior over time.}, journal = {Stress}, volume = {24}, year = {2021}, month = {2021 07}, pages = {474-480}, abstract = {

Stress may lead to augmented anxiety, which may, with time culminate in some form of anxiety disorder. Behavioral alterations related to increased anxiety can be broadly classified into two types-social, affecting interactions between individuals, and self-oriented, affecting the anxious individual only. While a growing body of literature now exists describing the effects of stress-induced anxiety on self-oriented behavior in animal models of anxiety disorders, the effects of such aberrant anxiety on social behavior has largely remained uncharacterized in these models. This study aims to fill this gap in our understanding by examining changes in social behavior following a single 2-hour episode of immobilization stress, which has been shown to cause delayed structural and functional changes in the amygdala. To this end, we examined social behavior, measured as active social interactions, anogenital sniffing, nose-to-nose contacts, allogrooming, actively following and crawling under, as well as self-oriented asocial behavior, manifested as self-grooming and rearing, in adult male rats. Stressed animals showed reduced social interaction 1 day after immobilization stress and this decrease was persistent for at least 10 days after stress. In contrast, individualistic behaviors were impaired only 10 days, but not 1 day later. Together, these results not only show that the same single episode of stress can elicit divergent effects on social and asocial measures of anxiety in the same animal, but also suggest that enhanced social anxiety soon after stress may also serve as an early indicator of its delayed behavioral effects.

}, keywords = {Amygdala, Animals, Anxiety, Anxiety Disorders, Behavior, Animal, Disease Models, Animal, Male, Rats, Social Behavior, Stress, Psychological}, issn = {1607-8888}, doi = {10.1080/10253890.2020.1855421}, author = {Saxena, Kapil and Chakraborty, Prabahan and Chattarji, Sumantra} } @article {2363, title = {SARS-CoV-2 B.1.617.2 Delta variant replication and immune evasion.}, journal = {Nature}, volume = {599}, year = {2021}, month = {2021 11}, pages = {114-119}, abstract = {

The B.1.617.2 (Delta) variant of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was first identified in the state of Maharashtra in late 2020 and spread throughout India, outcompeting pre-existing lineages including B.1.617.1 (Kappa) and B.1.1.7 (Alpha). In vitro, B.1.617.2 is sixfold less sensitive to serum neutralizing antibodies from recovered individuals, and eightfold less sensitive to vaccine-elicited antibodies, compared with wild-type Wuhan-1 bearing D614G. Serum neutralizing titres against B.1.617.2 were lower in ChAdOx1 vaccinees than in BNT162b2 vaccinees. B.1.617.2 spike pseudotyped viruses exhibited compromised sensitivity to monoclonal antibodies to the receptor-binding domain and the amino-terminal domain. B.1.617.2 demonstrated higher replication efficiency than B.1.1.7 in both airway organoid and human airway epithelial systems, associated with B.1.617.2 spike being in a predominantly cleaved state compared with B.1.1.7 spike. The B.1.617.2 spike protein was able to mediate highly efficient syncytium formation that was less sensitive to inhibition by neutralizing antibody, compared with that of wild-type spike. We also observed that B.1.617.2 had higher replication and spike-mediated entry than B.1.617.1, potentially explaining the B.1.617.2 dominance. In an analysis of more than 130 SARS-CoV-2-infected health care workers across three centres in India during a period of mixed lineage circulation, we observed reduced ChAdOx1 vaccine effectiveness against B.1.617.2 relative to non-B.1.617.2, with the caveat of possible residual confounding. Compromised vaccine efficacy against the highly fit and immune-evasive B.1.617.2 Delta variant warrants continued infection control measures in the post-vaccination era.

}, keywords = {Antibodies, Neutralizing, Cell Fusion, Cell Line, COVID-19 Vaccines, Female, Health Personnel, Humans, Immune Evasion, India, Kinetics, Male, SARS-CoV-2, Spike Glycoprotein, Coronavirus, Vaccination, Virus Replication}, issn = {1476-4687}, doi = {10.1038/s41586-021-03944-y}, author = {Mlcochova, Petra and Kemp, Steven A and Dhar, Mahesh Shanker and Papa, Guido and Meng, Bo and Ferreira, Isabella A T M and Datir, Rawlings and Collier, Dami A and Albecka, Anna and Singh, Sujeet and Pandey, Rajesh and Brown, Jonathan and Zhou, Jie and Goonawardane, Niluka and Mishra, Swapnil and Whittaker, Charles and Mellan, Thomas and Marwal, Robin and Datta, Meena and Sengupta, Shantanu and Ponnusamy, Kalaiarasan and Radhakrishnan, Venkatraman Srinivasan and Abdullahi, Adam and Charles, Oscar and Chattopadhyay, Partha and Devi, Priti and Caputo, Daniela and Peacock, Tom and Wattal, Chand and Goel, Neeraj and Satwik, Ambrish and Vaishya, Raju and Agarwal, Meenakshi and Mavousian, Antranik and Lee, Joo Hyeon and Bassi, Jessica and Silacci-Fegni, Chiara and Saliba, Christian and Pinto, Dora and Irie, Takashi and Yoshida, Isao and Hamilton, William L and Sato, Kei and Bhatt, Samir and Flaxman, Seth and James, Leo C and Corti, Davide and Piccoli, Luca and Barclay, Wendy S and Rakshit, Partha and Agrawal, Anurag and Gupta, Ravindra K} } @article {2243, title = {Spatial regulation and generation of diversity in signaling pathways.}, journal = {J Biosci}, volume = {46}, year = {2021}, month = {2021}, abstract = {

Signaling pathways orchestrate diverse cellular outcomes in the same tissue, spatially and temporally. These interactions, which are played out in micro-environments within cells and involve a relatively small number of core pathways, are the key to the development and function of multi-cellular organisms. How these outcomes are regulated has prompted interest in intracellular mechanisms that build diversity in signaling outcomes. This review specifically addresses spatial positioning of molecules as a means of enabling interactions and novel outcomes of signaling cascades. Using the Notch and Ras pathways as exemplars, we describe mechanisms that contribute to diverse signaling outcomes.

}, issn = {0973-7138}, author = {Saini, Neetu and Sarin, Apurva} } @article {2290, title = {Strategies to target SARS-CoV-2 entry and infection using dual mechanisms of inhibition by acidification inhibitors.}, journal = {PLoS Pathog}, volume = {17}, year = {2021}, month = {2021 07}, pages = {e1009706}, abstract = {

Many viruses utilize the host endo-lysosomal network for infection. Tracing the endocytic itinerary of SARS-CoV-2 can provide insights into viral trafficking and aid in designing new therapeutic strategies. Here, we demonstrate that the receptor binding domain (RBD) of SARS-CoV-2 spike protein is internalized via the pH-dependent CLIC/GEEC (CG) endocytic pathway in human gastric-adenocarcinoma (AGS) cells expressing undetectable levels of ACE2. Ectopic expression of ACE2 (AGS-ACE2) results in RBD traffic via both CG and clathrin-mediated endocytosis. Endosomal acidification inhibitors like BafilomycinA1 and NH4Cl, which inhibit the CG pathway, reduce the uptake of RBD and impede Spike-pseudoviral infection in both AGS and AGS-ACE2 cells. The inhibition by BafilomycinA1 was found to be distinct from Chloroquine which neither affects RBD uptake nor alters endosomal pH, yet attenuates Spike-pseudovirus entry. By screening a subset of FDA-approved inhibitors for functionality similar to BafilomycinA1, we identified Niclosamide as a SARS-CoV-2 entry inhibitor. Further validation using a clinical isolate of SARS-CoV-2 in AGS-ACE2 and Vero cells confirmed its antiviral effect. We propose that Niclosamide, and other drugs which neutralize endosomal pH as well as inhibit the endocytic uptake, could provide broader applicability in subverting infection of viruses entering host cells via a pH-dependent endocytic pathway.

}, keywords = {Ammonium Chloride, Angiotensin-Converting Enzyme 2, Animals, Antiviral Agents, Cell Line, Chlorocebus aethiops, Chloroquine, Clathrin, COVID-19, Drug Synergism, Endocytosis, Endosomes, Humans, Hydrogen-Ion Concentration, Hydroxychloroquine, Macrolides, Niclosamide, Protein Binding, Protein Domains, SARS-CoV-2, Spike Glycoprotein, Coronavirus, Vero Cells, Virus Internalization}, issn = {1553-7374}, doi = {10.1371/journal.ppat.1009706}, author = {Prabhakara, Chaitra and Godbole, Rashmi and Sil, Parijat and Jahnavi, Sowmya and Gulzar, Shah-E-Jahan and van Zanten, Thomas S and Sheth, Dhruv and Subhash, Neeraja and Chandra, Anchal and Shivaraj, Akshatha and Panikulam, Patricia and U, Ibrahim and Nuthakki, Vijay Kumar and Puthiyapurayil, Theja Parassini and Ahmed, Riyaz and Najar, Ashaq Hussain and Lingamallu, Sai Manoz and Das, Snigdhadev and Mahajan, Bhagyashri and Vemula, Praveen and Bharate, Sandip B and Singh, Parvinder Pal and Vishwakarma, Ram and Guha, Arjun and Sundaramurthy, Varadharajan and Mayor, Satyajit} } @article {2245, title = {Systematic evaluation of the impact of defacing on quality and volumetric assessments on T1-weighted MR-images.}, journal = {J Neuroradiol}, year = {2021}, month = {2021 Mar 13}, abstract = {

BACKGROUND AND PURPOSE: Facial features can be potentially reconstructed from structural magnetic resonance images, thereby compromising the confidentiality of study participants. Defacing methods can be applied to MRI images to ensure privacy of study participants. These methods remove facial features, thereby rendering the image unidentifiable. It is commonly assumed that defacing would not have any impact on quantitative assessments of the brain. In this study, we have assessed the impact of different defacing methods on quality and volumetric estimates.

MATERIALS AND METHODS: We performed SPM-, Freesurfer-, pydeface, and FSL-based defacing on 30 T1-weighted images. We statistically compared the change in quality measurements (from MRIQC) and volumes (from SPM, CAT, and Freesurfer) between non-defaced and defaced images. We also calculated the Dice coefficient of each tissue class between non-defaced and defaced images.

RESULTS: Almost all quality measurements and tissue volumes changed after defacing, irrespective of the method used. All tissue volumes decreased post-defacing for CAT, but no such consistent trend was seen for SPM and Freesurfer. Dice coefficients indicated that segmentations are relatively robust; however, partial volumes might be affected leading to changed volumetric estimates.

CONCLUSION: In this study, we demonstrated that volumes and quality measurements get affected differently by defacing methods. It is likely that this will have a significant impact on the reproducibility of experiments. We provide suggestions on ways to minimize the impact of defacing on outcome measurements. Our results warrant the need for robust handling of defaced images at different steps of image processing.

}, issn = {0150-9861}, doi = {10.1016/j.neurad.2021.03.001}, author = {Bhalerao, Gaurav Vivek and Parekh, Pravesh and Saini, Jitender and Venkatasubramanian, Ganesan and John, John P} } @article {2289, title = {Target identification for small-molecule discovery in the FOXO3a tumor-suppressor pathway using a biodiverse peptide library.}, journal = {Cell Chem Biol}, year = {2021}, month = {2021 Jun 01}, abstract = {

Genetic screening technologies to identify and validate macromolecular interactions (MMIs) essential for complex pathways remain an important unmet need for systems biology and therapeutics development. Here, we use a library of peptides from diverse prokaryal genomes to screen MMIs promoting the nuclear relocalization of Forkhead Box O3 (FOXO3a), a tumor suppressor more frequently inactivated by post-translational modification than mutation. A hit peptide engages the 14-3-3 family of signal regulators through a phosphorylation-dependent interaction, modulates FOXO3a-mediated transcription, and suppresses cancer cell growth. In a crystal structure, the hit peptide occupies the phosphopeptide-binding groove of 14-3-3ε in a conformation distinct from its natural peptide substrates. A biophysical screen identifies drug-like small molecules that displace the hit peptide from 14-3-3ε, providing starting points for structure-guided development. Our findings exemplify "protein interference," an approach using evolutionarily diverse, natural peptides to rapidly identify, validate, and develop chemical probes against MMIs essential for complex cellular phenotypes.

}, issn = {2451-9448}, doi = {10.1016/j.chembiol.2021.05.009}, author = {Emery, Amy and Hardwick, Bryn S and Crooks, Alex T and Milech, Nadia and Watt, Paul M and Mithra, Chandan and Kumar, Vikrant and Giridharan, Saranya and Sadasivam, Gayathri and Mathivanan, Subashini and Sudhakar, Sneha and Bairy, Sneha and Bharatham, Kavitha and Hurakadli, Manjunath A and Prasad, Thazhe K and Kamariah, Neelagandan and Muellner, Markus and Coelho, Miguel and Torrance, Christopher J and McKenzie, Grahame J and Venkitaraman, Ashok R} } @article {2216, title = {A 2-Tyr-1-carboxylate Mononuclear Iron Center Forms the Active Site of a Paracoccus Dimethylformamidase.}, journal = {Angew Chem Int Ed Engl}, volume = {59}, year = {2020}, month = {2020 09 21}, pages = {16961-16966}, abstract = {

N,N-dimethyl formamide (DMF) is an extensively used organic solvent but is also a potent pollutant. Certain bacterial species from genera such as Paracoccus, Pseudomonas, and Alcaligenes have evolved to use DMF as a sole carbon and nitrogen source for growth via degradation by a dimethylformamidase (DMFase). We show that DMFase from Paracoccus sp. strain DMF is a halophilic and thermostable enzyme comprising a multimeric complex of the α β or (α β ) type. One of the three domains of the large subunit and the small subunit are hitherto undescribed protein folds of unknown evolutionary origin. The active site consists of a mononuclear iron coordinated by two Tyr side-chain phenolates and one carboxylate from Glu. The Fe ion in the active site catalyzes the hydrolytic cleavage of the amide bond in DMF. Kinetic characterization reveals that the enzyme shows cooperativity between subunits, and mutagenesis and structural data provide clues to the catalytic mechanism.

}, issn = {1521-3773}, doi = {10.1002/anie.202005332}, author = {Arya, Chetan Kumar and Yadav, Swati and Fine, Jonathan and Casanal, Ana and Chopra, Gaurav and Ramanathan, Gurunath and Vinothkumar, Kutti R and Subramanian, Ramaswamy} } @article {2156, title = {Adverse childhood experiences in families with multiple members diagnosed to have psychiatric illnesses.}, journal = {Aust N Z J Psychiatry}, volume = {54}, year = {2020}, month = {2020 Nov}, pages = {1086-1094}, abstract = {

OBJECTIVE: Adverse childhood experiences are linked to the development of a number of psychiatric illnesses in adulthood. Our study examined the pattern of adverse childhood experiences and their relation to the age of onset of major psychiatric conditions in individuals from families that had ⩾2 first-degree relatives with major psychiatric conditions (multiplex families), identified as part of an ongoing longitudinal study.

METHODS: Our sample consisted of 509 individuals from 215 families. Of these, 268 were affected, i.e., diagnosed with bipolar disorder ( = 61), obsessive-compulsive disorder ( = 58), schizophrenia ( = 52), substance dependence ( = 59) or co-occurring diagnoses ( = 38), while 241 were at-risk first-degree relatives who were either unaffected ( = 210) or had other depressive or anxiety disorders ( = 31). All individuals were evaluated using the Adverse Childhood Experiences - International Questionnaire and total adverse childhood experiences exposure and severity scores were calculated.

RESULTS: It was seen that affected males, as a group, had the greatest adverse childhood experiences exposure and severity scores in our sample. A Cox mixed effects model fit by gender revealed that a higher total adverse childhood experiences severity score was associated with significantly increased risk for an earlier age of onset of psychiatric diagnoses in males. A similar model that evaluated the interaction of diagnosis revealed an earlier age of onset in obsessive-compulsive disorder and substance dependence, but not in schizophrenia and bipolar disorder.

CONCLUSION: Our study indicates that adverse childhood experiences were associated with an earlier onset of major psychiatric conditions in men and individuals diagnosed with obsessive-compulsive disorder and substance dependence. Ongoing longitudinal assessments in first-degree relatives from these families are expected to identify mechanisms underlying this relationship.

}, issn = {1440-1614}, doi = {10.1177/0004867420931157}, author = {Someshwar, Amala and Holla, Bharath and Pansari Agarwal, Preeti and Thomas, Anza and Jose, Anand and Joseph, Boban and Raju, Birudu and Karle, Hariprasad and Muthukumaran, M and Kodancha, Prabhath G and Kumar, Pramod and Reddy, Preethi V and Kumar Nadella, Ravi and Naik, Sanjay T and Mitra, Sayantanava and Mallappagiri, Sreenivasulu and Sreeraj, Vanteemar S and Balachander, Srinivas and Ganesh, Suhas and Murthy, Pratima and Benegal, Vivek and Reddy, Janardhan Yc and Jain, Sanjeev and Mahadevan, Jayant and Viswanath, Biju} } @article {2062, title = {The Aging Metabolome-Biomarkers to Hub Metabolites.}, journal = {Proteomics}, volume = {20}, year = {2020}, month = {2020 03}, pages = {e1800407}, abstract = {

Aging biology is intimately associated with dysregulated metabolism, which is one of the hallmarks of aging. Aging-related pathways such as mTOR and AMPK, which are major targets of anti-aging interventions including rapamcyin, metformin, and exercise, either directly regulate or intersect with metabolic pathways. In this review, numerous candidate bio-markers of aging that have emerged using metabolomics are outlined. Metabolomics studies also reveal that not all metabolites are created equally. A set of core "hub" metabolites are emerging as central mediators of aging. The hub metabolites reviewed here are nicotinamide adenine dinucleotide, reduced nicotinamide dinucleotide phosphate, α-ketoglutarate, and β-hydroxybutyrate. These "hub" metabolites have signaling and epigenetic roles along with their canonical roles as co-factors or intermediates of carbon metabolism. Together these hub metabolites suggest a central role of the TCA cycle in signaling and metabolic dysregulation associated with aging.

}, issn = {1615-9861}, doi = {10.1002/pmic.201800407}, author = {Sharma, Rishi and Ramanathan, Arvind} } @article {2248, title = {Allosteric inhibition of MTHFR prevents futile SAM cycling and maintains nucleotide pools in one-carbon metabolism.}, journal = {J Biol Chem}, volume = {295}, year = {2020}, month = {2020 11 20}, pages = {16037-16057}, abstract = {

Methylenetetrahydrofolate reductase (MTHFR) links the folate cycle to the methionine cycle in one-carbon metabolism. The enzyme is known to be allosterically inhibited by SAM for decades, but the importance of this regulatory control to one-carbon metabolism has never been adequately understood. To shed light on this issue, we exchanged selected amino acid residues in a highly conserved stretch within the regulatory region of yeast MTHFR to create a series of feedback-insensitive, deregulated mutants. These were exploited to investigate the impact of defective allosteric regulation on one-carbon metabolism. We observed a strong growth defect in the presence of methionine. Biochemical and metabolite analysis revealed that both the folate and methionine cycles were affected in these mutants, as was the transsulfuration pathway, leading also to a disruption in redox homeostasis. The major consequences, however, appeared to be in the depletion of nucleotides. C isotope labeling and metabolic studies revealed that the deregulated MTHFR cells undergo continuous transmethylation of homocysteine by methyltetrahydrofolate (CHTHF) to form methionine. This reaction also drives SAM formation and further depletes ATP reserves. SAM was then cycled back to methionine, leading to futile cycles of SAM synthesis and recycling and explaining the necessity for MTHFR to be regulated by SAM. The study has yielded valuable new insights into the regulation of one-carbon metabolism, and the mutants appear as powerful new tools to further dissect out the intersection of one-carbon metabolism with various pathways both in yeasts and in humans.

}, keywords = {Adenosine Triphosphate, Allosteric Regulation, Humans, Methylation, Methylenetetrahydrofolate Reductase (NADPH2), S-Adenosylmethionine, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins}, issn = {1083-351X}, doi = {10.1074/jbc.RA120.015129}, author = {Bhatia, Muskan and Thakur, Jyotika and Suyal, Shradha and Oniel, Ruchika and Chakraborty, Rahul and Pradhan, Shalini and Sharma, Monika and Sengupta, Shantanu and Laxman, Sunil and Masakapalli, Shyam Kumar and Bachhawat, Anand Kumar} } @article {1985, title = {Anabolic SIRT4 Exerts Retrograde Control over TORC1 Signaling by Glutamine Sparing in the Mitochondria.}, journal = {Mol Cell Biol}, volume = {40}, year = {2020}, month = {2020 Jan 03}, abstract = {

Anabolic and catabolic signaling mediated via mTOR and AMPK (AMP-activated kinase) have to be intrinsically coupled to mitochondrial functions for maintaining homeostasis and mitigate cellular/organismal stress. Although glutamine is known to activate mTOR, whether and how differential mitochondrial utilization of glutamine impinges on mTOR signaling has been less explored. Mitochondrial SIRT4, which unlike other sirtuins is induced in a fed state, is known to inhibit catabolic signaling/pathways through the AMPK-PGC1α/SIRT1-peroxisome proliferator-activated receptor α (PPARα) axis and negatively regulate glutamine metabolism via the tricarboxylic acid cycle. However, physiological significance of SIRT4 functions during a fed state is still unknown. Here, we establish SIRT4 as key anabolic factor that activates TORC1 signaling and regulates lipogenesis, autophagy, and cell proliferation. Mechanistically, we demonstrate that the ability of SIRT4 to inhibit anaplerotic conversion of glutamine to α-ketoglutarate potentiates TORC1. Interestingly, we also show that mitochondrial glutamine sparing or utilization is critical for differentially regulating TORC1 under fed and fasted conditions. Moreover, we conclusively show that differential expression of SIRT4 during fed and fasted states is vital for coupling mitochondrial energetics and glutamine utilization with anabolic pathways. These significant findings also illustrate that SIRT4 integrates nutrient inputs with mitochondrial retrograde signals to maintain a balance between anabolic and catabolic pathways.

}, issn = {1098-5549}, doi = {10.1128/MCB.00212-19}, author = {Shaw, Eisha and Talwadekar, Manasi and Rashida, Zeenat and Mohan, Nitya and Acharya, Aishwarya and Khatri, Subhash and Laxman, Sunil and Kolthur-Seetharam, Ullas} } @article {2217, title = {Comparison of CryoEM and X-ray structures of dimethylformamidase.}, journal = {Prog Biophys Mol Biol}, year = {2020}, month = {2020 Jul 28}, abstract = {

Dimethylformamidase (DMFase) catalyzes the hydrolysis of dimethylformamide, an industrial solvent, introduced into the environment by humans. Recently, we determined the structures of dimethylformamidase by electron cryo microscopy and X-ray crystallography revealing a tetrameric enzyme with a mononuclear iron at the active site. DMFase from Paracoccus sp. isolated from a waste water treatment plant around the city of Kanpur in India shows maximal activity at 54\ {\textdegree}C and is halotolerant. The structures determined by both techniques are mostly identical and the largest difference is in a loop near the active site. This loop could play a role in co-operativity between the monomers. A number of non-protein densities are observed in the EM map, which are modelled as water molecules. Comparison of the structures determined by the two methods reveals conserved water molecules that could play a structural role. The higher stability, unusual active site and negligible activity at low temperature makes this a very good model to study enzyme mechanism by cryoEM.

}, issn = {1873-1732}, doi = {10.1016/j.pbiomolbio.2020.06.008}, author = {Vinothkumar, Kutti R and Arya, Chetan Kumar and Ramanathan, Gurunath and Subramanian, Ramaswamy} } @article {2065, title = {Convolvulus pluricaulis extract can modulate synaptic plasticity in rat brain hippocampus.}, journal = {Neuroreport}, volume = {31}, year = {2020}, month = {2020 May 22}, pages = {597-604}, abstract = {

The memory-boosting property of Indian traditional herb, Convolvulus pluricaulis, has been documented in literature; however, its effect on synaptic plasticity has not yet been reported. Two important forms of synaptic plasticity known to be involved in the processes of memory formation are long-term potentiation (LTP) and long-term depression (LTD). In the present study, the effect of C. pluricaulis plant extract on LTP and LTD were evaluated. The adult male Wistar rats were fed orally with 250, 500 and 1000 mg/kg of this extract for 4 weeks and the effect was determined on LTP and LTD in the Schaffer collaterals of the hippocampal cornu ammonis region CA1. We found that the 500 mg/kg dose of the extract could significantly enhance LTP compared to the vehicle treated ones. Moreover, the same dose could also reduce LTD while used in a separate set of animals. Also, a fresh group of animals treated with the effective dose (500 mg/kg) of plant extract were examined for memory retention in two behavioral platforms namely, contextual fear conditioning (CFC) and novel object recognition test (NORT). Increased fear response to the conditioned stimulus and enhanced recognition of objects were observed in CFC and NORT, respectively, both indicating strengthening of memory. Following up, ex-vivo electrophysiology experiments were performed with the active single molecule scopoletin, present in C. pluricaulis extract and similar patterns in synaptic plasticity changes were obtained. These findings suggest that prolonged treatment of C. pluricaulis extract, at a specific dose in healthy animals, can augment memory functions by modulating hippocampal plasticity.

}, issn = {1473-558X}, doi = {10.1097/WNR.0000000000001446}, author = {Das, Rishi and Sengupta, Tathagata and Roy, Shubhrajit and Chattarji, Sumantra and Ray, Jharna} } @article {2110, title = {Cortical neurons derived from human pluripotent stem cells lacking FMRP display altered spontaneous firing patterns.}, journal = {Mol Autism}, volume = {11}, year = {2020}, month = {2020 Jun 19}, pages = {52}, abstract = {

BACKGROUND: Fragile X syndrome (FXS), a neurodevelopmental disorder, is a leading monogenetic cause of intellectual disability and autism spectrum disorder. Notwithstanding the extensive studies using rodent and other pre-clinical models of FXS, which have provided detailed mechanistic insights into the pathophysiology of this disorder, it is only relatively recently that human stem cell-derived neurons have been employed as a model system to further our understanding of the pathophysiological events that may underlie FXS. Our study assesses the physiological properties of human pluripotent stem cell-derived cortical neurons lacking fragile X mental retardation protein (FMRP).

METHODS: Electrophysiological whole-cell voltage- and current-clamp recordings were performed on two control and three FXS patient lines of human cortical neurons derived from induced pluripotent stem cells. In addition, we also describe the properties of an isogenic pair of lines in one of which FMR1 gene expression has been silenced.

RESULTS: Neurons lacking FMRP displayed bursts of spontaneous action potential firing that were more frequent but shorter in duration compared to those recorded from neurons expressing FMRP. Inhibition of large conductance Ca-activated K currents and the persistent Na current in control neurons phenocopies action potential bursting observed in neurons lacking FMRP, while in neurons lacking FMRP pharmacological potentiation of voltage-dependent Na channels phenocopies action potential bursting observed in control neurons. Notwithstanding the changes in spontaneous action potential firing, we did not observe any differences in the intrinsic properties of neurons in any of the lines examined. Moreover, we did not detect any differences in the properties of miniature excitatory postsynaptic currents in any of the lines.

CONCLUSIONS: Pharmacological manipulations can alter the action potential burst profiles in both control and FMRP-null human cortical neurons, making them appear like their genetic counterpart. Our studies indicate that FMRP targets that have been found in rodent models of FXS are also potential targets in a human-based model system, and we suggest potential mechanisms by which activity is altered.

}, issn = {2040-2392}, doi = {10.1186/s13229-020-00351-4}, author = {Das Sharma, Shreya and Pal, Rakhi and Reddy, Bharath Kumar and Selvaraj, Bhuvaneish T and Raj, Nisha and Samaga, Krishna Kumar and Srinivasan, Durga J and Ornelas, Loren and Sareen, Dhruv and Livesey, Matthew R and Bassell, Gary J and Svendsen, Clive N and Kind, Peter C and Chandran, Siddharthan and Chattarji, Sumantra and Wyllie, David J A} } @article {2146, title = {Differential flexibility leading to crucial microelastic properties of asymmetric lipid vesicles for cellular transfection: A combined spectroscopic and atomic force microscopy studies.}, journal = {Colloids Surf B Biointerfaces}, volume = {196}, year = {2020}, month = {2020 Sep 21}, pages = {111363}, abstract = {

The role of microscopic elasticity of nano-carriers in cellular uptake is an important aspect in biomedical research. Herein we have used AFM nano-indentation force spectroscopy and F{\"o}rster resonance energy transfer (FRET) measurements to probe microelastic properties of three novel cationic liposomes based on di-alkyl dihydroxy ethyl ammonium chloride based lipids having asymmetry in their hydrophobic chains (Lip1818, Lip1814 and Lip1810). AFM data reveals that symmetry in hydrophobic chains of a cationic lipid (Lip1818) imparts higher rigidity to the resulting liposomes than those based on asymmetric lipids (Lip1814 and Lip1810). The stiffness of the cationic liposomes is found to decrease with increasing asymmetry in the hydrophobic lipid chains in the order of Lip1818 \> Lip1814 \> lip1810. FRET measurements between Coumarin 500 (Donor) and Merocyanine 540 (Acceptor) have revealed that full width at half-maxima (hw) of the probability distribution (P(r)) of donor-acceptor distance (r), increases in an order Lip1818 \< Lip1814 \< Lip1810 with increasing asymmetry of the hydrophobic lipid chains. This increase in width (hw) of the donor-acceptor distance distributions is reflective of increasing flexibility of the liposomes with increasing asymmetry of their constituent lipids. Thus, the results from AFM and FRET studies are complementary to each other and indicates that an increase in asymmetry of the hydrophobic lipid chains increases elasticity and or flexibility of the corresponding liposomes. Cell biology experiments confirm that liposomal flexibility or rigidity directly influences their cellular transfection efficiency, where Lip1814 is found to be superior than the other two liposomes manifesting that a critical balance between flexibility and rigidity of the cationic liposomes is key to efficient cellular uptake. Taken together, our studies reveal how asymmetry in the molecular architecture of the hydrophobic lipid chains influences the microelastic properties of the liposomes, and hence, their cellular uptake efficiency.

}, issn = {1873-4367}, doi = {10.1016/j.colsurfb.2020.111363}, author = {Mukherjee, Dipanjan and Rakshit, Tatini and Singh, Priya and Mondal, Suman and Paul, Debashish and Ahir, Manisha and Adhikari, Arghya and Puthiyapurayil, Theja P and Vemula, Praveen Kumar and Senapati, Dulal and Das, Ranjan and Pal, Samir Kumar} } @article {2067, title = {Distinct regulation of bioenergetics and translation by group I mGluR and NMDAR.}, journal = {EMBO Rep}, year = {2020}, month = {2020 Apr 29}, pages = {e48037}, abstract = {

Neuronal activity is responsible for the high energy consumption in the brain. However, the cellular mechanisms draining ATP upon the arrival of a stimulus are yet to be explored systematically at the post-synapse. Here, we provide evidence that a significant fraction of ATP is consumed upon glutamate stimulation to energize mGluR-induced protein synthesis. We find that both mGluR and NMDAR alter protein synthesis and ATP consumption with distinct kinetics at the synaptic-dendritic compartments. While mGluR activation leads to a rapid and sustained reduction in neuronal ATP levels, NMDAR activation has no immediate impact on the same. ATP consumption correlates inversely with the kinetics of protein synthesis for both receptors. We observe a persistent elevation in protein synthesis within 5\ minutes of mGluR activation and a robust inhibition of the same within 2\ minutes of NMDAR activation, assessed by the phosphorylation status of eEF2 and metabolic labeling. However, a delayed protein synthesis-dependent ATP expenditure ensues after 15\ minutes of NMDAR stimulation. We identify a central role for AMPK in the correlation between protein synthesis and ATP consumption. AMPK is dephosphorylated and inhibited upon mGluR activation, while it is phosphorylated upon NMDAR activation. Perturbing AMPK activity disrupts receptor-specific modulations of eEF2 phosphorylation and protein synthesis. Our observations, therefore, demonstrate that the regulation of the AMPK-eEF2 signaling axis by glutamate receptors alters neuronal protein synthesis and bioenergetics.

}, issn = {1469-3178}, doi = {10.15252/embr.201948037}, author = {Ghosh Dastidar, Sudhriti and Das Sharma, Shreya and Chakraborty, Sumita and Chattarji, Sumantra and Bhattacharya, Aditi and Muddashetty, Ravi S} } @article {2151, title = {Dopamine requires unique residues to signal via the serotonin 2A receptor.}, journal = {Neuroscience}, volume = {439}, year = {2020}, month = {2020 07 15}, pages = {319-331}, abstract = {

Serotonin is an important neurotransmitter and neuromodulator. Disruption of the serotonergic system has been implicated in various psychiatric disorders such as schizophrenia and bipolar disorder. Most of the drugs targeting these neurotransmitter systems are classified primarily as agonists or inverse agonists/antagonists, with their described function being limited to activating the canonical signaling pathway(s), or inhibiting the pathway(s) respectively. Previous work with the human 5-HT has shown the receptor to be activated by dopamine, also an endogenous ligand. Dopamine is the cognate ligand of the dopaminergic system, which significantly overlaps with the serotonergic system in the brain. The two systems innervate many of the same brain areas, and the central serotonergic systems also regulate dopamine functions. Our aim was to investigate the downstream signaling set up by the receptor on being activated by dopamine. We show that dopamine is a functionally selective ligand at 5-HT and have examined dopamine as a ligand with respect to some receptor-dependent phenotypes. Our results show that dopamine acts as an agonist at the human serotonin 2A receptor and brings about its activation and internalization. Using in vitro assays, we have established differences in the signaling pathways set up by dopamine as compared to serotonin. Using site-specific mutagenesis we have identified residues important for this functional selectivity, shown by dopamine at this receptor. Our identification of specific residues important in the functional selectivity of dopamine at 5-HT could have far reaching implications for the field of GPCR signaling and drug-design. This article is part of a Special Issue entitled: Honoring Ricardo Miledi - outstanding neuroscientist of XX-XXI centuries.

}, issn = {1873-7544}, doi = {10.1016/j.neuroscience.2019.03.056}, author = {Soman, Shuchita and Bhattacharya, Aditi and Panicker, Mitradas M} } @article {2115, title = {A field-based quantitative analysis of sublethal effects of air pollution on pollinators.}, journal = {Proc Natl Acad Sci U S A}, volume = {117}, year = {2020}, month = {2020 Aug 25}, pages = {20653-20661}, abstract = {

While the impact of air pollution on human health is well studied, mechanistic impacts of air pollution on wild systems, including those providing essential ecosystem services, are largely unknown, but directly impact our health and well-being. India is the world{\textquoteright}s largest fruit producer, second most populous country, and contains 9 of the world{\textquoteright}s 10 most polluted cities. Here, we sampled Giant Asian honey bees, , at locations with varying air pollution levels in Bangalore, India. We observed significant correlations between increased respirable suspended particulate matter (RSPM) deposition and changes in bee survival, flower visitation, heart rate, hemocyte levels, and expression of genes related to lipid metabolism, stress, and immunity. Lab-reared exposed to these same sites also exhibited similar molecular and physiological differences. Our study offers a quantitative analysis on the current impacts of air pollution on insects, and indicates the urgency for more nonhuman studies to accurately assess the effects of pollution on our natural world.

}, issn = {1091-6490}, doi = {10.1073/pnas.2009074117}, author = {Thimmegowda, Geetha G and Mullen, Susan and Sottilare, Katie and Sharma, Ankit and Mohanta, Saptashi Soham and Brockmann, Axel and Dhandapany, Perundurai S and Olsson, Shannon B} } @article {2251, title = {Generic Molding Platform for Simple, Low-Cost Fabrication of Polymeric Microneedles}, journal = {Macromolecular Materials and Engineering}, volume = {305}, year = {2020}, abstract = {

Micromolding technology is widely used for the fabrication of polymer microneedles for transdermal and intradermal drug delivery applications. Geometric features of microneedles in molding are solely determined by geometry of the master mold template. Fabrication of master mold template usually involves costly and cumbersome technologies due to small feature sizes typical of microneedles. In this research, a novel molding platform is designed that is fabricated using low-cost and simple techniques with flexibility of producing large number of microneedle geometries. The proposed molding platform eliminates need for developing multiple mold templates for fabrication of various geometries of polymer microneedles. Utility of this molding platform is demonstrated in polylactic acid-based solid thermoplastic microneedles and polyacrylic acid-based dissolvable microneedles with various aspect ratio settings. Various microneedles fabricated at heights differing with resolution of as low as 100 {\textmu}m are successfully achieved using specified settings in the molding platform. The suitability of fabricated microneedles for drug delivery applications is evaluated by in vitro and in vivo testing.

}, author = {7. Badnikar, K. and Jayadevi, S. N. and Pahal, S. and Sripada, S. and Nayak, M. M. and Vemula, P. K. and Subrahmanyam, D. N.} } @article {2155, title = {Genetic, clinical, molecular, and pathogenic aspects of the South Asian-specific polymorphic MYBPC3 variant.}, journal = {Biophys Rev}, volume = {12}, year = {2020}, month = {2020 Aug}, pages = {1065-1084}, abstract = {

Hypertrophic cardiomyopathy (HCM) is a cardiac genetic disease characterized by ventricular enlargement, diastolic dysfunction, and increased risk for sudden cardiac death. Sarcomeric genetic defects are the predominant known cause of HCM. In particular, mutations in the myosin-binding protein C gene (MYBPC3) are associated with ~ 40\% of all HCM cases in which a genetic basis has been established. A decade ago, our group reported a 25-base pair deletion in intron 32 of MYBPC3 (MYBPC3) that is uniquely prevalent in South Asians and is associated with autosomal dominant cardiomyopathy. Although our studies suggest that this deletion results in left ventricular dysfunction, cardiomyopathies, and heart failure, the precise mechanism by which this variant predisposes to heart disease remains unclear. Increasingly appreciated, however, is the contribution of secondary risk factors, additional mutations, and lifestyle choices in augmenting or modifying the HCM phenotype in MYBPC3 carriers. Therefore, the goal of this review article is to summarize the current research dedicated to understanding the molecular pathophysiology of HCM in South Asians with the MYBPC3 variant. An emphasis is to review the latest techniques currently applied to explore the MYBPC3 pathogenesis and to provide a foundation for developing new diagnostic strategies and advances in therapeutics.

}, issn = {1867-2450}, doi = {10.1007/s12551-020-00725-1}, author = {Arif, Mohammed and Nabavizadeh, Pooneh and Song, Taejeong and Desai, Darshini and Singh, Rohit and Bazrafshan, Sholeh and Kumar, Mohit and Wang, Yigang and Gilbert, Richard J and Dhandapany, Perundurai S and Becker, Richard C and Kranias, Evangelia G and Sadayappan, Sakthivel} } @article {2148, title = {Genetically encoded live-cell sensor for tyrosinated microtubules.}, journal = {J Cell Biol}, volume = {219}, year = {2020}, month = {2020 Oct 05}, abstract = {

Microtubule cytoskeleton exists in various biochemical forms in different cells due to tubulin posttranslational modifications (PTMs). Tubulin PTMs are known to affect microtubule stability, dynamics, and interaction with MAPs and motors in a specific manner, widely known as tubulin code hypothesis. At present, there exists no tool that can specifically mark tubulin PTMs in living cells, thus severely limiting our understanding of their dynamics and cellular functions. Using a yeast display library, we identified a binder against terminal tyrosine of α-tubulin, a unique PTM site. Extensive characterization validates the robustness and nonperturbing nature of our binder as tyrosination sensor, a live-cell tubulin nanobody specific towards tyrosinated microtubules. Using this sensor, we followed nocodazole-, colchicine-, and vincristine-induced depolymerization events of tyrosinated microtubules in real time and found each distinctly perturbs the microtubule polymer. Together, our work describes a novel tyrosination sensor and its potential applications to study the dynamics of microtubule and their PTM processes in living cells.

}, issn = {1540-8140}, doi = {10.1083/jcb.201912107}, author = {Kesarwani, Shubham and Lama, Prakash and Chandra, Anchal and Reddy, P Purushotam and Jijumon, A S and Bodakuntla, Satish and Rao, Balaji M and Janke, Carsten and Das, Ranabir and Sirajuddin, Minhajuddin} } @article {2203, title = {Genome-scale reconstruction of Gcn4/ATF4 networks driving a growth program.}, journal = {PLoS Genet}, volume = {16}, year = {2020}, month = {2020 12}, pages = {e1009252}, abstract = {

Growth and starvation are considered opposite ends of a spectrum. To sustain growth, cells use coordinated gene expression programs and manage biomolecule supply in order to match the demands of metabolism and translation. Global growth programs complement increased ribosomal biogenesis with sufficient carbon metabolism, amino acid and nucleotide biosynthesis. How these resources are collectively managed is a fundamental question. The role of the Gcn4/ATF4 transcription factor has been best studied in contexts where cells encounter amino acid starvation. However, high Gcn4 activity has been observed in contexts of rapid cell proliferation, and the roles of Gcn4 in such growth contexts are unclear. Here, using a methionine-induced growth program in yeast, we show that Gcn4/ATF4 is the fulcrum that maintains metabolic supply in order to sustain translation outputs. By integrating matched transcriptome and ChIP-Seq analysis, we decipher genome-wide direct and indirect roles for Gcn4 in this growth program. Genes that enable metabolic precursor biosynthesis indispensably require Gcn4; contrastingly ribosomal genes are partly repressed by Gcn4. Gcn4 directly binds promoter-regions and transcribes a subset of metabolic genes, particularly driving lysine and arginine biosynthesis. Gcn4 also globally represses lysine and arginine enriched transcripts, which include genes encoding the translation machinery. The Gcn4 dependent lysine and arginine supply thereby maintains the synthesis of the translation machinery. This is required to maintain translation capacity. Gcn4 consequently enables metabolic-precursor supply to bolster protein synthesis, and drive a growth program. Thus, we illustrate how growth and starvation outcomes are both controlled using the same Gcn4 transcriptional outputs that function in distinct contexts.

}, keywords = {Basic-Leucine Zipper Transcription Factors, Cell Proliferation, Gene Expression Regulation, Fungal, Gene Regulatory Networks, Genome, Fungal, Ribosomes, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Transcriptional Activation}, issn = {1553-7404}, doi = {10.1371/journal.pgen.1009252}, author = {Srinivasan, Rajalakshmi and Walvekar, Adhish S and Rashida, Zeenat and Seshasayee, Aswin and Laxman, Sunil} } @article {2078, title = {Improved detection of RNA foci in amyotrophic lateral sclerosis post-mortem tissue using BaseScope{\texttrademark} shows a lack of association with cognitive dysfunction.}, journal = {Brain Commun}, volume = {2}, year = {2020}, month = {2020 Jan 31}, pages = {fcaa009}, abstract = {

The hexanucleotide repeat expansion is the commonest known genetic mutation in amyotrophic lateral sclerosis. A neuropathological hallmark is the intracellular accumulation of RNA foci. The role that RNA foci play in the pathogenesis of amyotrophic lateral sclerosis is widely debated. Historically, RNA foci have been identified using hybridization. Here, we have implemented BaseScope{\texttrademark}, a high-resolution modified hybridization technique. We demonstrate that previous studies have underestimated the abundance of RNA foci in neurons and glia. This improved detection allowed us to investigate the abundance, regional distribution and cell type specificity of antisense RNA foci in post-mortem brain and spinal cord tissue of six deeply clinically phenotyped patients and six age- and sex-matched controls. We find a correlation between RNA foci and the accumulation of transactive response DNA-binding protein of 43 kDa in spinal motor neurons ( = 0.93; = 0.008), but not in glia or cortical motor neurons. We also demonstrate that there is no correlation between the presence of RNA foci and the accumulation of transactive response DNA binding protein of 43 kDa in extra-motor brain regions. Furthermore, there is no association between the presence of RNA foci and cognitive indices. These results highlight the utility of BaseScope{\texttrademark} in the clinicopathological assessment of the role of antisense RNA foci in .

}, issn = {2632-1297}, doi = {10.1093/braincomms/fcaa009}, author = {Mehta, Arpan R and Selvaraj, Bhuvaneish T and Barton, Samantha K and McDade, Karina and Abrahams, Sharon and Chandran, Siddharthan and Smith, Colin and Gregory, Jenna M} } @article {2068, title = {Lithium response in bipolar disorder correlates with improved cell viability of patient derived cell lines.}, journal = {Sci Rep}, volume = {10}, year = {2020}, month = {2020 May 04}, pages = {7428}, abstract = {

Lithium is an effective, well-established treatment for bipolar disorder (BD). However, the mechanisms of its action, and reasons for variations in clinical response, are unclear. We used neural precursor cells (NPCs) and lymphoblastoid cell lines (LCLs), from BD patients characterized for clinical response to lithium (using the "Alda scale" and "NIMH Retrospective Life chart method"), to interrogate cellular phenotypes related to both disease and clinical lithium response. NPCs from two biologically related BD patients who differed in their clinical response to lithium were compared with healthy controls. RNA-Seq and analysis, mitochondrial membrane potential (MMP), cell viability, and cell proliferation parameters were assessed, with and without in vitro lithium. These parameters were also examined in LCLs from 25 BD patients (16 lithium responders and 9 non-responders), and 12 controls. MMP was lower in both NPCs and LCLs from BD; but it was reversed with in vitro lithium only in LCLs, and this was unrelated to clinical lithium response. The higher cell proliferation observed in BD was unaffected by in vitro lithium. Cell death was greater in BD. However, LCLs from clinical lithium responders could be rescued by addition of in vitro lithium. In vitro lithium also enhanced BCL2 and GSK3B expression in these cells. Our findings indicate cellular phenotypes related to the disease (MMP, cell proliferation) in both NPCs and LCLs; and those related to clinical lithium response (cell viability, BCL2/GSK3B expression) in LCLs.

}, issn = {2045-2322}, doi = {10.1038/s41598-020-64202-1}, author = {Paul, Pradip and Iyer, Shruti and Nadella, Ravi Kumar and Nayak, Rashmitha and Chellappa, Anirudh S and Ambardar, Sheetal and Sud, Reeteka and Sukumaran, Salil K and Purushottam, Meera and Jain, Sanjeev and Viswanath, Biju} } @article {2208, title = {Metabolic control of cellular immune-competency by odors in .}, journal = {Elife}, volume = {9}, year = {2020}, month = {2020 12 29}, abstract = {

Studies in different animal model systems have revealed the impact of odors on immune cells; however, any understanding on why and how odors control cellular immunity remained unclear. We find that employ an olfactory-immune cross-talk to tune a specific cell type, the lamellocytes, from hematopoietic-progenitor cells. We show that neuronally released GABA derived upon olfactory stimulation is utilized by blood-progenitor cells as a metabolite and through its catabolism, these cells stabilize Sima/HIFα protein. Sima capacitates blood-progenitor cells with the ability to initiate lamellocyte differentiation. This systemic axis becomes relevant for larvae dwelling in wasp-infested environments where chances of infection are high. By co-opting the olfactory route, the preconditioned animals elevate their systemic GABA levels leading to the upregulation of blood-progenitor cell Sima expression. This elevates their immune-potential and primes them to respond rapidly when infected with parasitic wasps. The present work highlights the importance of the olfaction in immunity and shows how odor detection during animal development is utilized to establish a long-range axis in the control of blood-progenitor competency and immune-priming.

}, issn = {2050-084X}, doi = {10.7554/eLife.60376}, author = {Madhwal, Sukanya and Shin, Mingyu and Kapoor, Ankita and Goyal, Manisha and Joshi, Manish K and Ur Rehman, Pirzada Mujeeb and Gor, Kavan and Shim, Jiwon and Mukherjee, Tina} } @article {2144, title = {Methylated PP2A stabilizes Gcn4 to enable a methionine-induced anabolic program.}, journal = {J Biol Chem}, year = {2020}, month = {2020 Oct 29}, abstract = {

Methionine, through S-adenosylmethionine, activates a multifaceted growth program in which ribosome biogenesis, carbon metabolism, amino acid and nucleotide biosynthesis are induced. This growth program requires the activity of the Gcn4 transcription factor (called ATF4 in mammals), which facilitates the supply of metabolic precursors that are essential for anabolism. However, how Gcn4 itself is regulated in the presence of methionine is unknown. Here, we discover that Gcn4 protein levels are increased by methionine, despite conditions of high cell growth and translation (where the roles of Gcn4 are not well studied). We demonstrate that this mechanism of Gcn4 induction is independent of transcription, as well as the conventional Gcn2/eIF2α-mediated increased translation of Gcn4. Instead, when methionine is abundant, Gcn4 phosphorylation is decreased, which reduces its ubiquitination and therefore degradation. Gcn4 is dephosphorylated by the protein phosphatase PP2A; our data show that when methionine is abundant, the conserved methyltransferase Ppm1 methylates and alters the activity of the catalytic subunit of PP2A, shifting the balance of Gcn4 towards a dephosphorylated, stable state. The absence of Ppm1 or the loss of the PP2A methylation destabilizes Gcn4 even when methionine is abundant, leading to collapse of the Gcn4-dependent anabolic program. These findings reveal a novel, methionine-dependent signaling and regulatory axis. Here methionine directs a conserved methyltransferase Ppm1, via its target phosphatase PP2A, to selectively stabilize Gcn4. Through this, cells conditionally modify a major phosphatase to stabilize a metabolic master-regulator and drive anabolism.

}, issn = {1083-351X}, doi = {10.1074/jbc.RA120.014248}, author = {Walvekar, Adhish S and Kadamur, Ganesh and Sreedharan, Sreesa and Gupta, Ritu and Srinivasan, Rajalakshmi and Laxman, Sunil} } @article {2205, title = {Methylated PP2A stabilizes Gcn4 to enable a methionine-induced anabolic program.}, journal = {J Biol Chem}, volume = {295}, year = {2020}, month = {2020 Dec 25}, pages = {18390-18405}, abstract = {

Methionine, through S-adenosylmethionine, activates a multifaceted growth program in which ribosome biogenesis, carbon metabolism, and amino acid and nucleotide biosynthesis are induced. This growth program requires the activity of the Gcn4 transcription factor (called ATF4 in mammals), which facilitates the supply of metabolic precursors that are essential for anabolism. However, how Gcn4 itself is regulated in the presence of methionine is unknown. Here, we discover that Gcn4 protein levels are increased by methionine, despite conditions of high cell growth and translation (in which the roles of Gcn4 are not well-studied). We demonstrate that this mechanism of Gcn4 induction is independent of transcription, as well as the conventional Gcn2/eIF2α-mediated increased translation of Gcn4. Instead, when methionine is abundant, Gcn4 phosphorylation is decreased, which reduces its ubiquitination and therefore degradation. Gcn4 is dephosphorylated by the protein phosphatase 2A (PP2A); our data show that when methionine is abundant, the conserved methyltransferase Ppm1 methylates and alters the activity of the catalytic subunit of PP2A, shifting the balance of Gcn4 toward a dephosphorylated, stable state. The absence of Ppm1 or the loss of the PP2A methylation destabilizes Gcn4 even when methionine is abundant, leading to collapse of the Gcn4-dependent anabolic program. These findings reveal a novel, methionine-dependent signaling and regulatory axis. Here methionine directs the conserved methyltransferase Ppm1 via its target phosphatase PP2A to selectively stabilize Gcn4. Through this, cells conditionally modify a major phosphatase to stabilize a metabolic master regulator and drive anabolism.

}, issn = {1083-351X}, doi = {10.1074/jbc.RA120.014248}, author = {Walvekar, Adhish S and Kadamur, Ganesh and Sreedharan, Sreesa and Gupta, Ritu and Srinivasan, Rajalakshmi and Laxman, Sunil} } @article {2107, title = {Microbial Metabolite Urolithin B Inhibits Recombinant Human Monoamine Oxidase A Enzyme.}, journal = {Metabolites}, volume = {10}, year = {2020}, month = {2020 Jun 19}, abstract = {

Urolithins are gut microbial metabolites derived from ellagitannins (ET) and ellagic acid (EA), and shown to exhibit anticancer, anti-inflammatory, anti-microbial, anti-glycative and anti-oxidant activities. Similarly, the parent molecules, ET and EA are reported for their neuroprotection and antidepressant activities. Due to the poor bioavailability of ET and EA, the in vivo functional activities cannot be attributed exclusively to these compounds. Elevated monoamine oxidase (MAO) activities are responsible for the inactivation of monoamine neurotransmitters in neurological disorders, such as depression and Parkinson{\textquoteright}s disease. In this study, we examined the inhibitory effects of urolithins (A, B and C) and EA on MAO activity using recombinant human MAO-A and MAO-B enzymes. Urolithin B was found to be a better MAO-A enzyme inhibitor among the tested urolithins and EA with an IC value of 0.88 {\textmu}M, and displaying a mixed mode of inhibition. However, all tested compounds exhibited higher IC (\>100 {\textmu}M) for MAO-B enzyme.

}, issn = {2218-1989}, doi = {10.3390/metabo10060258}, author = {Singh, Rajbir and Chandrashekharappa, Sandeep and Vemula, Praveen Kumar and Haribabu, Bodduluri and Jala, Venkatakrishna Rao} } @article {2109, title = {Modulation of β-catenin levels regulates cranial neural crest patterning and dispersal into first pharyngeal arch.}, journal = {Dev Dyn}, year = {2020}, month = {2020 May 19}, abstract = {

BACKGROUND: Vertebrate cranial neural crest cells (CNCCs) are multipotent, proximal to the source CNCC form the cranial ganglia. Distally, in the pharyngeal arches, they give rise to the craniofacial skeleton and connective tissues. Fate choices are made as CNCC pattern into distinct destination compartments. In spite of this importance, the mechanism patterning CNCC is poorly defined.

RESULTS: Here, we report that a novel β-catenin-dependent regulation of N-Cadherin levels may drive CNCC patterning. In mouse embryos, at the first pharyngeal arch axial level, membrane β-catenin levels correlate with the extent of N-cadherin-mediated adhesion and thus suggest the presence of collective and dispersed states of CNCC. Using in vitro human neural crest model and chemical modulators of β-catenin levels, we show a requirement for down-modulating β-catenin for regulating N-cadherin levels and cell-cell adhesion. Similarly, in β-catenin gain-of-function mutant mouse embryos, CNCC fail to lower N-cadherin levels. This indicates a failure to reduce cell-cell adhesion, which may underlie the failure of mutant CNCC to populate first pharyngeal arch.

CONCLUSION: We suggest that β-catenin-mediated regulation of CNCC adhesion, a previously underappreciated mechanism, underlies the patterning of CNCC into fate-specific compartments.

}, issn = {1097-0177}, doi = {10.1002/dvdy.208}, author = {Javali, Alok and Lakshmanan, Vairavan and Palakodeti, Dasaradhi and Sambasivan, Ramkumar} } @article {2063, title = {N-terminal variant Asp14Asn of the human p70 S6 Kinase 1 enhances translational signaling causing different effects in developing and mature neuronal cells.}, journal = {Neurobiol Learn Mem}, volume = {171}, year = {2020}, month = {2020 May}, pages = {107203}, abstract = {

The ribosomal p70 S6 Kinase 1 (S6K1) has been implicated in the etiology of complex neurological diseases including autism, depression and dementia. Though no major gene disruption has been reported in humans in RPS6KB1, single nucleotide variants (SNVs) causing missense mutations have been identified, which have not been assessed for their impact on protein function. These S6K1 mutations have the potential to influence disease progression and treatment response. We mined the Simon Simplex Collection (SSC) and SPARK autism database to find inherited SNVs in S6K1 and characterized the effect of two missense SNVs, Asp14Asn (allele frequency\ =\ 0.03282\%) and Glu44Gln (allele frequency\ =\ 0.0008244\%), on S6K1 function in HEK293, human ES cells and primary neurons. Expressing Asp14Asn in HEK293 cells resulted in increased basal phosphorylation of downstream targets of S6K1 and increased de novo translation. This variant also showed blunted response to the specific S6K1 inhibitor, FS-115. In human embryonic cell line Shef4, Asp14Asn enhanced spontaneous neural fate specification in the absence of differentiating growth factors. In addition to enhanced translation, neurons expressing Asp14Asn exhibited impaired dendritic arborization and increased levels of phosphorylated ERK 1/2. Finally, in the SSC families tracked, Asp14Asn segregated with lower IQ scores when found in the autistic individual rather than the unaffected sibling. The Glu44Gln mutation showed a milder, but opposite phenotype in HEK cells as compared to Asp14Asn. Although the Glu44Gln mutation displayed increased neuronal translation, it had no impact on neuronal morphology. Our results provide the first characterization of naturally occurring human S6K1 variants on cognitive phenotype, neuronal morphology and maturation, underscoring again the importance of translation control in neural development and plasticity.

}, issn = {1095-9564}, doi = {10.1016/j.nlm.2020.107203}, author = {Venkatasubramani, Janani Priya and Subramanyam, Prakash and Pal, Rakhi and Reddy, Bharath K and Srinivasan, Durga Jeyalakshmi and Chattarji, Sumantra and Iossifov, Ivan and Klann, Eric and Bhattacharya, Aditi} } @article {2038, title = {Nucleolar localization of the Notch4 intracellular domain underpins its regulation of the cellular response to genotoxic stressors.}, journal = {Cell Death Discov}, volume = {6}, year = {2020}, month = {2020}, pages = {7}, abstract = {

Cell survival is one of the many cellular processes regulated by Notch family of proteins. A comparison of human breast cancer cell lines, which differ in the levels of endogenous Notch4, implicated the protein in regulating susceptibility to apoptosis triggered by genomic damage. In agreement with this observation, increased susceptibility to genotoxic damage was observed following siRNA ablations of Notch4 in two breast cancer cell lines. Further, overexpressing Notch4 intracellular domain (NIC4) tagged to GFP (NIC4-GFP), protected cells from apoptosis triggered by genotoxic drugs. In cells immune-stained for endogenous Notch4, protein was detected in the nucleolus and nucleoplasm, which was also confirmed by the co-localization of NIC4-GFP with RFP-tagged nucleolar proteins in breast cancer cells or the unrelated HEK cell line. Linking functional outcomes to nucleolar localization, NIC4-GFP protection from apoptosis, required the nucleolar proteins Nucleolin and Fibrillarin. Consistently, immunoprecipitation analysis revealed associations between nucleolar proteins-Nucleolin and Nucleophosmin-and Notch4. Microscopy-based biophysical analysis of live cells showed that nucleolar and nucleoplasmic pools of NIC4-GFP are mobile, with some sequestration of nucleolar NIC4-GFP pools. A nucleolar excluded form, NIC4_3RA-GFP, generated by site-directed mutagenesis of the nucleolar localization sequence in NIC4, could not protect from apoptosis triggered by genotoxic stressors. However, transcriptional activity or protection from apoptosis triggered by endoplasmic stress was comparable in cells expressing NIC4_3RA-GFP or NIC4-GFP. Together, the data show that nucleolar localization of NIC4 is critical for the regulation of genomic damage and may be uncoupled from its activities in the nucleoplasm. This study identifies intrinsic features of NIC4 that regulate signaling outcomes activated by the receptor by controlling its spatial localization.

}, issn = {2058-7716}, doi = {10.1038/s41420-020-0242-y}, author = {Saini, Neetu and Sarin, Apurva} } @article {2152, title = {Optimization of Protocols for Detection of De Novo Protein Synthesis in Whole Blood Samples via Azide-Alkyne Cycloaddition.}, journal = {J Proteome Res}, volume = {19}, year = {2020}, month = {2020 Sep 04}, pages = {3856-3866}, abstract = {

Aberrant protein synthesis and protein expression are a hallmark of many conditions ranging from cancer to Alzheimer{\textquoteright}s. Blood-based biomarkers indicative of changes in proteomes have long been held to be potentially useful with respect to disease prognosis and treatment. However, most biomarker efforts have focused on unlabeled plasma proteomics that include nonmyeloid origin proteins with no attempt to dynamically tag acute changes in proteomes. Herein we report a method for evaluating de novo protein synthesis in whole blood liquid biopsies. Using a modification of the "bioorthogonal noncanonical amino acid tagging" (BONCAT) protocol, rodent whole blood samples were incubated with l-azidohomoalanine (AHA) to allow incorporation of this selectively reactive non-natural amino acid within nascent polypeptides. Notably, failure to incubate the blood samples with EDTA prior to implementation of azide-alkyne "click" reactions resulted in the inability to detect probe incorporation. This live-labeling assay was sensitive to inhibition with anisomycin and nascent, tagged polypeptides were localized to a variety of blood cells using FUNCAT. Using labeled rodent blood, these tagged peptides could be consistently identified through standard LC/MS-MS detection of known blood proteins across a variety of experimental conditions. Furthermore, this assay could be expanded to measure de novo protein synthesis in human blood samples. Overall, we present a rapid and convenient de novo protein synthesis assay that can be used with whole blood biopsies that can quantify translational change as well as identify differentially expressed proteins that may be useful for clinical applications.

}, issn = {1535-3907}, doi = {10.1021/acs.jproteome.0c00299}, author = {Bowling, Heather L and Kasper, Amanda and Patole, Chhaya and Venkatasubramani, Janani Priya and Leventer, Sarah Parker and Carmody, Erin and Sharp, Kevin and Berry-Kravis, Elizabeth and Kirshenbaum, Kent and Klann, Eric and Bhattacharya, Aditi} } @article {2061, title = {A perspective on challenges and opportunities in characterizing oral cancer stem cells.}, journal = {Front Biosci (Landmark Ed)}, volume = {25}, year = {2020}, month = {2020 Mar 01}, pages = {1011-1021}, abstract = {

Cancer stem cells (CSCs) or tumor-initiating cells (TICs) represent a minority population of cells in a tumor that can self-renew and re-create the heterogeneity of the entire tumor. Cell lines, patient-derived tumor cells, and patient-derived xenografts have all been used to isolate presumptive CSC populations from different tumor types. Because of their purported roles in tumor recurrence and prognosis, numerous efforts have centered around reliably identifying CSCs using cell surface markers, and in using genomics tools to identify molecular features unique to these cells. In this brief review, we will discuss different markers, CD44, ALDH1, CD271 and others that have used for identifying and isolating CSCs from primary head \& neck and oral squamous cell carcinomas. In particular, we focus on the challenges associated with these experiments as this will be useful to researchers attempting similar isolations. We also discuss some important molecular features gleaned from studying these CSCs such as the expression of stem cell-related markers, loss of cell adhesion and terminal differentiation markers, and the presence of both epithelial and epithelial-to-mesenchymal transition (EMT) features.

}, issn = {1093-4715}, author = {Sadasivam, Subhashini and Subramanian, Ramaswamy} } @article {2150, title = {The Rad53-Spt21 and Tel1 axes couple glucose tolerance to histone dosage and subtelomeric silencing.}, journal = {Nat Commun}, volume = {11}, year = {2020}, month = {2020 08 19}, pages = {4154}, abstract = {

The DNA damage response (DDR) coordinates DNA metabolism with nuclear and non-nuclear processes. The DDR kinase Rad53 controls histone degradation to assist DNA repair. However, Rad53 deficiency causes histone-dependent growth defects in the absence of DNA damage, pointing out unknown physiological functions of the Rad53-histone axis. Here we show that histone dosage control by Rad53 ensures metabolic homeostasis. Under physiological conditions, Rad53 regulates histone levels through inhibitory phosphorylation of the transcription factor Spt21 on Ser276. Rad53-Spt21 mutants display severe glucose dependence, caused by excess histones through two separable mechanisms: dampening of acetyl-coenzyme A-dependent carbon metabolism through histone hyper-acetylation, and Sirtuin-mediated silencing of starvation-induced subtelomeric domains. We further demonstrate that repression of subtelomere silencing by physiological Tel1 and Rpd3 activities coveys tolerance to glucose restriction. Our findings identify DDR mutations, histone imbalances and aberrant subtelomeric chromatin as interconnected causes of glucose dependence, implying that DDR kinases coordinate metabolism and epigenetic changes.

}, keywords = {Acetylation, Ataxia Telangiectasia Mutated Proteins, Cell Cycle Proteins, Checkpoint Kinase 2, DNA Damage, DNA Repair, Gene Silencing, Glucose, Histone Deacetylases, Histones, Intracellular Signaling Peptides and Proteins, Mutation, Phosphorylation, Protein-Serine-Threonine Kinases, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Serine, Telomere, Transcription Factors}, issn = {2041-1723}, doi = {10.1038/s41467-020-17961-4}, author = {Bruhn, Christopher and Ajazi, Arta and Ferrari, Elisa and Lanz, Michael Charles and Batrin, Renaud and Choudhary, Ramveer and Walvekar, Adhish and Laxman, Sunil and Longhese, Maria Pia and Fabre, Emmanuelle and Smolka, Marcus Bustamente and Foiani, Marco} } @article {2204, title = {Resource plasticity-driven carbon-nitrogen budgeting enables specialization and division of labor in a clonal community.}, journal = {Elife}, volume = {9}, year = {2020}, month = {2020 09 02}, abstract = {

Previously, we found that in glucose-limited colonies, metabolic constraints drive cells into groups exhibiting gluconeogenic or glycolytic states. In that study, threshold amounts of trehalose - a limiting, produced carbon-resource, controls the emergence and self-organization of cells exhibiting the glycolytic state, serving as a carbon source that fuels glycolysis (Varahan et al., 2019). We now discover that the plasticity of use of a non-limiting resource, aspartate, controls both resource production and the emergence of heterogeneous cell states, based on differential metabolic budgeting. In gluconeogenic cells, aspartate is a carbon source for trehalose production, while in glycolytic cells using trehalose for carbon, aspartate is predominantly a nitrogen source for nucleotide synthesis. This metabolic plasticity of aspartate enables carbon-nitrogen budgeting, thereby driving the biochemical self-organization of distinct cell states. Through this organization, cells in each state exhibit true division of labor, providing growth/survival advantages for the whole community.

}, issn = {2050-084X}, doi = {10.7554/eLife.57609}, author = {Varahan, Sriram and Sinha, Vaibhhav and Walvekar, Adhish and Krishna, Sandeep and Laxman, Sunil} } @article {2250, title = {Role of connexins in female reproductive system and endometriosis.}, journal = {J Gynecol Obstet Hum Reprod}, volume = {49}, year = {2020}, month = {2020 Jun}, pages = {101705}, abstract = {

Gap junction form channels between the cells and facilitate the function of cellular cross talk. Connexins, the gap junction proteins play an essential role in female reproductive health and its expression anomalies are correlated with female reproductive disorders like polycystic ovarian syndrome, recurrent miscarriage, pre-term birth and endometriosis. Endometriosis is a chronic gynecologic disorder caused by ectopic endometrial lesions growing outside the uterine cavity. Embryonic implantation is adversely affected in case of endometriosis leading to infertility. Endometriosis also interferes with ovulatory functions, reduces fertilization and impaires blastocyst implantation. There lies a lacunae in understanding of the role of gap junctions protein connexins in endometriosis. Therefore, this study discusses the role of connexins in improving female fertility by taming the processes of oogenesis, germ line development, uterine receptivity, placental growth, implantation, decidualization and concludes by focusing the role of connexins in endometriosis.

}, keywords = {Connexins, Decidua, Embryo Implantation, Endometriosis, Endometrium, Female, Gap Junctions, Genitalia, Female, Humans, Infertility, Female, Oogenesis, Ovulation, Pregnancy, Uterus}, issn = {2468-7847}, doi = {10.1016/j.jogoh.2020.101705}, author = {Kaushik, Tripti and Mishra, Rakesh and Singh, Rakesh K and Bajpai, Surabhi} } @article {2106, title = {Structural insights into actin filament recognition by commonly used cellular actin markers.}, journal = {EMBO J}, volume = {39}, year = {2020}, month = {2020 Jul 15}, pages = {e104006}, abstract = {

Cellular studies of filamentous actin (F-actin) processes commonly utilize fluorescent versions of toxins, peptides, and proteins that bind actin. While the choice of these markers has been largely based on availability and ease, there is a severe dearth of structural data for an informed judgment in employing suitable F-actin markers for a particular requirement. Here, we describe the electron cryomicroscopy structures of phalloidin, lifeAct, and utrophin bound to F-actin, providing a comprehensive high-resolution structural comparison of widely used actin markers and their influence towards F-actin. Our results show that phalloidin binding does not induce specific conformational change and lifeAct specifically recognizes closed D-loop conformation, i.e., ADP-Pi or ADP states of F-actin. The structural models aided designing of minimal utrophin and a shorter lifeAct, which can be utilized as F-actin marker. Together, our study provides a structural perspective, where the binding sites of utrophin and lifeAct overlap with majority of actin-binding proteins and thus offering an invaluable resource for researchers in choosing appropriate actin markers and generating new marker variants.

}, issn = {1460-2075}, doi = {10.15252/embj.2019104006}, author = {Kumari, Archana and Kesarwani, Shubham and Javoor, Manjunath G and Vinothkumar, Kutti R and Sirajuddin, Minhajuddin} } @article {2064, title = {Temporal specificity and heterogeneity of Drosophila immune cells.}, journal = {EMBO J}, year = {2020}, month = {2020 Mar 12}, pages = {e104486}, abstract = {

Immune cells provide defense against non-self and have recently been shown to also play key roles in diverse processes such as development, metabolism, and tumor progression. The heterogeneity of Drosophila immune cells (hemocytes) remains an open question. Using bulk RNA sequencing, we find that the hemocytes display distinct features in the embryo, a closed and rapidly developing system, compared to the larva, which is exposed to environmental and metabolic challenges. Through single-cell RNA sequencing, we identify fourteen hemocyte clusters present in unchallenged larvae and associated with distinct processes, e.g., proliferation, phagocytosis, metabolic homeostasis, and humoral response. Finally, we characterize the changes occurring in the hemocyte clusters upon wasp infestation, which triggers the differentiation of a novel hemocyte type, the lamellocyte. This first molecular atlas of hemocytes provides insights and paves the way to study the biology of the Drosophila immune cells in physiological and pathological conditions.

}, issn = {1460-2075}, doi = {10.15252/embj.2020104486}, author = {Cattenoz, Pierre B and Sakr, Rosy and Pavlidaki, Alexia and Delaporte, Claude and Riba, Andrea and Molina, Nacho and Hariharan, Nivedita and Mukherjee, Tina and Giangrande, Angela} } @article {2039, title = {Temporal specificity and heterogeneity of the fly immune cells{\textquoteright} transcriptional landscape}, journal = {The EMBO Journal (in press)}, year = {2020}, abstract = {

Immune cells provide defense against the non-self, however recent data suggest roles well beyond innate immunity, in processes as diverse as development, metabolism and tumor progression. Nevertheless, the heterogeneity of these cells remains an open question. Using bulk RNA sequencing we find that the Drosophila immune cells (hemocytes) display distinct features in the embryo, a closed and rapidly developing system, compared to the larva, which is exposed to environmental and metabolic challenges. Through single cell RNA sequencing we identify fourteen hemocyte clusters present in unchallenged larvae and associated with distinct cellular processes e.g. proliferation, phagocytosis, metabolic homeostasis and humoral response. Finally, we characterize the changes occurring in the hemocyte clusters upon wasp infestation that triggers the differentiation of a novel cell type, the lamellocyte. This first molecular atlas provides precious insights and paves the way to study the biology of the Drosophila immune cells in physiological and pathological conditions.

}, author = {Cattenoz, Pierre B. and Sakr, Rosy and Pavlidaki, Alexia and Delaporte, Claude and Riba, Andrea and Molina, Nacho and Hariharan, Nivedita and Mukherjee, Tina and Giangrande, Angela} } @article {2210, title = {VEGFA Promoter Polymorphisms rs699947 and rs35569394 Are Associated With the Risk of Anterior Cruciate Ligament Ruptures Among Indian Athletes: A Cross-sectional Study.}, journal = {Orthop J Sports Med}, volume = {8}, year = {2020}, month = {2020 Dec}, pages = {2325967120964472}, abstract = {

Background: Associations of genetic variants within certain fibril-forming genes have previously been observed with anterior cruciate ligament (ACL) injuries. Evidence suggests a significant role of angiogenesis-associated cytokines in remodeling the ligament fibril matrix after mechanical loading and maintaining structural and functional integrity of the ligament. Functional polymorphisms within the vascular endothelial growth factor A (VEGFA) gene have emerged as plausible candidates owing to their role in the regulation of angiogenic responses.

Hypothesis: VEGFA promoter polymorphisms rs699947 and rs35569394 are associated with ACL injury risk among athletes.

Study Design: Cross-sectional study; Level of evidence, 3.

Methods: A total of 90 Indian athletes with radiologically confirmed or surgically proven isolated ACL tears and 76 matched-control athletes were selected for the present cross-sectional genetic association study. Oral mouthwash samples were collected from all the case and control athletes and genotyped for VEGFA rs699947 and rs35569394 using the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method.

Results: The A allele (rs699947) was significantly overrepresented in the ACL group (C vs A allele: odds ratio [OR], 1.68 [95\% CI, 1.08-2.60]; = .021) (CC vs CA + AA: OR, 2.69 [95\% CI, 1.37-5.26]; = .004). There was a greater frequency of the AA genotype in the ACL group in comparison with the control group (OR, 3.38 [95\% CI, 1.23-9.28]; = .016) when only male athletes were compared. Likewise, there was a greater frequency of the I allele (rs35569394) in the ACL group (D vs I allele: OR, 1.64 [95\% CI, 1.06-2.55]; = .025) (DD vs ID + II: OR, 2.61 [95\% CI, 1.31-5.21]; = .006). The A-I haplotype was overrepresented in the ACL group compared with the control group (OR, 1.68 [95\% CI, 1.08-2.60]; χ = 5.320; = .021), and both the polymorphisms were found to be in complete linkage disequilibrium ( = 0.929; logarithm of the odds score = 63.74; D{\textquoteright} = 1.0). Female athletes did not show any difference in genotype or allele frequency.

Conclusion: This is the first study to investigate the association of VEGFA promoter polymorphisms in ACL tears among Indian athletes. Increased frequencies of the A allele (rs699947) and I allele (rs35569394) were observed in the ACL group. These results suggest that sequence variants in the VEGF gene are associated with ACL injury risk among athletes. Further research with long-term follow-ups measuring VEGF expression levels during recovery is warranted to establish its role in ACL injuries and healing.

}, issn = {2325-9671}, doi = {10.1177/2325967120964472}, author = {Shukla, Manish and Gupta, Rahul and Pandey, Vivek and Rochette, Jacques and Dhandapany, Perundurai S and Tiwari, Pramod Kumar and Amrathlal, Rabbind Singh} } @article {1645, title = {Altered steady state and activity-dependent de novo protein expression in fragile X syndrome.}, journal = {Nat Commun}, volume = {10}, year = {2019}, month = {2019 Apr 12}, pages = {1710}, abstract = {

Whether fragile X mental retardation protein (FMRP) target mRNAs and neuronal activity contributing to elevated basal neuronal protein synthesis in fragile X syndrome (FXS) is unclear. Our proteomic experiments reveal that the de novo translational profile in FXS model mice is altered at steady state and in response to metabotropic glutamate receptor (mGluR) stimulation, but the proteins expressed differ under these conditions. Several altered proteins, including Hexokinase\ 1 and Ras, also are expressed in the blood of\ FXS model mice and pharmacological treatments previously reported to ameliorate phenotypes modify their abundance in blood. In addition, plasma levels of Hexokinase\ 1 and Ras differ between FXS patients and healthy volunteers. Our data suggest that brain-based de novo proteomics in FXS model mice can be used to find altered expression of proteins in blood that could serve as disease-state biomarkers in individuals with FXS.

}, issn = {2041-1723}, doi = {10.1038/s41467-019-09553-8}, author = {Bowling, Heather and Bhattacharya, Aditi and Zhang, Guoan and Alam, Danyal and Lebowitz, Joseph Z and Bohm-Levine, Nathaniel and Lin, Derek and Singha, Priyangvada and Mamcarz, Maggie and Puckett, Rosemary and Zhou, Lili and Aryal, Sameer and Sharp, Kevin and Kirshenbaum, Kent and Berry-Kravis, Elizabeth and Neubert, Thomas A and Klann, Eric} } @article {1935, title = {Biomaterials for topical and transdermal drug delivery in reconstructive transplantation.}, journal = {Nanomedicine (Lond)}, year = {2019}, month = {2019 Oct 23}, abstract = {

Lifelong systemic immunosuppression remains the biggest challenge in vascularized composite allotransplantation (VCA) due to the adverse effects it causes. Since VCA is a life-enhancing procedure as compared with solid organ transplant which is life-saving;\ one needs to weigh the benefits and risks carefully. Thus, there is a huge unmet clinical need to design biomaterial-based vehicles that can deliver drugs more efficiently, topically and locally to eliminate adverse effects of systemic immune suppression. This review discusses several biomaterial-based systems that have been carefully designed, conceived and attempted to make VCA a more patient compliant approach. Variety of promising preclinical studies has shown the feasibility of the approaches, and clinical trials are required to bridge the gap. Several challenges for the future and new approaches have been discussed.

}, issn = {1748-6963}, doi = {10.2217/nnm-2019-0137}, author = {Dhayani, Ashish and Kalita, Sanjeeb and Mahato, Manohar and Srinath, Preethem and Vemula, Praveen K} } @article {1734, title = {Characterization of new variant human ES line VH9 hESC (INSTEMe001-a): a tool for human stem cell and cancer research.}, journal = {Stem Cell Res}, volume = {37}, year = {2019}, month = {2019 May}, pages = {101444}, abstract = {

Human pluripotent stem cells (hPSCs) acquire changes at the genomic level upon proliferation and differentiation (Peterson and Loring, 2014). Studies from International Stem Cell Initiative and independent laboratories identified a copy number variant (CNV) in hES cell lines displaying a normal karyotype, which provided a selective advantage to hES cells in culture. In our laboratory we have identified variant H9-hESC (derived from H9-hESC) with normal karyotype, pluripotency expression, differentiation profile but with altered traits of high cell survival and low E-CADHERIN expression.

}, issn = {1876-7753}, doi = {10.1016/j.scr.2019.101444}, author = {Arasala, Radhika Rao and Jayaram, Manjunath and Chattai, Jagamohan and Kumarasamy, Thangaraj and Sambasivan, Ramkumar and Rampalli, Shravanti} } @article {1737, title = {Cytoplasmic sequestration of the RhoA effector mDiaphanous1 by Prohibitin2 promotes muscle differentiation.}, journal = {Sci Rep}, volume = {9}, year = {2019}, month = {2019 Jun 05}, pages = {8302}, abstract = {

Muscle differentiation is controlled by adhesion and growth factor-dependent signalling through common effectors that regulate muscle-specific transcriptional programs. Here we report that mDiaphanous1, an effector of adhesion-dependent RhoA-signalling, negatively regulates myogenesis at the level of Myogenin expression. In myotubes, over-expression of mDia1ΔN3, a RhoA-independent mutant, suppresses Myogenin promoter activity and expression. We investigated mDia1-interacting proteins that may counteract mDia1 to permit Myogenin expression and timely differentiation. Using yeast two-hybrid and mass-spectrometric analysis, we report that mDia1 has a stage-specific interactome, including Prohibitin2, MyoD, Akt2, and β-Catenin, along with a number of proteosomal and mitochondrial components. Of these interacting partners, Prohibitin2 colocalises with mDia1 in cytoplasmic punctae in myotubes. We mapped the interacting domains of mDia1 and Phb2, and used interacting (mDia1ΔN3/Phb2 FL or mDia1ΔN3/Phb2-Carboxy) and non-interacting pairs (mDia1H + P/Phb2 FL or mDia1ΔN3/Phb2-Amino) to dissect the functional consequences of this partnership on Myogenin promoter activity. Co-expression of full-length as well as mDia1-interacting domains of Prohibitin2 reverse the anti-myogenic effects of mDia1ΔN3, while non-interacting regions do not. Our results suggest that Prohibitin2 sequesters mDia1, dampens its anti-myogenic activity and fine-tunes RhoA-mDia1 signalling to promote differentiation. Overall, we report that mDia1 is multi-functional signalling effector whose anti-myogenic activity is modulated by a differentiation-dependent interactome.\ The data have been deposited to the ProteomeXchange with identifier PXD012257.

}, issn = {2045-2322}, doi = {10.1038/s41598-019-44749-4}, author = {Saleh, Amena and Subramaniam, Gunasekaran and Raychaudhuri, Swasti and Dhawan, Jyotsna} } @article {1604, title = {Derivation of iPSC lines from two patients with familial Alzheimer{\textquoteright}s disease from India.}, journal = {Stem Cell Res}, volume = {34}, year = {2019}, month = {2019 Jan}, pages = {101370}, abstract = {

The current prevalence of diagnosable dementia in India is 1\% of people over 60 years (~3.7 million people), but is estimated to increase significantly, as ~15\% world{\textquoteright}s aged population (\>65 years) would be resident here by 2020 (Shah et al., 2016). While several mutations that pose a familial risk have been identified, the ethnic background may influence disease susceptibility, clinical presentation and treatment response. In this study, we report a detailed characterization of two representative HiPSC lines from a well-characterized dementia cohort from India. Availability of these lines, and associated molecular and clinical information, would be useful in the detailed exploration of the genomic contribution(s) to AD.

}, issn = {1876-7753}, doi = {10.1016/j.scr.2018.101370}, author = {Najar, Ashaq H and Sneha, K M and Ashok, Aparna and Babu, Swathy and Subramaniam, Anand G and Kannan, Ramkrishnan and Viswanath, Biju and Purushottam, Meera and Varghese, Mathew and Parvez, Suhel and Panicker, Mitradas M and Mukherjee, Odity and Jain, Sanjeev} } @article {1982, title = {Differential Regulation of Translation by FMRP Modulates eEF2 Mediated Response on NMDAR Activity.}, journal = {Front Mol Neurosci}, volume = {12}, year = {2019}, month = {2019}, pages = {97}, abstract = {

SYNGAP1, a Synaptic Ras-GTPase activating protein, regulates synapse maturation during a critical developmental window. Heterozygous mutation in () has been shown to cause Intellectual Disability (ID) in children. Recent studies have provided evidence for altered neuronal protein synthesis in a mouse model of . However, the molecular mechanism behind the same is unclear. Here, we report the reduced expression of a known translation regulator, FMRP, during a specific developmental period in mice. Our results demonstrate that FMRP interacts with and regulates the translation of mRNA. We further show reduced translation leads to decreased FMRP level during development in which results in an increase in translation. These developmental changes are reflected in the altered response of eEF2 phosphorylation downstream of NMDA Receptor (NMDAR)-mediated signaling. In this study, we propose a cross-talk between FMRP and SYNGAP1 mediated signaling which can also explain the compensatory effect of impaired signaling observed in mice.

}, issn = {1662-5099}, doi = {10.3389/fnmol.2019.00097}, author = {Paul, Abhik and Nawalpuri, Bharti and Shah, Devanshi and Sateesh, Shruthi and Muddashetty, Ravi S and Clement, James P} } @article {1740, title = {Dynamic expression of tRNA-derived small RNAs define cellular states.}, journal = {EMBO Rep}, volume = {20}, year = {2019}, month = {2019 Jul}, pages = {e47789}, abstract = {

Transfer RNA (tRNA)-derived small RNAs (tsRNAs) have recently emerged as important regulators of protein translation and shown to have diverse biological functions. However, the underlying cellular and molecular mechanisms of tsRNA function in the context of dynamic cell-state transitions remain unclear. Expression analysis of tsRNAs in distinct heterologous cell and tissue models of stem vs. differentiated states revealed a differentiation-dependent enrichment of 5{\textquoteright}-tsRNAs. We report the identification of a set of 5{\textquoteright}-tsRNAs that is upregulated in differentiating mouse embryonic stem cells (mESCs). Notably, interactome studies with differentially enriched 5{\textquoteright}-tsRNAs revealed a switch in their association with "effector" RNPs and "target" mRNAs in different cell states. We demonstrate that specific 5{\textquoteright}-tsRNAs can preferentially interact with the RNA-binding protein, Igf2bp1, in the RA-induced differentiated state. This association influences the transcript stability and thereby translation of the pluripotency-promoting factor, c-Myc, thus providing a mechanistic basis for how 5{\textquoteright}-tsRNAs can modulate stem cell states in mESCs. Together our study highlights the role of 5{\textquoteright}-tsRNAs in defining distinct cell states.

}, issn = {1469-3178}, doi = {10.15252/embr.201947789}, author = {Krishna, Srikar and Yim, Daniel Gr and Lakshmanan, Vairavan and Tirumalai, Varsha and Koh, Judice Ly and Park, Jung Eun and Cheong, Jit Kong and Low, Joo Leng and Lim, Michelle Js and Sze, Siu Kwan and Shivaprasad, Padubidri and Gulyani, Akash and Raghavan, Srikala and Palakodeti, Dasaradhi and DasGupta, Ramanuj} } @article {1603, title = {Enhancement of the gut barrier integrity by a microbial metabolite through the Nrf2 pathway.}, journal = {Nat Commun}, volume = {10}, year = {2019}, month = {2019 01 09}, pages = {89}, abstract = {

The importance of gut microbiota in human health and pathophysiology is undisputable. Despite the abundance of metagenomics data, the functional dynamics of gut microbiota in human health and disease remain elusive. Urolithin A (UroA), a major microbial metabolite derived from polyphenolics of berries and pomegranate fruits displays anti-inflammatory, anti-oxidative, and anti-ageing activities. Here, we show that UroA and its potent synthetic analogue (UAS03) significantly enhance gut barrier function and inhibit unwarranted inflammation. We demonstrate that UroA and UAS03 exert their barrier functions through activation of aryl hydrocarbon receptor (AhR)- nuclear factor erythroid 2-related factor 2 (Nrf2)-dependent pathways to upregulate epithelial tight junction proteins. Importantly, treatment with these compounds attenuated colitis in pre-clinical models by remedying barrier dysfunction in addition to anti-inflammatory activities. Cumulatively, the results highlight how microbial metabolites provide two-pronged beneficial activities at gut epithelium by enhancing barrier functions and reducing inflammation to protect from colonic diseases.

}, keywords = {Animals, Basic Helix-Loop-Helix Transcription Factors, Caco-2 Cells, Coumarins, Epithelial Cells, Gene Expression Regulation, HT29 Cells, Humans, Intestinal Mucosa, Macrophages, Mice, Mice, Inbred C57BL, Mice, Knockout, NF-E2-Related Factor 2, Receptors, Aryl Hydrocarbon, Specific Pathogen-Free Organisms, Tight Junction Proteins}, issn = {2041-1723}, doi = {10.1038/s41467-018-07859-7}, author = {Singh, Rajbir and Chandrashekharappa, Sandeep and Bodduluri, Sobha R and Baby, Becca V and Hegde, Bindu and Kotla, Niranjan G and Hiwale, Ankita A and Saiyed, Taslimarif and Patel, Paresh and Vijay-Kumar, Matam and Langille, Morgan G I and Douglas, Gavin M and Cheng, Xi and Rouchka, Eric C and Waigel, Sabine J and Dryden, Gerald W and Alatassi, Houda and Zhang, Huang-Ge and Haribabu, Bodduluri and Vemula, Praveen K and Jala, Venkatakrishna R} } @article {1607, title = {Exome sequencing in families with severe mental illness identifies novel and rare variants in genes implicated in Mendelian neuropsychiatric syndromes.}, journal = {Psychiatry Clin Neurosci}, volume = {73}, year = {2019}, month = {2019 Jan}, pages = {11-19}, abstract = {

AIM: Severe mental illnesses (SMI), such as bipolar disorder and schizophrenia, are highly heritable, and have a complex pattern of inheritance. Genome-wide association studies detect a part of the heritability, which can be attributed to common genetic variation. Examination of rare variants with next-generation sequencing may add to the understanding of the genetic architecture of SMI.

METHODS: We analyzed 32 ill subjects from eight multiplex families and 33 healthy individuals using whole-exome sequencing. Prioritized variants were selected by a three-step filtering process, which included: deleteriousness by five in silico algorithms; sharing within families by affected individuals; rarity in South Asian sample estimated using the Exome Aggregation Consortium data; and complete absence of these variants in control individuals from the same gene pool.

RESULTS: We identified 42 rare, non-synonymous deleterious variants (~5 per pedigree) in this study. None of the variants were shared across families, indicating a {\textquoteright}private{\textquoteright} mutational profile. Twenty (47.6\%) of the variant harboring genes were previously reported to contribute to the risk of diverse neuropsychiatric syndromes, nine (21.4\%) of which were of Mendelian inheritance. These included genes carrying novel deleterious variants, such as the GRM1 gene implicated in spinocerebellar ataxia 44 and the NIPBL gene implicated in Cornelia de Lange syndrome.

CONCLUSION: Next-generation sequencing approaches in family-based studies are useful to identify novel and rare variants in genes for complex disorders like SMI. The findings of the study suggest a potential phenotypic burden of rare variants in Mendelian disease genes, indicating pleiotropic effects in the etiology of SMI.

}, keywords = {Bipolar Disorder, Exome, Female, Genetic Predisposition to Disease, Genetic Variation, Genome-Wide Association Study, Humans, Male, Pedigree, Phenotype, Schizophrenia}, issn = {1440-1819}, doi = {10.1111/pcn.12788}, author = {Ganesh, Suhas and Ahmed P, Husayn and Nadella, Ravi K and More, Ravi P and Seshadri, Manasa and Viswanath, Biju and Rao, Mahendra and Jain, Sanjeev and Mukherjee, Odity} } @article {1602, title = {Generation of a set of isogenic, gene-edited iPSC lines homozygous for all main APOE variants and an APOE knock-out line.}, journal = {Stem Cell Res}, volume = {34}, year = {2019}, month = {2019 Jan}, pages = {101349}, abstract = {

Alzheimer{\textquoteright}s disease (AD) is the most frequent neurodegenerative disease amongst the elderly. The SNPs rs429358 and rs7412 in the APOE gene are the most common risk factor for sporadic AD, and there are three different alleles commonly referred to as APOE-ε2, APOE-ε3 and APOE-ε4. Induced pluripotent stem cells (iPSCs) hold great promise to model AD as such cells can be differentiated in vitro to the required cell type. Here we report the use of CRISPR/Cas9 technology employed on iPSCs from a healthy individual with an APOE-ε3/ε4 genotype to obtain isogenic APOE-ε2/ε2, APOE-ε3/ε3, APOE-ε4/ε4 lines as well as an APOE-knock-out line.

}, issn = {1876-7753}, doi = {10.1016/j.scr.2018.11.010}, author = {Schmid, Benjamin and Prehn, Kennie R and Nimsanor, Natakarn and Garcia, Blanca Irene Aldana and Poulsen, Ulla and J{\o}rring, Ida and Rasmussen, Mikkel A and Clausen, Christian and Mau-Holzmann, Ulrike A and Ramakrishna, Sarayu and Muddashetty, Ravi and Steeg, Rachel and Bruce, Kevin and Mackintosh, Peter and Ebneth, Andreas and Holst, Bj{\o}rn and Cabrera-Socorro, Alfredo} } @article {1609, title = {Generation of two iPSC lines with either a heterozygous V717I or a heterozygous KM670/671NL mutation in the APP gene.}, journal = {Stem Cell Res}, volume = {34}, year = {2019}, month = {2019 Jan}, pages = {101368}, abstract = {

Alzheimer{\textquoteright}s disease (AD) is the most common form of dementia, affecting millions of people worldwide. Mutations in the genes PSEN1, PSEN2 or APP are known to cause familial forms of AD with an early age of onset. In this study, specific pathogenic mutations in the APP gene were introduced into an iPSC line from a healthy individual by the use of CRISPR-Cas9. The study resulted in the generation of two new cell lines, one carrying the V717I APP mutation and one with the KM670/671NL APP mutation.

}, issn = {1876-7753}, doi = {10.1016/j.scr.2018.101368}, author = {Frederiksen, Henriette R and Holst, Bj{\o}rn and Ramakrishna, Sarayu and Muddashetty, Ravi and Schmid, Benjamin and Freude, Kristine} } @article {1608, title = {Graft-implanted, enzyme responsive, tacrolimus-eluting hydrogel enables long-term survival of orthotopic porcine limb vascularized composite allografts: A proof of concept study.}, journal = {PLoS One}, volume = {14}, year = {2019}, month = {2019}, pages = {e0210914}, abstract = {

BACKGROUND: Currently, patients receiving vascularized composite allotransplantation (VCA) grafts must take long-term systemic immunosuppressive therapy to prevent immunologic rejection. The morbidity and mortality associated with these medications is the single greatest barrier to more patients being able to receive these life-enhancing transplants. In contrast to solid organs, VCA, exemplified by hand or face transplants, allow visual diagnosis of clinical acute rejection (AR), directed biopsy and targeted graft therapies. Local immunosuppression in VCA could reduce systemic drug exposure and limit adverse effects. This proof of concept study evaluated, in a large animal forelimb VCA model, the efficacy and tolerability of a novel graft-implanted enzyme-responsive, tacrolimus (TAC)-eluting hydrogel platform, in achieving long-term graft survival.

METHODS: Orthotopic forelimb VCA were performed in single haplotype mismatched mini-swine. Controls (n = 2) received no treatment. Two groups received TAC hydrogel: high dose (n = 4, 91 mg TAC) and low dose (n = 4, 49 mg TAC). The goal was to find a dose that was tolerable and resulted in long-term graft survival. Limbs were evaluated for clinical and histopathological signs of AR. TAC levels were measured in serial blood and skin tissue samples. Tolerability of the dose was evaluated by monitoring animal feeding behavior and weight.

RESULTS: Control limbs underwent Banff Grade IV AR by post-operative day six. Low dose TAC hydrogel treatment resulted in long-term graft survival time to onset of Grade IV AR ranging from 56 days to 93 days. High dose TAC hydrogel also resulted in long-term graft survival (24 to 42 days), but was not well tolerated.

CONCLUSION: Graft-implanted TAC-loaded hydrogel delays the onset of Grade IV AR of mismatched porcine forelimb VCA grafts, resulting in long term graft survival and demonstrates dose-dependent tolerability.

}, issn = {1932-6203}, doi = {10.1371/journal.pone.0210914}, author = {Fries, C Anton and Lawson, Shari D and Wang, Lin C and Slaughter, Kai V and Vemula, Praveen K and Dhayani, Ashish and Joshi, Nitin and Karp, Jeffrey M and Rickard, Rory F and Gorantla, Vijay S and Davis, Michael R} } @article {1741, title = {Metabolic constraints drive self-organization of specialized cell groups.}, journal = {Elife}, volume = {8}, year = {2019}, month = {2019 Jun 26}, abstract = {

How phenotypically distinct states in isogenic cell populations appear and stably co-exist remains unresolved. We find that within a mature, clonal yeast colony developing in low glucose, cells arrange into metabolically disparate cell groups. Using this system, we model and experimentally identify metabolic constraints sufficient to drive such self-assembly. Beginning in a uniformly gluconeogenic state, cells exhibiting a contrary, high pentose phosphate pathway activity state, spontaneously appear and proliferate, in a spatially constrained manner. Gluconeogenic cells in the colony produce and provide a resource, which we identify as trehalose. Above threshold concentrations of external trehalose, cells switch to the new metabolic state and proliferate. A self-organized system establishes, where cells in this new state are sustained by trehalose consumption, which thereby restrains other cells in the trehalose producing, gluconeogenic state. Our work suggests simple physico-chemical principles that determine how isogenic cells spontaneously self-organize into structured assemblies in complimentary, specialized states.

}, issn = {2050-084X}, doi = {10.7554/eLife.46735}, author = {Varahan, Sriram and Walvekar, Adhish and Sinha, Vaibhhav and Krishna, Sandeep and Laxman, Sunil} } @article {1843, title = {Molecular basis for metabolite channeling in a ring opening enzyme of the phenylacetate degradation pathway.}, journal = {Nat Commun}, volume = {10}, year = {2019}, month = {2019 Sep 11}, pages = {4127}, abstract = {

Substrate channeling is a mechanism for the internal transfer of hydrophobic, unstable or toxic intermediates from the active site of one enzyme to another. Such transfer has previously been described to be mediated by a hydrophobic tunnel, the use of electrostatic highways or pivoting and by conformational changes. The enzyme PaaZ is used by many bacteria to degrade environmental pollutants. PaaZ is a bifunctional enzyme that catalyzes the ring opening of oxepin-CoA and converts it to 3-oxo-5,6-dehydrosuberyl-CoA. Here we report the structures of PaaZ determined by electron cryomicroscopy with and without bound ligands. The structures reveal that three domain-swapped dimers of the enzyme form a trilobed structure. A combination of small-angle X-ray scattering (SAXS), computational studies, mutagenesis and microbial growth experiments suggests that the key intermediate is transferred from one active site to the other by a mechanism of electrostatic pivoting of the CoA moiety, mediated by a set of conserved positively charged residues.

}, issn = {2041-1723}, doi = {10.1038/s41467-019-11931-1}, author = {Sathyanarayanan, Nitish and Cannone, Giuseppe and Gakhar, Lokesh and Katagihallimath, Nainesh and Sowdhamini, Ramanathan and Ramaswamy, Subramanian and Vinothkumar, Kutti R} } @article {1644, title = {Myocardin ablation in a cardiac-renal rat model.}, journal = {Sci Rep}, volume = {9}, year = {2019}, month = {2019 Apr 10}, pages = {5872}, abstract = {

Cardiorenal syndrome is defined by primary heart failure conditions influencing or leading to renal injury or dysfunction. Dilated cardiomyopathy (DCM) is a major co-existing form of heart failure (HF) with renal diseases. Myocardin (MYOCD), a cardiac-specific co-activator of serum response factor (SRF), is increased in DCM porcine and patient cardiac tissues and plays a crucial role in the pathophysiology of DCM. Inhibiting the increased MYOCD has shown to be partially rescuing the DCM phenotype in porcine model. However, expression levels of MYOCD in the cardiac tissues of the cardiorenal syndromic patients and the effect of inhibiting MYOCD in a cardiorenal syndrome model remains to be explored. Here, we analyzed the expression levels of MYOCD in the DCM patients with and without renal diseases. We also explored, whether cardiac specific silencing of MYOCD expression could ameliorate the cardiac remodeling and improve cardiac function in a renal artery ligated rat model (RAL). We observed an increase in MYOCD levels in the endomyocardial biopsies of DCM patients associated with renal failure compared to DCM alone. Silencing of MYOCD in RAL rats by a cardiac homing peptide conjugated MYOCD siRNA resulted in attenuation of cardiac hypertrophy, fibrosis and restoration of the left ventricular functions. Our data suggest hyper-activation of MYOCD in the pathogenesis of the cardiorenal failure cases. Also, MYOCD silencing showed beneficial effects by rescuing cardiac hypertrophy, fibrosis, size and function in a cardiorenal rat model.

}, issn = {2045-2322}, doi = {10.1038/s41598-019-42009-z}, author = {Mittal, Anupam and Rana, Santanu and Sharma, Rajni and Kumar, Akhilesh and Prasad, Rishikesh and Raut, Satish K and Sarkar, Sagartirtha and Saikia, Uma Nahar and Bahl, Ajay and Dhandapany, Perundurai S and Khullar, Madhu} } @article {1617, title = {Natural monomers: A mine for functional and sustainable materials {\textendash} Occurrence, chemical modification and polymerization}, journal = {Progress in Polymer Science}, volume = {92}, year = {2019}, abstract = {

Owing to the natural abundance, variety of structural features, and highly specific functions, natural monomers render themselves as potential candidates for production of high performance functional polymers. The emerging concept of the biorefinery and development of new biosynthetic routes to synthesize a versatile and broad spectrum of natural monomers and polymers continues to gain momentum. The production of high quality polymers from renewable feedstocks requires innovative chemical modifications and catalytic transformations to achieve higher yields in an efficient manner. A fresh look into monomers available from natural resources such as terpenes, rosin, glycerol, furans, tannins, suberin, their derivatives and miscellaneous monomers may inspire future applications with impactful biobased materials. There are also many areas that require urgent discussion and review pertaining to recent developments in the field; this includes monomer sources that give molecules having special structural features. In particular, cardanol, a naturally occurring low-molecular-weight compound is unique as it contains a phenolic head group and a hydrocarbon chain with different degrees of unsaturation. This molecule possesses functional groups that are amenable for classical chemical modification, which is instrumental in developing a wide range of functional monomers and polymers. A large number of soft and hard materials have been developed from cardanol-based monomers. During the past, a large number of industrial grade materials have been developed from plant-based monomers, including development from microbial and fermentation processes (i.e. lactic acid). This review provides a comprehensive study and survey on recent developments on monomers and polymers derived from urushiol and cardanol based monomers and polymers, vegetable oil-based monomers and polymers, microbially produced monomers and polymers. These all represent emerging fronts giving a vast scope while highlighting important potential material and reagent opportunities.

}, url = {https://www.sciencedirect.com/science/article/pii/S0079670018300273}, author = {John, G., and Nagarajan, S., and Vemula, P. K., and Silverman, J.R., and Pillai, C. K. S.} } @article {1984, title = {Novel Series of Methyl 3-(Substituted Benzoyl)-7-Substituted-2-Phenylindolizine-1-Carboxylates as Promising Anti-Inflammatory Agents: Molecular Modeling Studies.}, journal = {Biomolecules}, volume = {9}, year = {2019}, month = {2019 Oct 28}, abstract = {

The cyclooxygenase-2 (COX-2) enzyme is considered to be an important target for developing novel anti-inflammatory agents. Selective COX-2 inhibitors offer the advantage of lower adverse effects that are commonly associated with non-selective COX inhibitors. In this work, a novel series of methyl 3-(substituted benzoyl)-7-substituted-2-phenylindolizine-1-carboxylates was synthesized and evaluated for COX-2 inhibitory activity. Compound was identified as the most active compound of the series with an IC of 6.71 M, which is comparable to the IC of indomethacin, a marketed non-steroidal anti-inflammatory drug (NSAID). Molecular modeling and crystallographic studies were conducted to further characterize the compounds and gain better understanding of the binding interactions between the compounds and the residues at the active site of the COX-2 enzyme. The pharmacokinetic properties and potential toxic effects were predicted for all the synthesized compounds, which indicated good drug-like properties. Thus, these synthesized compounds can be considered as potential lead compounds for developing effective anti-inflammatory therapeutic agents.

}, issn = {2218-273X}, doi = {10.3390/biom9110661}, author = {Venugopala, Katharigatta N and Al-Attraqchi, Omar H A and Tratrat, Christophe and Nayak, Susanta K and Morsy, Mohamed A and Aldhubiab, Bandar E and Attimarad, Mahesh and Nair, Anroop B and Sreeharsha, Nagaraja and Venugopala, Rashmi and Haroun, Michelyne and Girish, Meravanige B and Chandrashekharappa, Sandeep and Alwassil, Osama I and Odhav, Bharti} } @article {1614, title = {Post-transcriptional regulation in planarian stem cells.}, journal = {Semin Cell Dev Biol}, volume = {87}, year = {2019}, month = {2019 Mar}, pages = {69-78}, abstract = {

Planarians are known for their immense regenerative abilities. A pluripotent stem cell population provides the cellular source for this process, as well as for the homeostatic cell turnover of the animals. These stem cells, known as neoblasts, present striking similarities at the morphological and molecular level to germ cells, but however, give rise to somatic tissue. Many RNA binding proteins known to be important for germ cell biology are also required for neoblast function, highlighting the importance of post-transcriptional regulation for stem cell control. Many of its aspects, including alternative splicing, alternative polyadenylation, translational control and mRNA deadenylation, as well as small RNAs such as microRNAs and piRNA are critical for stem cells. Their inhibition often abrogates both regeneration and cell turnover, resulting in lethality. Some of aspects of post-transcriptional regulation are conserved from planarian to mammalian stem cells.

}, issn = {1096-3634}, doi = {10.1016/j.semcdb.2018.05.013}, author = {Krishna, Srikar and Palakodeti, Dasaradhi and Solana, Jordi} } @article {1838, title = {A question of lineage.}, journal = {Elife}, volume = {8}, year = {2019}, month = {2019 May 07}, abstract = {

In the ventral nerve cord of fruit flies, neurons from the same hemilineage use the same neurotransmitter.

}, issn = {2050-084X}, doi = {10.7554/eLife.47162}, author = {Sen, Sonia} } @article {1742, title = {The resolution revolution reaches India.}, journal = {Biophys Rev}, year = {2019}, month = {2019 Jul 02}, issn = {1867-2450}, doi = {10.1007/s12551-019-00558-7}, author = {Subramanian, Ramaswamy and Mayor, Satyajit and Vinothkumar, Kutti R} } @article {1844, title = {Serotonin is essential for eye regeneration in planaria Schmidtea mediterranea.}, journal = {FEBS Lett}, year = {2019}, month = {2019 Sep 17}, abstract = {

Planaria is an ideal system to study factors involved in regeneration and tissue homeostasis. Little is known about the role of metabolites and small molecules in stem cell maintenance and lineage specification in planarians. Using liquid chromatography and mass spectrometry (LC-MS)-based quantitative metabolomics, we determined the relative levels of metabolites in stem cells, progenitors, and differentiated cells of the planarian Schmidtea mediterranea. Tryptophan and its metabolic product serotonin are significantly enriched in stem cells and progenitor population. Serotonin biosynthesis in these cells is brought about by a non-canonical enzyme, phenylalanine hydroxylase (PAH). Knockdown of Smed-pah leads to complete disappearance of eyes in regenerating planaria, while exogenous supply of serotonin and its precursor rescue the eyeless phenotype. Our results demonstrate a key role for serotonin in eye regeneration.

}, issn = {1873-3468}, doi = {10.1002/1873-3468.13607}, author = {Sarkar, Arunabha and Mukundan, Namita and Sowndarya, Sai and Dubey, Vinay Kumar and Babu, Rosana and Lakshmanan, Vairavan and Rangiah, Kannan and Panicker, Mitradas M and Palakodeti, Dasaradhi and Subramanian, Sabarinath Peruvemba and Ramaswamy, Subramanian} } @article {1983, title = {Structural and functional characterization of CMP-N-acetylneuraminate synthetase from Vibrio cholerae.}, journal = {Acta Crystallogr D Struct Biol}, volume = {75}, year = {2019}, month = {2019 Jun 01}, pages = {564-577}, abstract = {

Several pathogenic bacteria utilize sialic acid, including host-derived N-acetylneuraminic acid (Neu5Ac), in at least two ways: they use it as a nutrient source and as a host-evasion strategy by coating themselves with Neu5Ac. Given the significant role of sialic acid in pathogenesis and host-gut colonization by various pathogenic bacteria, including Neisseria meningitidis, Haemophilus influenzae, Pasteurella multocida and Vibrio cholerae, several enzymes of the sialic acid catabolic, biosynthetic and incorporation pathways are considered to be potential drug targets. In this work, findings on the structural and functional characterization of CMP-N-acetylneuraminate synthetase (CMAS), a key enzyme in the incorporation pathway, from Vibrio cholerae are reported. CMAS catalyzes the synthesis of CMP-sialic acid by utilizing CTP and sialic acid. Crystal structures of the apo and the CDP-bound forms of the enzyme were determined, which allowed the identification of the metal cofactor Mg in the active site interacting with CDP and the invariant Asp215 residue. While open and closed structural forms of the enzyme from eukaryotic and other bacterial species have already been characterized, a partially closed structure of V.\ cholerae CMAS (VcCMAS) observed upon CDP binding, representing an intermediate state, is reported here. The kinetic data suggest that VcCMAS is capable of activating the two most common sialic acid derivatives, Neu5Ac and Neu5Gc. Amino-acid sequence and structural comparison of the active site of VcCMAS with those of eukaryotic and other bacterial counterparts reveal a diverse hydrophobic pocket that interacts with the C5 substituents of sialic acid. Analyses of the thermodynamic signatures obtained from the binding of the nucleotide (CTP) and the product (CMP-sialic acid) to VcCMAS provide fundamental information on the energetics of the binding process.

}, keywords = {Amino Acid Sequence, Bacterial Proteins, Binding Sites, Catalytic Domain, Crystallization, Crystallography, X-Ray, Cytidine Diphosphate, Cytidine Monophosphate N-Acetylneuraminic Acid, Cytidine Triphosphate, N-Acylneuraminate Cytidylyltransferase, Protein Interaction Domains and Motifs, Protein Structure, Quaternary, Sialic Acids, Vibrio cholerae}, issn = {2059-7983}, doi = {10.1107/S2059798319006831}, author = {Bose, Sucharita and Purkait, Debayan and Joseph, Deepthi and Nayak, Vinod and Subramanian, Ramaswamy} } @article {1840, title = {Structure-Guided Synthesis and Evaluation of Small-Molecule Inhibitors Targeting Protein-Protein Interactions of BRCA1 tBRCT Domain.}, journal = {ChemMedChem}, volume = {14}, year = {2019}, month = {2019 Sep 18}, pages = {1620-1632}, abstract = {

The tandem BRCT domains (tBRCT) of BRCA1 engage phosphoserine-containing motifs in target proteins to propagate intracellular signals initiated by DNA damage, thereby controlling cell cycle arrest and DNA repair. Recently, we identified Bractoppin, the first small-molecule inhibitor of the BRCA1 tBRCT domain, which selectively interrupts BRCA1-mediated cellular responses evoked by DNA damage. Here, we combine structure-guided chemical elaboration, protein mutagenesis and cellular assays to define the structural features responsible for Bractoppin{\textquoteright}s activity. Bractoppin fails to bind mutant forms of BRCA1 tBRCT bearing K1702A, a key residue mediating phosphopeptide recognition, or F1662R or L1701K that adjoin the pSer-recognition site. However, the M1775R mutation, which engages the Phe residue in the consensus phosphopeptide motif pSer-X-X-Phe, does not affect Bractoppin binding, confirming a binding mode distinct from the substrate phosphopeptide binding. We explored these structural features through structure-guided chemical elaboration and characterized structure-activity relationships (SARs) in biochemical assays. Two analogues, CCBT2088 and CCBT2103 were effective in abrogating BRCA1 foci formation and inhibiting G2 arrest induced by irradiation of cells. Collectively, our findings reveal structural features underlying the activity of a novel inhibitor of phosphopeptide recognition by the BRCA1 tBRCT domain, providing fresh insights to guide the development of inhibitors that target protein-protein interactions.

}, issn = {1860-7187}, doi = {10.1002/cmdc.201900300}, author = {Kurdekar, Vadiraj and Giridharan, Saranya and Subbarao, Jasti and Nijaguna, Mamatha B and Periasamy, Jayaprakash and Boggaram, Sanjana and Shivange, Amol V and Sadasivam, Gayathri and Padigaru, Muralidhara and Potluri, Vijay and Venkitaraman, Ashok R and Bharatham, Kavitha} } @article {1738, title = {Sustained correction of associative learning deficits after brief, early treatment in a rat model of Fragile X Syndrome.}, journal = {Sci Transl Med}, volume = {11}, year = {2019}, month = {2019 May 29}, abstract = {

Fragile X Syndrome (FXS) is one of the most common monogenic forms of autism and intellectual disability. Preclinical studies in animal models have highlighted the potential of pharmaceutical intervention strategies for alleviating the symptoms of FXS. However, whether treatment strategies can be tailored to developmental time windows that define the emergence of particular phenotypes is unknown. Similarly, whether a brief, early intervention can have long-lasting beneficial effects, even after treatment cessation, is also unknown. To address these questions, we first examined the developmental profile for the acquisition of associative learning in a rat model of FXS. Associative memory was tested using a range of behavioral paradigms that rely on an animal{\textquoteright}s innate tendency to explore novelty. knockout (KO) rats showed a developmental delay in their acquisition of object-place recognition and did not demonstrate object-place-context recognition paradigm at any age tested (up to 23 weeks of age). Treatment of KO rats with lovastatin between 5 and 9 weeks of age, during the normal developmental period that this associative memory capability is established, prevents the emergence of deficits but has no effect in wild-type animals. Moreover, we observe no regression of cognitive performance in the FXS rats over several months after treatment. This restoration of the normal developmental trajectory of cognitive function is associated with the sustained rescue of both synaptic plasticity and altered protein synthesis. The findings provide proof of concept that the impaired emergence of the cognitive repertoire in neurodevelopmental disorders may be prevented by brief, early pharmacological intervention.

}, issn = {1946-6242}, doi = {10.1126/scitranslmed.aao0498}, author = {Asiminas, Antonis and Jackson, Adam D and Louros, Susana R and Till, Sally M and Spano, Teresa and Dando, Owen and Bear, Mark F and Chattarji, Sumantra and Hardingham, Giles E and Osterweil, Emily K and Wyllie, David J A and Wood, Emma R and Kind, Peter C} } @article {1986, title = {Sustained Secretion of the Antimicrobial Peptide S100A7 Is Dependent on the Downregulation of Caspase-8.}, journal = {Cell Rep}, volume = {29}, year = {2019}, month = {2019 Nov 26}, pages = {2546-2555.e4}, abstract = {

Antimicrobial peptides (AMPs) are the body{\textquoteright}s natural innate immune defense against a spectrum of pathogens and can also modulate cell proliferation, chemotaxis, angiogenesis, wound healing, and immune cell activity. Harnessing these diverse functions for prophylactic use is contingent upon understanding the regulatory mechanisms governing their unconventional secretion from cells. Analysis of the secretion of S100A7 (Psoriasin), an abundant AMP stored in differentiated keratinocytes of the skin, has revealed an unexpected biphasic secretory response to bacterial exposure. The core components regulating S100A7 secretion are NFκB/p38MAPK, caspase-1, and interleukin (IL)-1α. The initial activation of this core machinery is mediated by Toll-like receptor signaling, whereas the chronic response is mediated by Caspase-8 downregulation. Interestingly, there is a concomitant downregulation of Caspase-8 in inflammatory skin diseases wherein S100A7 is constitutively released. These results highlight the potential of targeting these components to control the release of AMPs from the skin in both homeostatic and disease conditions.

}, issn = {2211-1247}, doi = {10.1016/j.celrep.2019.10.090}, author = {Bhatt, Tanay and Bhosale, Aishwarya and Bajantri, Bhavya and Mathapathi, Mruthyunjaya Swamy and Rizvi, Abrar and Scita, Giorgio and Majumdar, Amitabha and Jamora, Colin} } @article {1959, title = {Sustained Secretion of the Antimicrobial Peptide S100A7 Is Dependent on the Downregulation of Caspase-8.}, year = {2019}, month = {09/2019}, abstract = {

Antimicrobial peptides (AMPs) are the body{\textquoteright}s natural innate immune defense against a spectrum of pathogens and can also modulate cell proliferation, chemotaxis, angiogenesis, wound healing, and immune cell activity. Harnessing these diverse functions for prophylactic use is contingent upon understanding the regulatory mechanisms governing their unconventional secretion from cells. Analysis of the secretion of S100A7 (Psoriasin), an abundant AMP stored in differentiated keratinocytes of the skin, has revealed an unexpected biphasic secretory response to bacterial exposure. The core components regulating S100A7 secretion are NFκB/p38MAPK, caspase-1, and interleukin (IL)-1α. The initial activation of this core machinery is mediated by Toll-like receptor signaling, whereas the chronic response is mediated by Caspase-8 downregulation. Interestingly, there is a concomitant downregulation of Caspase-8 in inflammatory skin diseases wherein S100A7 is constitutively released. These results highlight the potential of targeting these components to control the release of AMPs from the skin in both homeostatic and disease conditions.

Copyright {\textcopyright} 2019 The Author(s). Published by Elsevier Inc. All rights reserved.

}, author = {Bhatt,T and Bhosale,A and Bajantri,B and Mathapathi,MS and Rizvi,A and Scita,G and Majumdar,A and Jamora,C} } @article {1743, title = {A tRNA modification balances carbon and nitrogen metabolism by regulating phosphate homeostasis.}, journal = {Elife}, volume = {8}, year = {2019}, month = {2019 Jul 01}, abstract = {

Cells must appropriately sense and integrate multiple metabolic resources to commit to proliferation. Here, we report that cells regulate carbon and nitrogen metabolic homeostasis through tRNA U-thiolation. Despite amino acid sufficiency, tRNA-thiolation deficient cells appear amino acid starved. In these cells, carbon flux towards nucleotide synthesis decreases, and trehalose synthesis increases, resulting in a starvation-like metabolic signature. Thiolation mutants have only minor translation defects. However, in these cells phosphate homeostasis genes are strongly down-regulated, resulting in an effectively phosphate-limited state. Reduced phosphate enforces a metabolic switch, where glucose-6-phosphate is routed towards storage carbohydrates. Notably, trehalose synthesis, which releases phosphate and thereby restores phosphate availability, is central to this metabolic rewiring. Thus, cells use thiolated tRNAs to perceive amino acid sufficiency, balance carbon and amino acid metabolic flux and grow optimally, by controlling phosphate availability. These results further biochemically explain how phosphate availability determines a switch to a {\textquoteright}starvation-state{\textquoteright}.

}, issn = {2050-084X}, doi = {10.7554/eLife.44795}, author = {Gupta, Ritu and Walvekar, Adhish and Liang, Shun and Rashida, Zeenat and Shah, Premal and Laxman, Sunil} } @article {1582, title = {Urolithin A, a Novel Natural Compound to Target PI3K/AKT/mTOR Pathway in Pancreatic Cancer.}, journal = {Mol Cancer Ther}, volume = {18}, year = {2019}, month = {2019 Feb}, pages = {301-311}, abstract = {

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy and is highly resistant to standard treatment regimens. Targeted therapies against , a mutation present in an overwhelming majority of PDAC cases, have been largely ineffective. However, inhibition of downstream components in the KRAS signaling cascade provides promising therapeutic targets in the management of PDAC and warrants further exploration. Here, we investigated Urolithin A (Uro A), a novel natural compound derived from pomegranates, which targets numerous kinases downstream of KRAS, in particular the PI3K/AKT/mTOR signaling pathways. We showed that treatment of PDAC cells with Uro A blocked the phosphorylation of AKT and p70S6K successfully inhibited the growth of tumor xenografts, and increased overall survival of Ptf1a;LSL-Kras;Tgfbr2 (PKT) mice compared with vehicle or gemcitabine therapy alone. Histologic evaluation of these Uro A-treated tumor samples confirmed mechanistic actions of Uro A via decreased phosphorylation of AKT and p70S6K, reduced proliferation, and increased cellular apoptosis in both xenograft and PKT mouse models. In addition, Uro A treatment reprogrammed the tumor microenvironment, as evidenced by reduced levels of infiltrating immunosuppressive cell populations such as myeloid-derived suppressor cells, tumor-associated macrophages, and regulatory T cells. Overall, this work provides convincing preclinical evidence for the utility of Uro A as a therapeutic agent in PDAC through suppression of the PI3K/AKT/mTOR pathway.

}, issn = {1538-8514}, doi = {10.1158/1535-7163.MCT-18-0464}, author = {Totiger, Tulasigeri M and Srinivasan, Supriya and Jala, Venkatakrishna R and Lamichhane, Purushottam and Dosch, Austin R and Gaidarski, Alexander A and Joshi, Chandrashekhar and Rangappa, Shobith and Castellanos, Jason and Vemula, Praveen Kumar and Chen, Xi and Kwon, Deukwoo and Kashikar, Nilesh and VanSaun, Michael and Merchant, Nipun B and Nagathihalli, Nagaraj S} } @article {1192, title = {Automation aided optimization of cloning, expression and purification of enzymes of the bacterial sialic acid catabolic and sialylation pathways enzymes for structural studies.}, journal = {Microb Biotechnol}, volume = {11}, year = {2018}, month = {2018 Mar}, pages = {420-428}, abstract = {

The process of obtaining a well-expressing, soluble and correctly folded constructs can be made easier and quicker by automating the optimization of cloning, expression and purification. While there are many semiautomated pipelines available for cloning, expression and purification, there is hardly any pipeline that involves complete automation. Here, we achieve complete automation of all the steps involved in cloning and in\ vivo expression screening. This is demonstrated using 18 genes involved in sialic acid catabolism and the surface sialylation pathway. Our main objective was to clone these genes into a His-tagged Gateway vector, followed by their small-scale expression optimization in\ vivo. The constructs that showed best soluble expression were then selected for purification studies and scaled up for crystallization studies. Our technique allowed us to quickly find conditions for producing significant quantities of soluble proteins in Escherichia coli, their large-scale purification and successful crystallization of a number of these proteins. The method can be implemented in other cases where one needs to screen a large number of constructs, clones and expression vectors for successful recombinant production of functional proteins.

}, issn = {1751-7915}, doi = {10.1111/1751-7915.13041}, author = {Bairy, Sneha and Gopalan, Lakshmi Narayanan and Setty, Thanuja Gangi and Srinivasachari, Sathya and Manjunath, Lavanyaa and Kumar, Jay Prakash and Guntupalli, Sai R and Bose, Sucharita and Nayak, Vinod and Ghosh, Swagatha and Sathyanarayanan, Nitish and Caing-Carlsson, Rhawnie and Wahlgren, Weixiao Yuan and Friemann, Rosmarie and Ramaswamy, S and Neerathilingam, Muniasamy} } @article {1587, title = {Bioresponsive drug delivery systems in intestinal inflammation: State-of-the-art and future perspectives.}, journal = {Adv Drug Deliv Rev}, year = {2018}, month = {2018 Jun 29}, abstract = {

Oral colon-specific delivery systems emerged as the main therapeutic cargos by making a significant impact in the field of modern medicine for local drug delivery in intestinal inflammation. The site-specific delivery of therapeutics (aminosalicylates, glucocorticoids, biologics) to the ulcerative mucus tissue can provide prominent advantages in mucosal healing (MH). Attaining gut mucosal healing and anti-fibrosis are main treatment outcomes in inflammatory bowel disease (IBD). The pharmaceutical strategies that are commonly used to achieve a colon-specific drug delivery system include time, pH-dependent polymer coating, prodrug, colonic microbiota-activated delivery systems and a combination of these approaches. Amongst the different approaches reported, the use of biodegradable polysaccharide coated systems holds great promise in delivering drugs to the ulcerative regions. The present review focuses on major physiological gastro-intestinal tract challenges involved in altering the pharmacokinetics of delivery systems, pathophysiology of MH and fibrosis, reported drug-polysaccharide cargos and focusing on conventional to advanced disease responsive delivery strategies, highlighting their limitations and future perspectives in intestinal inflammation therapy.

}, issn = {1872-8294}, doi = {10.1016/j.addr.2018.06.021}, author = {Kotla, Niranjan G and Rana, Shubhasmin and Sivaraman, Gandhi and Sunnapu, Omprakash and Vemula, Praveen K and Pandit, Abhay and Rochev, Yury} } @article {1217, title = {Chemically diverse small molecule fluorescent chemosensors for copper ion}, journal = {Coordination Chemistry Reviews}, volume = {357}, year = {2018}, pages = {50 - 104}, keywords = {Chemosensors, Fluorescence, Live cell imaging, Small molecules, Turn-off, Turn-on}, issn = {0010-8545}, doi = {https://doi.org/10.1016/j.ccr.2017.11.020}, url = {http://www.sciencedirect.com/science/article/pii/S0010854517306537}, author = {Gandhi Sivaraman and Murugan Iniya and Thangaraj Anand and Niranjan G. Kotla and Omprakash Sunnapu and Subramanian Singaravadivel and Akash Gulyani and Duraisamy Chellappa} } @article {1184, title = {Decreased expression of cell adhesion genes in cancer stem-like cells isolated from primary oral squamous cell carcinomas.}, journal = {Tumour Biol}, volume = {40}, year = {2018}, month = {2018 May}, pages = {1010428318780859}, abstract = {

The goal of this study was to isolate cancer stem-like cells marked by high expression of CD44, a putative cancer stem cell marker, from primary oral squamous cell carcinomas and identify distinctive gene expression patterns in these cells. From 1 October 2013 to 4 September 2015, 76 stage III-IV primary oral squamous cell carcinoma of the gingivobuccal sulcus were resected. In all, 13 tumours were analysed by immunohistochemistry to visualise CD44-expressing cells. Expression of CD44 within The Cancer Genome Atlas-Head and Neck Squamous Cell Carcinoma RNA-sequencing data was also assessed. Seventy resected tumours were dissociated into single cells and stained with antibodies to CD44 as well as CD45 and CD31 (together referred as Lineage/Lin). From 45 of these, CD44Lin and CD44Lin subpopulations were successfully isolated using fluorescence-activated cell sorting, and good-quality RNA was obtained from 14 such sorted pairs. Libraries from five pairs were sequenced and the results analysed using bioinformatics tools. Reverse transcription quantitative polymerase chain reaction was performed to experimentally validate the differential expression of selected candidate genes identified from the transcriptome sequencing in the same 5 and an additional 9 tumours. CD44 was expressed on the surface of poorly differentiated tumour cells, and within the The Cancer Genome Atlas-Head and Neck Squamous Cell Carcinoma samples, its messenger RNA levels were higher in tumours compared to normal. Transcriptomics revealed that 102 genes were upregulated and 85 genes were downregulated in CD44Lin compared to CD44Lin cells in at least 3 of the 5 tumours sequenced. The upregulated genes included those involved in immune regulation, while the downregulated genes were enriched for genes involved in cell adhesion. Decreased expression of PCDH18, MGP, SPARCL1 and KRTDAP was confirmed by reverse transcription quantitative polymerase chain reaction. Lower expression of the cell-cell adhesion molecule PCDH18 correlated with poorer overall survival in the The Cancer Genome Atlas-Head and Neck Squamous Cell Carcinoma data highlighting it as a potential negative prognostic factor in this cancer.

}, issn = {1423-0380}, doi = {10.1177/1010428318780859}, author = {Mishra, Amrendra and Sriram, Harshini and Chandarana, Pinal and Tanavde, Vivek and Kumar, Rekha V and Gopinath, Ashok and Govindarajan, Raman and Ramaswamy, S and Sadasivam, Subhashini} } @article {1149, title = {Discovery biology of neuropsychiatric syndromes (DBNS): a center for integrating clinical medicine and basic science.}, journal = {BMC Psychiatry}, volume = {18}, year = {2018}, month = {2018 Apr 18}, pages = {106}, abstract = {

BACKGROUND: There is emerging evidence that there are shared genetic, environmental and developmental risk factors in psychiatry, that cut across traditional diagnostic boundaries. With this background, the Discovery biology of neuropsychiatric syndromes (DBNS) proposes to recruit patients from five different syndromes (schizophrenia, bipolar disorder, obsessive compulsive disorder, Alzheimer{\textquoteright}s dementia and substance use disorders), identify those with multiple affected relatives, and invite these families to participate in this study. The families will be assessed: 1) To compare neuro-endophenotype measures between patients, first degree relatives (FDR) and healthy controls., 2) To identify cellular phenotypes which differentiate the groups., 3) To examine the longitudinal course of neuro-endophenotype measures., 4) To identify measures which correlate with outcome, and 5) To create a unified digital database and biorepository.

METHODS: The identification of the index participants will occur at well-established specialty clinics. The selected individuals will have a strong family history (with at least another affected FDR) of mental illness. We will also recruit healthy controls without family history of such illness. All recruited individuals (N = 4500) will undergo brief clinical assessments and a blood sample will be drawn for isolation of DNA and peripheral blood mononuclear cells (PBMCs). From among this set, a subset of 1500 individuals (300 families and 300 controls) will be assessed on several additional assessments [detailed clinical assessments, endophenotype measures (neuroimaging- structural and functional, neuropsychology, psychophysics-electroencephalography, functional near infrared spectroscopy, eye movement tracking)], with the intention of conducting repeated measurements every alternate year. PBMCs from this set will be used to generate lymphoblastoid cell lines, and a subset of these would be converted to induced pluripotent stem cell lines and also undergo whole exome sequencing.

DISCUSSION: We hope to identify unique and overlapping brain endophenotypes for major psychiatric syndromes. In a proportion of subjects, we expect these neuro-endophenotypes to progress over time and to predict treatment outcome. Similarly, cellular assays could differentiate cell lines derived from such groups. The repository of biomaterials as well as digital datasets of clinical parameters, will serve as a valuable resource for the broader scientific community who wish to address research questions in the area.

}, issn = {1471-244X}, doi = {10.1186/s12888-018-1674-2}, author = {Viswanath, Biju and Rao, Naren P and Narayanaswamy, Janardhanan C and Sivakumar, Palanimuthu T and Kandasamy, Arun and Kesavan, Muralidharan and Mehta, Urvakhsh Meherwan and Venkatasubramanian, Ganesan and John, John P and Mukherjee, Odity and Purushottam, Meera and Kannan, Ramakrishnan and Mehta, Bhupesh and Kandavel, Thennarasu and Binukumar, B and Saini, Jitender and Jayarajan, Deepak and Shyamsundar, A and Moirangthem, Sydney and Vijay Kumar, K G and Thirthalli, Jagadisha and Chandra, Prabha S and Gangadhar, Bangalore N and Murthy, Pratima and Panicker, Mitradas M and Bhalla, Upinder S and Chattarji, Sumantra and Benegal, Vivek and Varghese, Mathew and Reddy, Janardhan Y C and Raghu, Padinjat and Rao, Mahendra and Jain, Sanjeev} } @article {1212, title = {Distinguishing States of Arrest: Genome-Wide Descriptions of Cellular Quiescence Using ChIP-Seq and RNA-Seq Analysis.}, journal = {Methods Mol Biol}, volume = {1686}, year = {2018}, month = {2018}, pages = {215-239}, abstract = {

Regenerative potential in adult stem cells is closely associated with the establishment of-and exit from-a temporary state of quiescence. Emerging evidence not only provides a rationale for the link between lineage determination programs and cell cycle regulation but also highlights the understanding of quiescence as an actively maintained cellular program, encompassing networks and mechanisms beyond mitotic inactivity or metabolic restriction. Interrogating the quiescent genome and transcriptome using deep-sequencing technologies offers an unprecedented view of the global mechanisms governing this reversibly arrested cellular state and its importance for cell identity. While many efforts have identified and isolated pure target stem cell populations from a variety of adult tissues, there is a growing appreciation that their isolation from the stem cell niche in vivo leads to activation and loss of hallmarks of quiescence. Thus, in vitro models that recapitulate the dynamic reversibly arrested stem cell state in culture and lend themselves to comparison with the activated or differentiated state are useful templates for genome-wide analysis of the quiescence network.In this chapter, we describe the methods that can be adopted for whole genome epigenomic and transcriptomic analysis of cells derived from one such established culture model where mouse myoblasts are triggered to enter or exit quiescence as homogeneous populations. The ability to synchronize myoblasts in G permits insights into the genome in "deep quiescence." The culture methods for generating large populations of quiescent myoblasts in either 2D or 3D culture formats are described in detail in a previous chapter in this series (Arora et al. Methods Mol Biol 1556:283-302, 2017). Among the attractive features of this model are that genes isolated from quiescent myoblasts in culture mark satellite cells in vivo (Sachidanandan et al., J Cell Sci 115:2701-2712, 2002) providing a validation of its approximation of the molecular state of true stem cells. Here, we provide our working protocols for ChIP-seq and RNA-seq analysis, focusing on those experimental elements that require standardization for optimal analysis of chromatin and RNA from quiescent myoblasts, and permitting useful and revealing comparisons with proliferating myoblasts or differentiated myotubes.

}, issn = {1940-6029}, doi = {10.1007/978-1-4939-7371-2_16}, author = {Srivastava, Surabhi and Gala, Hardik P and Mishra, Rakesh K and Dhawan, Jyotsna} } @article {1216, title = {A double helical motif in OCIAD2 is essential for its localization, interactions and STAT3 activation}, journal = {Scientific Reports}, volume = {8}, year = {2018}, month = {2018/05/09}, pages = {7362}, abstract = {

The Ovarian Carcinoma Immunoreactive Antigen domain (OCIAD) - containing proteins OCIAD1/Asrij and OCIAD2, are implicated in several cancers and neurodegenerative diseases. While Asrij has a conserved role in facilitating STAT3 activation for JAK/STAT signaling, the expression and function of OCIAD2 in non-cancerous contexts remains unknown. Here, we report that ociad2 neighbors ociad1/asrij in most vertebrate genomes, and the two genes likely arose by tandem gene duplication, probably somewhere between the Ordovician and Silurian eras. We show that ociad2 expression is higher in the mouse kidney, liver and brain relative to other tissues. OCIAD2 localizes to early endosomes and mitochondria, and interacts with Asrij and STAT3. Knockdown and overexpression studies showed that OCIAD2 is essential for STAT3 activation and cell migration, which could contribute to its role in tumor metastasis. Structure prediction programs, protein disruption studies, biochemical and functional assays revealed a double helical motif in the OCIA domain that is necessary and sufficient for its localization, interactions and STAT3 activation. Given the importance of JAK/STAT signaling in development and disease, our studies shed light on the evolution and conserved function of the OCIA domain in regulating this pathway and will be critical for understanding this clinically important protein family.

}, isbn = {2045-2322}, url = {https://doi.org/10.1038/s41598-018-25667-3}, author = {Sinha, Saloni and Bheemsetty, Venkata Anudeep and Inamdar, Maneesha S.} } @article {1585, title = {Experiential contributions to social dominance in a rat model of fragile-X syndrome.}, journal = {Proc Biol Sci}, volume = {285}, year = {2018}, month = {2018 Jun 13}, abstract = {

Social withdrawal is one phenotypic feature of the monogenic neurodevelopmental disorder fragile-X. Using a {\textquoteright}knockout{\textquoteright} rat model of fragile-X, we examined whether deletion of the gene that causes this condition would affect the ability to form and express a social hierarchy as measured in a tube test. Male fragile-X {\textquoteright}knockout{\textquoteright} rats living together could successfully form a social dominance hierarchy, but were significantly subordinate to wild-type animals in mixed group cages. Over 10 days of repeated testing, the fragile-X mutant rats gradually showed greater variance and instability of rank during their tube-test encounters. This affected the outcome of future encounters with stranger animals from other cages, with the initial phenotype of wild-type dominance lost to a more complex picture that reflected, regardless of genotype, the prior experience of winning or losing. Our findings offer a novel insight into the complex dynamics of social interactions between laboratory living groups of fragile-X and wild-type rats. Even though this is a monogenic condition, experience has an impact upon future interactions with other animals. Gene/environment interactions should therefore be considered in the development of therapeutics.

}, issn = {1471-2954}, doi = {10.1098/rspb.2018.0294}, author = {Saxena, K and Webster, J and Hallas-Potts, A and Mackenzie, R and Spooner, P A and Thomson, D and Kind, P and Chatterji, S and Morris, R G M} } @article {1584, title = {Extinction recall of fear memories formed before stress is not affected despite higher theta activity in the amygdala.}, journal = {Elife}, volume = {7}, year = {2018}, month = {2018 08 13}, abstract = {

Stress is known to exert its detrimental effects not only by enhancing fear, but also by impairing its extinction. However, in earlier studies stress exposure preceded both processes. Thus, compared to unstressed animals, stressed animals had to extinguish fear memories that were strengthened by prior exposure to stress. Here, we dissociate the two processes to examine if stress specifically impairs the acquisition and recall of fear extinction. Strikingly, when fear memories were formed before stress exposure, thereby allowing animals to initiate extinction from comparable levels of fear, recall of fear extinction was unaffected. Despite this, we observed a persistent increase in theta activity in the BLA. Theta activity in the mPFC, by contrast, was normal. Stress also disrupted mPFC-BLA theta-frequency synchrony and directional coupling. Thus, in the absence of the fear-enhancing effects of stress, the expression of fear during and after extinction reflects normal regulation of theta activity in the mPFC, not theta hyperactivity in the amygdala.

}, keywords = {Amygdala, Animals, Extinction, Psychological, Fear, Male, Memory, Mental Recall, Prefrontal Cortex, Rats, Sprague-Dawley, Stress, Physiological, Theta Rhythm}, issn = {2050-084X}, doi = {10.7554/eLife.35450}, author = {Rahman, Mohammed Mostafizur and Shukla, Ashutosh and Chattarji, Sumantra} } @article {1598, title = {FMRP Interacts with C/D Box snoRNA in the Nucleus and Regulates Ribosomal RNA Methylation.}, journal = {iScience}, volume = {9}, year = {2018}, month = {2018 Nov 30}, pages = {399-411}, abstract = {

FMRP is an RNA-binding protein that is known to localize in the cytoplasm and in the nucleus. Here, we have identified an interaction of FMRP with a specific set of C/D box snoRNAs in the nucleus. C/D box snoRNAs guide 2{\textquoteright}O methylations of ribosomal RNA (rRNA) on defined sites, and this modification regulates rRNA folding and assembly of ribosomes. 2{\textquoteright}O methylation of rRNA is partial on several sites in human embryonic stem cells, which results in ribosomes with differential methylation patterns. FMRP-snoRNA interaction affects rRNA methylation on several of these sites, and in the absence of FMRP, differential methylation pattern of rRNA is significantly altered. We found that FMRP recognizes ribosomes carrying specific methylation patterns on rRNA and the recognition of methylation pattern by FMRP may potentially determine the translation status of its target mRNAs. Thus, FMRP integrates its function in the nucleus and in the cytoplasm.

}, issn = {2589-0042}, doi = {10.1016/j.isci.2018.11.007}, author = {D{\textquoteright}Souza, Michelle Ninochka and Gowda, Naveen Kumar Chandappa and Tiwari, Vishal and Babu, Rosana Ottakandathil and Anand, Praveen and Dastidar, Sudhriti Ghosh and Singh, Randhir and James, Owen G and Selvaraj, Bhuvaneish and Pal, Rakhi and Ramesh, Arati and Chattarji, Sumantra and Chandran, Siddharthan and Gulyani, Akash and Palakodeti, Dasaradhi and Muddashetty, Ravi S} } @article {1153, title = {Identification of multiple isomeric core chitobiose-modified high-mannose and paucimannose -glycans in the planarian .}, journal = {J Biol Chem}, volume = {293}, year = {2018}, month = {2018 May 04}, pages = {6707-6720}, abstract = {

Cell surface-associated glycans mediate many cellular processes, including adhesion, migration, signaling, and extracellular matrix organization. The galactosylation of core fucose (GalFuc epitope) in paucimannose and complex-type -glycans is characteristic of protostome organisms, including flatworms (planarians). Although uninvestigated, the structures of these glycans may play a role in planarian regeneration. Whole-organism MALDI-MS analysis of -linked oligosaccharides from the planarian revealed the presence of multiple isomeric high-mannose and paucimannose structures with unusual mono-, di-, and polygalactosylated ( = 3-5) core fucose structures; the latter structures have not been reported in other systems. Di- and trigalactosylated core fucoses were the most dominant glycomers. -Glycans showed extensive, yet selective, methylation patterns, ranging from non-methylated to polymethylated glycoforms. Although the majority of glycoforms were polymethylated, a small fraction also consisted of non-methylated glycans. Remarkably, monogalactosylated core fucose remained unmethylated, whereas its polygalactosylated forms were methylated, indicating structurally selective methylation. Using database searches, we identified two potential homologs of the Galβ1-4Fuc-synthesizing enzyme from nematodes (GALT-1) that were expressed in the prepharyngeal, pharyngeal, and mesenchymal regions in The presence of two GALT-1 homologs suggests different requirements for mono- and polygalactosylation of core fucose for the formation of multiple isomers. Furthermore, we observed variations in core fucose glycosylation patterns in different planarian strains, suggesting evolutionary adaptation in fucose glycosylation. The various core chitobiose modifications and methylations create \>60 different glycoforms in These results contribute greatly to our understanding of -glycan biosynthesis and suggest the presence of a GlcNAc-independent biosynthetic pathway in

}, issn = {1083-351X}, doi = {10.1074/jbc.RA117.000782}, author = {Subramanian, Sabarinath Peruvemba and Babu, Ponnusamy and Palakodeti, Dasaradhi and Subramanian, Ramaswamy} } @article {1581, title = {Infectivity of adeno-associated virus serotypes in mouse testis.}, journal = {BMC Biotechnol}, volume = {18}, year = {2018}, month = {2018 Nov 01}, pages = {70}, abstract = {

BACKGROUND: Recombinant adeno-associated viruses (AAVs) are emerging as favoured transgene delivery vectors for both research applications and gene therapy. In this context, a thorough investigation of the potential of various AAV serotypes to transduce specific cell types is valuable. Here, we rigorously tested the infectivity of a number of AAV serotypes in murine testis by direct testicular injection.

RESULTS: We report the tropism of serotypes AAV2, 5, 8, 9 and AAVrh10 in mouse testis. We reveal unique infectivity of AAV2 and AAV9, which preferentially target intertubular testosterone-producing Leydig cells. Remarkably, AAV2 TM, a mutant for capsid designed to increase transduction, displayed a dramatic alteration in tropism; it infiltrated seminiferous tubules unlike wildtype AAV2 and transduced Sertoli cells. However, none of the AAVs tested infected spermatogonial cells.

CONCLUSIONS: In spite of direct testicular injection, none of the tested AAVs appeared to infect sperm progenitors as assayed by reporter expression. This lends support to the current view that AAVs are safe gene-therapy vehicles. However, testing the presence of rAAV genomic DNA in germ cells is necessary to assess the risk of individual serotypes.

}, issn = {1472-6750}, doi = {10.1186/s12896-018-0479-1}, author = {Rajasekaran, Santhanasabapathy and Thatte, Jayashree and Periasamy, Jayaprakash and Javali, Alok and Jayaram, Manjunath and Sen, Dwaipayan and Krishnagopal, Akshaya and Jayandharan, Giridhara R and Sambasivan, Ramkumar} } @article {1154, title = {Life as an early career researcher: interview with Minhaj Sirajuddin.}, journal = {Future Sci OA}, volume = {4}, year = {2018}, month = {2018 Feb}, pages = {FSO258}, abstract = {

Minhaj Sirajuddin is currently an Assistant Investigator at the Institute for Stem Cell Biology and Regenerative Medicine (InStem), Bangalore, India. His lab works on understanding biological motility mediated by cytoskeleton elements using biophysics and cell biology. Previously his tenures were at UCSF (CA, USA) for his postdoctoral work and the Max Planck Institute (Dortmund, Germany) for his PhD. Minhaj is a recipient of Wellcome Trust - DBT India Alliance Intermediate Fellowship and the EMBO Young Investigator Award. He was also a finalist in the inaugural Future Science Early Career Research Award.

}, issn = {2056-5623}, doi = {10.4155/fsoa-2017-0106}, author = {Sirajuddin, Minhaj} } @article {1596, title = {Methionine coordinates a hierarchically organized anabolic program enabling proliferation.}, journal = {Mol Biol Cell}, year = {2018}, month = {2018 Oct 24}, pages = {mbcE18080515}, abstract = {

Methionine availability during overall amino acid limitation metabolically reprograms cells to support proliferation, the underlying basis for which remains unclear. Here, we construct the organization of this methionine mediated anabolic program, using yeast. Combining comparative transcriptome analysis, biochemical and metabolic flux based approaches, we discover that methionine rewires overall metabolic outputs by increasing the activity of a key regulatory node. This comprises of: the pentose phosphate pathway (PPP) coupled with reductive biosynthesis, the glutamate dehydrogenase (GDH) dependent synthesis of glutamate/glutamine, and pyridoxal-5-phosphate (PLP) dependent transamination capacity. This PPP-GDH-PLP node provides the required cofactors and/or substrates for subsequent rate-limiting reactions in the synthesis of amino acids, and therefore nucleotides. These rate-limiting steps in amino acid biosynthesis are also induced in a methionine-dependent manner. This thereby results in a biochemical cascade establishing a hierarchically organized anabolic program. For this methionine mediated anabolic program to be sustained, cells co-opt a "starvation stress response" regulator, Gcn4p. Collectively, our data suggest a hierarchical metabolic framework explaining how methionine mediates an anabolic switch.

}, issn = {1939-4586}, doi = {10.1091/mbc.E18-08-0515}, author = {Walvekar, Adhish S and Srinivasan, Rajalakshmi and Gupta, Ritu and Laxman, Sunil} } @article {1583, title = {Mutation burden profile in familial Alzheimer{\textquoteright}s disease cases from India.}, journal = {Neurobiol Aging}, volume = {64}, year = {2018}, month = {2018 04}, pages = {158.e7-158.e13}, abstract = {

This study attempts to identify coding risk variants in genes previously implicated in Alzheimer{\textquoteright}s disease (AD) pathways, through whole-exome sequencing of subjects (N\ = 17) with AD, with a positive family history of dementia (familial AD). We attempted to evaluate the mutation burden in genes encoding amyloid precursor protein metabolism and previously linked to risk of dementias. Novel variants were identified in genes involved in amyloid precursor protein metabolism such as PSEN1 (chr 14:73653575, W161C, tgg \> tgT), PLAT (chr 8:42039530,G272R), and SORL1 (chr11:121414373,G601D). The mutation burden assessment of dementia-related genes for all 17 cases revealed 45 variants, which were either shared across subjects, or were present in just the 1 patient. The study shows that the clinical characteristics, and genetic correlates, obtained in this sample are broadly comparable to the other studies that have investigated familial forms of AD. Our study identifies rare deleterious genetic variations, in the coding region of genes involved in amyloid signaling, and other dementia-associated pathways.

}, keywords = {Aged, Alzheimer Disease, Amyloid beta-Protein Precursor, Genetic Association Studies, Genetic Predisposition to Disease, Genetic Variation, Humans, India, LDL-Receptor Related Proteins, Membrane Transport Proteins, Middle Aged, Mutation, Presenilin-1, Risk, Signal Transduction, Tissue Plasminogen Activator, Whole Exome Sequencing}, issn = {1558-1497}, doi = {10.1016/j.neurobiolaging.2017.12.002}, author = {Syama, Adhikarla and Sen, Somdatta and Kota, Lakshmi Narayanan and Viswanath, Biju and Purushottam, Meera and Varghese, Mathew and Jain, Sanjeev and Panicker, Mitradas M and Mukherjee, Odity} } @article {1186, title = {Networks that link cytoskeletal regulators and diaphragm proteins underpin filtration function in Drosophila nephrocytes.}, journal = {Exp Cell Res}, volume = {364}, year = {2018}, month = {2018 Mar 15}, pages = {234-242}, abstract = {

Insect nephrocytes provide a valuable model for kidney disease, as they are structurally and functionally homologous to mammalian kidney podocytes. They possess an exceptional macromolecular assembly, the nephrocyte diaphragm (ND), which serves as a filtration barrier and helps maintain tissue homeostasis by filtering out wastes and toxic products. However, the elements that maintain nephrocyte architecture and the ND are not understood. We show that Drosophila nephrocytes have a unique cytoplasmic cluster of F-actin, which is maintained by the microtubule cytoskeleton and Rho-GTPases. A balance of Rac1 and Cdc42 activity as well as proper microtubule organization and endoplasmic reticulum structure, are required to position the actin cluster. Further, ND proteins Sns and Duf also localize to this cluster and regulate organization of the actin and microtubule cytoskeleton. Perturbation of any of these inter-dependent components impairs nephrocyte ultrafiltration. Thus cytoskeletal components, Rho-GTPases and ND proteins work in concert to maintain the specialized nephrocyte architecture and function.

}, issn = {1090-2422}, doi = {10.1016/j.yexcr.2018.02.015}, author = {Muraleedharan, Simi and Sam, Aksah and Skaer, Helen and Inamdar, Maneesha S} } @article {1589, title = {OCIAD1 Controls Electron Transport Chain Complex I Activity to Regulate Energy Metabolism in Human Pluripotent Stem Cells.}, journal = {Stem Cell Reports}, volume = {11}, year = {2018}, month = {2018 Jul 10}, pages = {128-141}, abstract = {

Pluripotent stem cells (PSCs) derive energy predominantly from glycolysis and not the energy-efficient oxidative phosphorylation (OXPHOS). Differentiation is initiated with energy metabolic shift from glycolysis to OXPHOS. We investigated the role of mitochondrial energy metabolism in human PSCs using molecular, biochemical, genetic, and pharmacological approaches. We show that the carcinoma protein OCIAD1 interacts with and regulates mitochondrial complex I activity. Energy metabolic assays on live pluripotent cells showed that OCIAD1-depleted cells have increased OXPHOS and may be poised for differentiation. OCIAD1 maintains human embryonic stem cells, and its depletion by CRISPR/Cas9-mediated knockout leads to rapid and increased differentiation upon induction, whereas OCIAD1 overexpression has the opposite effect. Pharmacological alteration of complex I activity was able to rescue the defects of OCIAD1 modulation. Thus, hPSCs can exist in energy metabolic substates. OCIAD1 provides a target to screen for additional modulators of mitochondrial activity to promote transient multipotent precursor expansion or enhance differentiation.

}, issn = {2213-6711}, doi = {10.1016/j.stemcr.2018.05.015}, author = {Shetty, Deeti K and Kalamkar, Kaustubh P and Inamdar, Maneesha S} } @article {1152, title = {PAI1 mediates fibroblast-mast cell interactions in skin fibrosis.}, journal = {J Clin Invest}, volume = {128}, year = {2018}, month = {2018 May 01}, pages = {1807-1819}, abstract = {

Fibrosis is a prevalent pathological condition arising from the chronic activation of fibroblasts. This activation results from the extensive intercellular crosstalk mediated by both soluble factors and direct cell-cell connections. Prominent among these are the interactions of fibroblasts with immune cells, in which the fibroblast-mast cell connection, although acknowledged, is relatively unexplored. We have used a Tg mouse model of skin fibrosis, based on expression of the transcription factor Snail in the epidermis, to probe the mechanisms regulating mast cell activity and the contribution of these cells to this pathology. We have discovered that Snail-expressing keratinocytes secrete plasminogen activator inhibitor type 1 (PAI1), which functions as a chemotactic factor to increase mast cell infiltration into the skin. Moreover, we have determined that PAI1 upregulates intercellular adhesion molecule type 1 (ICAM1) expression on dermal fibroblasts, rendering them competent to bind to mast cells. This heterotypic cell-cell adhesion, also observed in the skin fibrotic disorder scleroderma, culminates in the reciprocal activation of both mast cells and fibroblasts, leading to the cascade of events that promote fibrogenesis. Thus, we have identified roles for PAI1 in the multifactorial program of fibrogenesis that expand its functional repertoire beyond its canonical role in plasmin-dependent processes.

}, issn = {1558-8238}, doi = {10.1172/JCI99088}, author = {Pincha, Neha and Hajam, Edries Yousaf and Badarinath, Krithika and Batta, Surya Prakash Rao and Masudi, Tafheem and Dey, Rakesh and Andreasen, Peter and Kawakami, Toshiaki and Samuel, Rekha and George, Renu and Danda, Debashish and Jacob, Paul Mazhuvanchary and Jamora, Colin} } @inbook {1193, title = {Prevention of Metal Exposure: Chelating Agents and Barrier Creams}, booktitle = {Metal Allergy: From Dermatitis to Implant and Device Failure}, year = {2018}, pages = {227{\textendash}246}, publisher = {Springer International Publishing}, organization = {Springer International Publishing}, address = {Cham}, abstract = {

Metals are a group of elements which are ubiquitous in modern life. They are used in the fields of cosmetics, water purification, medicine, paint, food products, pesticides, and almost innumerable others. As the use of metals has increased in recent decades, so has human exposure to these elements. Metals such as mercury, lead, arsenic, nickel, and others have been implicated in negatively affecting human homeostasis by causing chronic inflammatory diseases, among other serious conditions. Both acute and chronic metal toxicity in vital organs could arise from local or systemic exposure to numerous metals. Although some metals have health benefits, overaccumulation of metals in body tissues can result in deleterious, toxic effects. Most exposure to metals occurs via cutaneous, inhalation, or oral routes. At the highest risk of negative effects of exposure are pregnant women and children. To ameliorate or prevent the toxic effects of metals, chelating agents and barrier creams are used widely in medical practice today. In this chapter, we will discuss preventing metal toxicity from overexposure via chelation therapy and skin barrier creams.

}, isbn = {978-3-319-58503-1}, doi = {10.1007/978-3-319-58503-1_18}, url = {https://doi.org/10.1007/978-3-319-58503-1_18}, author = {Mahato, Manohar and Sherman, Nicholas E. and Kiran Kumar Mudnakudu, N. and Joshi, Nitin and Briand, Elisabeth and Karp, Jeffrey M. and Vemula, Praveen Kumar}, editor = {Chen, Jennifer K and Thyssen, Jacob P.} } @article {1595, title = {Prevention of pesticide-induced neuronal dysfunction and mortality with nucleophilic poly-Oxime topical gel.}, journal = {Sci Adv}, volume = {4}, year = {2018}, month = {2018 Oct}, pages = {eaau1780}, abstract = {

Organophosphate-based pesticides inhibit acetylcholinesterase (AChE), which plays a pivotal role in neuromuscular function. While spraying in the field, farmworkers get exposed to pesticides through the dermal route. Internalized pesticide inhibits AChE, which leads to neurotoxicity, cardiotoxicity, cognitive dysfunction, loss of endurance, and death in severe cases. Here, we present a nucleophilic pyridine-2-aldoxime-functionalized chitosan-based topical gel (-Oxime gel) that rapidly deactivates organophosphates, methyl parathion (MPT), on the skin of rats, which leads to reduced AChE inhibition in the blood and tissues. Testing the robustness of -Oxime gel, we report reduction in AChE inhibition following repeated dermal administration of MPT in the presence of -Oxime gel. Furthermore, -Oxime gel prevented MPT-induced neuromuscular dysfunction, loss of endurance, and locomotor coordination. We observe a 100\% survival in rats following topical MPT administration in the presence of -Oxime gel. This prophylactic gel may therefore help farmworkers by limiting pesticide-induced toxicity and mortality.

}, issn = {2375-2548}, doi = {10.1126/sciadv.aau1780}, author = {Thorat, Ketan and Pandey, Subhashini and Chandrashekharappa, Sandeep and Vavilthota, Nikitha and Hiwale, Ankita A and Shah, Purna and Sreekumar, Sneha and Upadhyay, Shubhangi and Phuntsok, Tenzin and Mahato, Manohar and Mudnakudu-Nagaraju, Kiran K and Sunnapu, Omprakash and Vemula, Praveen K} } @article {1151, title = {Rudhira/BCAS3 is essential for mouse development and cardiovascular patterning.}, journal = {Sci Rep}, volume = {8}, year = {2018}, month = {2018 Apr 04}, pages = {5632}, abstract = {

Rudhira/Breast Carcinoma Amplified Sequence 3 (BCAS3) is a cytoskeletal protein that promotes directional cell migration and angiogenesis in vitro and is implicated in human carcinomas and coronary artery disease. To study the role of Rudhira during development in vivo, we generated the first knockout mouse for rudhira and show that Rudhira is essential for mouse development. Rudhira null embryos die at embryonic day (E) 9.5 accompanied by severe vascular patterning defects in embryonic and extra-embryonic tissues. To identify the molecular processes downstream of rudhira, we analyzed the transcriptome of intact knockout yolk sacs. Genome-wide transcriptome analysis showed that Rudhira functions in angiogenesis and its related processes such as cell adhesion, extracellular matrix organization, peptidase activity and TGFβ signaling. Since Rudhira is also expressed in endothelial cells\ (ECs), we further generated Tie2Cre-mediated endothelial knockout (CKO) of rudhira. CKO embryos survive to E11.5 and similar to the global knockout, display gross vascular patterning defects, showing that endothelial Rudhira is vital for development. Further, Rudhira knockdown ECs in culture fail to sprout in a spheroid-sprouting assay, strongly supporting its role in vascular patterning. Our study identifies an essential role for Rudhira in blood vessel remodeling and provides a mouse model for cardiovascular development.

}, issn = {2045-2322}, doi = {10.1038/s41598-018-24014-w}, author = {Shetty, Ronak and Joshi, Divyesh and Jain, Mamta and Vasudevan, Madavan and Paul, Jasper Chrysolite and Bhat, Ganesh and Banerjee, Poulomi and Abe, Takaya and Kiyonari, Hiroshi and VijayRaghavan, K and Inamdar, Maneesha S} } @article {1187, title = {Sequence diversity of tubulin isotypes in regulation of the mitochondrial voltage-dependent anion channel.}, journal = {J Biol Chem}, year = {2018}, month = {2018 May 18}, abstract = {

The microtubule protein tubulin is a heterodimer comprising α/β subunits, in which each subunit features multiple isotypes in vertebrates. For example, seven α-tubulin and eight β-tubulin isotypes in the human tubulin gene family vary mostly in the length and primary sequence of the disordered anionic C-terminal tails (CTTs). The biological reason for such sequence diversity remains a topic of vigorous enquiry. Here, we demonstrate that it may be a key feature of tubulin{\textquoteright}s role in regulation of the permeability of the mitochondrial outer membrane voltage-dependent anion channel (VDAC). Using recombinant yeast α/β-tubulin constructs with α-CTTs, β-CTTs, or both from various human tubulin isotypes, we probed their interactions with VDAC reconstituted into planar lipid bilayers. A comparative study of the blockage kinetics revealed that either α-CTTs or β-CTTs block VDAC pore and that the efficiency of blockage by individual CTTs spans two orders of magnitude, depending on the CTT isotype. β-Tubulin constructs, notably β3, blocked VDAC most effectively. We quantitatively describe these experimental results using a physical model that accounts only for the number and distribution of charges in the CTT, and not for the interactions between specific residues on the CTT and VDAC pore. Based on these results, we speculate that the effectiveness of VDAC regulation by tubulin depends on the predominant tubulin isotype in a cell. Consequently, the fluxes of ATP/ADP through the channel could vary significantly depending on the isotype, thus suggesting an intriguing link between VDAC regulation and the diversity of tubulin isotypes present in vertebrates.

}, issn = {1083-351X}, doi = {10.1074/jbc.RA117.001569}, author = {Rostovtseva, Tatiana K and Gurnev, Philip A and Hoogerheide, David P and Rovini, Amandine and Sirajuddin, Minhajuddin and Bezrukov, Sergey M} } @article {1588, title = {The Sodium Sialic Acid Symporter From Has Altered Substrate Specificity.}, journal = {Front Chem}, volume = {6}, year = {2018}, month = {2018}, pages = {233}, abstract = {

Mammalian cell surfaces are decorated with complex glycoconjugates that terminate with negatively charged sialic acids. Commensal and pathogenic bacteria can use host-derived sialic acids for a competitive advantage, but require a functional sialic acid transporter to import the sugar into the cell. This work investigates the sodium sialic acid symporter (SiaT) from (SiaT). We demonstrate that SiaT rescues an strain lacking its endogenous sialic acid transporter when grown on the sialic acids -acetylneuraminic acid (Neu5Ac) or -glycolylneuraminic acid (Neu5Gc). We then develop an expression, purification and detergent solubilization system for SiaT and demonstrate that the protein is largely monodisperse in solution with a stable monomeric oligomeric state. Binding studies reveal that SiaT has a higher affinity for Neu5Gc over Neu5Ac, which was unexpected and is not seen in another SiaT homolog. We develop a homology model and use comparative sequence analyses to identify substitutions in the substrate-binding site of SiaT that may explain the altered specificity. SiaT is shown to be electrogenic, and transport is dependent upon more than one Na ion for every sialic acid molecule. A functional sialic acid transporter is essential for the uptake and utilization of sialic acid in a range of pathogenic bacteria, and developing new inhibitors that target these transporters is a valid mechanism for inhibiting bacterial growth. By demonstrating a route to functional recombinant SiaT, and developing the and assay systems, our work underpins the design of inhibitors to this transporter.

}, issn = {2296-2646}, doi = {10.3389/fchem.2018.00233}, author = {North, Rachel A and Wahlgren, Weixiao Y and Remus, Daniela M and Scalise, Mariafrancesca and Kessans, Sarah A and Dunevall, Elin and Claesson, Elin and Soares da Costa, Tatiana P and Perugini, Matthew A and Ramaswamy, S and Allison, Jane R and Indiveri, Cesare and Friemann, Rosmarie and Dobson, Renwick C J} } @article {1599, title = {A strategy to identify a ketoreductase that preferentially synthesizes pharmaceutically relevant (S)-alcohols using whole-cell biotransformation.}, journal = {Microb Cell Fact}, volume = {17}, year = {2018}, month = {2018 Dec 03}, pages = {192}, abstract = {

INTRODUCTION: Chemical industries are constantly in search of an expeditious and environmentally benign method for producing chiral synthons. Ketoreductases have been used as catalysts for enantioselective conversion of desired prochiral ketones to their corresponding alcohol. We chose reported promiscuous ketoreductases belonging to different protein families and expressed them in E.\ coli to evaluate their ability as whole-cell catalysts for obtaining chiral alcohol intermediates of pharmaceutical importance. Apart from establishing a method to produce high value (S)-specific alcohols that have not been evaluated before, we propose an in silico analysis procedure\ to predict product chirality.

RESULTS: Six enzymes originating from Sulfolobus\ sulfotaricus, Zygosaccharomyces\ rouxii, Hansenula\ polymorpha, Corynebacterium sp. ST-10, Synechococcus sp. PCC\ 7942 and Bacillus sp. ECU0013 with reported efficient activity for dissimilar substrates are compared here to arrive at an optimal enzyme for the method. Whole-cell catalysis of ketone intermediates for drugs like Aprepitant, Sitagliptin and Dolastatin using E.\ coli over-expressing these enzymes yielded (S)-specific chiral alcohols. We explain this chiral specificity for the best-performing enzyme, i.e., Z.\ rouxii ketoreductase using in silico modelling and MD simulations. This rationale was applied to five additional ketones that are used in the synthesis of Crizotinib, MA-20565\ (an antifungal agent), Sulopenem, Rivastigmine, Talampanel and Barnidipine and predicted the yield of (S) enantiomers. Experimental evaluation matched the in silico analysis wherein ~ 95\% (S)-specific alcohol with a chemical yield of 23-79\% was obtained through biotransformation. Further, the cofactor re-cycling was optimized by switching the carbon source from glucose to sorbitol that improved the chemical yield to 85-99\%.

CONCLUSIONS: Here, we present a strategy to synthesize pharmaceutically relevant chiral alcohols by ketoreductases using a cofactor balanced whole-cell catalysis scheme that is useful for the industry. Based on the results obtained in these trials, Zygosaccharomyces\ rouxii ketoreductase was identified as a proficient enzyme to obtain (S)-specific alcohols from their respective ketones. The whole-cell catalyst when combined with nutrient modulation of using sorbitol as a carbon source helped obtain high enantiomeric and chemical yield.

}, keywords = {Biotransformation, Catalysis, Ethanol, Ketones}, issn = {1475-2859}, doi = {10.1186/s12934-018-1036-2}, author = {Haq, Saiful F and Shanbhag, Anirudh P and Karthikeyan, Subbulakshmi and Hassan, Imran and Thanukrishnan, Kannan and Ashok, Abhishek and Sukumaran, Sunilkumar and Ramaswamy, S and Bharatham, Nagakumar and Datta, Santanu and Samant, Shalaka and Katagihallimath, Nainesh} } @article {1147, title = {Substrate-bound outward-open structure of a Na-coupled sialic acid symporter reveals a new Na site.}, journal = {Nat Commun}, volume = {9}, year = {2018}, month = {2018 May 01}, pages = {1753}, abstract = {

Many pathogenic bacteria utilise sialic acids as an energy source or use them as an external coating to evade immune detection. As such, bacteria that colonise sialylated environments deploy specific transporters to mediate import of scavenged sialic acids. Here, we report a substrate-bound 1.95 {\r A} resolution structure and subsequent characterisation of SiaT, a sialic acid transporter from Proteus mirabilis. SiaT is a secondary active transporter of the sodium solute symporter (SSS) family, which use Na gradients to drive the uptake of extracellular substrates. SiaT adopts the LeuT-fold and is in an outward-open conformation in complex with the sialic acid N-acetylneuraminic acid and two Na ions. One Na binds to the conserved Na2 site, while the second Na binds to a new position, termed Na3, which is conserved in many SSS family members. Functional and molecular dynamics studies validate the substrate-binding site and demonstrate that both Na sites regulate N-acetylneuraminic acid transport.

}, issn = {2041-1723}, doi = {10.1038/s41467-018-04045-7}, author = {Wahlgren, Weixiao Y and Dunevall, Elin and North, Rachel A and Paz, Aviv and Scalise, Mariafrancesca and Bisignano, Paola and Bengtsson-Palme, Johan and Goyal, Parveen and Claesson, Elin and Caing-Carlsson, Rhawnie and Andersson, Rebecka and Beis, Konstantinos and Nilsson, Ulf J and Farewell, Anne and Pochini, Lorena and Indiveri, Cesare and Grabe, Michael and Dobson, Renwick C J and Abramson, Jeff and Ramaswamy, S and Friemann, Rosmarie} } @article {1580, title = {Targeting Phosphopeptide Recognition by the Human BRCA1 Tandem BRCT Domain to Interrupt BRCA1-Dependent Signaling.}, journal = {Cell Chem Biol}, volume = {25}, year = {2018}, month = {2018 06 21}, pages = {677-690.e12}, abstract = {

Intracellular signals triggered by DNA breakage flow through proteins containing BRCT (BRCA1 C-terminal) domains. This family, comprising 23 conserved phosphopeptide-binding modules in man, is inaccessible to small-molecule chemical inhibitors. Here, we develop Bractoppin, a drug-like inhibitor of phosphopeptide recognition by the human BRCA1 tandem (t)BRCT domain, which selectively inhibits substrate binding with nanomolar potency in\ vitro. Structure-activity exploration suggests that Bractoppin engages BRCA1 tBRCT residues recognizing pSer in the consensus motif, pSer-Pro-Thr-Phe, plus an abutting hydrophobic pocket that is distinct in structurally related BRCT domains, conferring selectivity. In cells, Bractoppin inhibits substrate recognition detected by F{\"o}rster resonance energy transfer, and diminishes BRCA1 recruitment to DNA breaks, in turn suppressing damage-induced G2 arrest and assembly of the recombinase, RAD51. But damage-induced MDC1 recruitment, single-stranded DNA (ssDNA) generation, and TOPBP1 recruitment remain unaffected. Thus, an inhibitor of phosphopeptide recognition selectively interrupts BRCA1 tBRCT-dependent signals evoked by DNA damage.

}, issn = {2451-9448}, doi = {10.1016/j.chembiol.2018.02.012}, author = {Periasamy, Jayaprakash and Kurdekar, Vadiraj and Jasti, Subbarao and Nijaguna, Mamatha B and Boggaram, Sanjana and Hurakadli, Manjunath A and Raina, Dhruv and Kurup, Lokavya Meenakshi and Chintha, Chetan and Manjunath, Kavyashree and Goyal, Aneesh and Sadasivam, Gayathri and Bharatham, Kavitha and Padigaru, Muralidhara and Potluri, Vijay and Venkitaraman, Ashok R} } @article {1146, title = {Towards an arthritis flare-responsive drug delivery system.}, journal = {Nat Commun}, volume = {9}, year = {2018}, month = {2018 Apr 03}, pages = {1275}, abstract = {

Local delivery of therapeutics for the treatment of inflammatory arthritis (IA) is limited by short intra-articular half-lives. Since IA severity often fluctuates over time, a local drug delivery method that titrates drug release to arthritis activity would represent an attractive paradigm in IA therapy. Here we report the development of a hydrogel platform that exhibits disassembly and drug release controlled by the concentration of enzymes expressed during arthritis flares. In vitro, hydrogel loaded with triamcinolone acetonide (TA) releases drug on-demand upon exposure to enzymes or synovial fluid from patients with rheumatoid arthritis. In arthritic mice, hydrogel loaded with a fluorescent dye demonstrates flare-dependent disassembly measured as loss of fluorescence. Moreover, a single dose of TA-loaded hydrogel but not the equivalent dose of locally injected free TA reduces arthritis activity in the injected paw. Together, our data suggest flare-responsive hydrogel as a promising next-generation drug delivery approach for the treatment of IA.

}, issn = {2041-1723}, doi = {10.1038/s41467-018-03691-1}, author = {Joshi, Nitin and Yan, Jing and Levy, Seth and Bhagchandani, Sachin and Slaughter, Kai V and Sherman, Nicholas E and Amirault, Julian and Wang, Yufeng and Riegel, Logan and He, Xueyin and Rui, Tan Shi and Valic, Michael and Vemula, Praveen K and Miranda, Oscar R and Levy, Oren and Gravallese, Ellen M and Aliprantis, Antonios O and Ermann, Joerg and Karp, Jeffrey M} } @article {1191, title = {Tunable Emission from Fluorescent Organic Nanoparticles in Water: Insight into the Nature of Self-Assembly and Photoswitching}, journal = {Chemistry {\textendash} A European Journal}, volume = {24}, year = {2018}, pages = {2643-2652}, abstract = {

Abstract Excitation-dependent tuning of the emission behavior of fluorescent organic nanoparticles (FONs) with two simple luminescent pyrenyl{\textendash}pyridyl conjugates as model systems is demonstrated. In the case of the compound with a flexible bis-picolyl moiety, the simultaneous presence of multiple ground-state species with distinct absorption and emission characteristics can be observed. The relative ratios of these species can easily be modulated, and it is possible to selectively stimulate any one of them individually by choosing an appropriate excitation channel. Moreover, at high concentration, a drastic change in the nature of the self-assembly is observed, which shifts from donor{\textendash}acceptor-type self-assembly to exciplex-type self-agglomeration. On the contrary, the compound containing a rigid terpyridine unit has only a single ground state and shows no such tunable emission. However, it can exhibit multiple emission bands in water, whereby the positions of their emission maxima depend on the extent of aggregation-induced planarization of the probe molecules. Overall, this work demonstrates multimodal modulation of FON emission and a gives insight into how molecular order can translate into complete switching of nanoparticle self-assembly and photophysics.

}, keywords = {aggregation, Fluorescence, nanoparticles, self-assembly}, doi = {10.1002/chem.201704607}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/chem.201704607}, author = {Gulyani Akash and Dey Nilanjan and Bhattacharya Santanu} } @article {1158, title = {Co-expression of Tbx6 and Sox2 identifies a novel transient neuromesoderm progenitor cell state.}, journal = {Development}, volume = {144}, year = {2017}, month = {2017 12 15}, pages = {4522-4529}, abstract = {

Elongation of the body axis is a key aspect of body plan development. Bipotential neuromesoderm progenitors (NMPs) ensure axial growth of embryos by contributing both to the spinal cord and mesoderm. The current model for the mechanism controlling NMP deployment invokes Tbx6, a T-box factor, to drive mesoderm differentiation of NMPs. Here, we identify a new population of Tbx6 cells in a subdomain of the NMP niche in mouse embryos. Based on co-expression of a progenitor marker, Sox2, we identify this population as representing a transient cell state in the mesoderm-fated NMP lineage. Genetic lineage tracing confirms the presence of the NMP cell state. Furthermore, we report a novel aspect of the documented mutant phenotype, namely an increase from two to four ectopic neural tubes, corresponding to the switch in NMP niche, thus highlighting the importance of function in NMP fate decision. This study emphasizes the function of Tbx6 as a bistable switch that turns mesoderm fate {\textquoteright}on{\textquoteright} and progenitor state {\textquoteright}off{\textquoteright}, and thus has implications for the molecular mechanism driving NMP fate choice.

}, keywords = {Animals, Body Patterning, Cell Differentiation, Cell Lineage, Embryonic Stem Cells, Gene Expression Regulation, Developmental, Mesoderm, Mice, Mice, Transgenic, Neural Tube, SOXB1 Transcription Factors, Spinal Cord, Transcription Factors}, issn = {1477-9129}, doi = {10.1242/dev.153262}, author = {Javali, Alok and Misra, Aritra and Leonavicius, Karolis and Acharyya, Debalina and Vyas, Bhakti and Sambasivan, Ramkumar} } @article {1185, title = {Crystal structure of N-acetylmannosamine kinase from Fusobacterium nucleatum.}, journal = {Acta Crystallogr F Struct Biol Commun}, volume = {73}, year = {2017}, month = {2017 Jun 01}, pages = {356-362}, abstract = {

Sialic acids comprise a varied group of nine-carbon amino sugars that are widely distributed among mammals and higher metazoans. Some human commensals and bacterial pathogens can scavenge sialic acids from their environment and degrade them for use as a carbon and nitrogen source. The enzyme N-acetylmannosamine kinase (NanK; EC 2.7.1.60) belongs to the transcriptional repressors, uncharacterized open reading frames and sugar kinases (ROK) superfamily. NanK catalyzes the second step of the sialic acid catabolic pathway, transferring a phosphate group from adenosine 5{\textquoteright}-triphosphate to the C6 position of N-acetylmannosamine to generate N-acetylmannosamine 6-phosphate. The structure of NanK from Fusobacterium nucleatum was determined to 2.23 {\r A} resolution by X-ray crystallography. Unlike other NanK enzymes and ROK family members, F. nucleatum NanK does not have a conserved zinc-binding site. In spite of the absence of the zinc-binding site, all of the major structural features of enzymatic activity are conserved.

}, keywords = {Adenosine Triphosphate, Amino Acid Sequence, Bacterial Proteins, Binding Sites, Cloning, Molecular, Crystallography, X-Ray, Escherichia coli, Fusobacterium nucleatum, Gene Expression, Genetic Vectors, Hexosamines, Models, Molecular, Phosphotransferases (Alcohol Group Acceptor), Protein Binding, Protein Conformation, alpha-Helical, Protein Conformation, beta-Strand, Protein Interaction Domains and Motifs, Protein Multimerization, Recombinant Proteins, Sequence Alignment, Sequence Homology, Amino Acid, Substrate Specificity}, issn = {2053-230X}, doi = {10.1107/S2053230X17007439}, author = {Caing-Carlsson, Rhawnie and Goyal, Parveen and Sharma, Amit and Ghosh, Swagatha and Setty, Thanuja Gangi and North, Rachel A and Friemann, Rosmarie and Ramaswamy, S} } @article {1210, title = {C-State: an interactive web app for simultaneous multi-gene visualization and comparative epigenetic pattern search.}, journal = {BMC Bioinformatics}, volume = {18}, year = {2017}, month = {2017 Sep 13}, pages = {392}, abstract = {

BACKGROUND: Comparative epigenomic analysis across multiple genes presents a bottleneck for bench biologists working with NGS data. Despite the development of standardized peak analysis algorithms, the identification of novel epigenetic patterns and their visualization across gene subsets remains a challenge.

RESULTS: We developed a fast and interactive web app, C-State (Chromatin-State), to query and plot chromatin landscapes across multiple loci and cell types. C-State has an interactive, JavaScript-based graphical user interface and runs locally in modern web browsers that are pre-installed on all computers, thus eliminating the need for cumbersome data transfer, pre-processing and prior programming knowledge.

CONCLUSIONS: C-State is unique in its ability to extract and analyze multi-gene epigenetic information. It allows for powerful GUI-based pattern searching and visualization. We include a case study to demonstrate its potential for identifying user-defined epigenetic trends in context of gene expression profiles.

}, keywords = {Algorithms, Embryonic Stem Cells, Epigenomics, Genes, Genomics, HeLa Cells, Humans, Internet, K562 Cells, Promoter Regions, Genetic, Software, Transcription, Genetic, Web Browser}, issn = {1471-2105}, doi = {10.1186/s12859-017-1786-6}, author = {Sowpati, Divya Tej and Srivastava, Surabhi and Dhawan, Jyotsna and Mishra, Rakesh K} } @article {1160, title = {Cytoplasmic poly (A)-binding protein critically regulates epidermal maintenance and turnover in the planarian .}, journal = {Development}, volume = {144}, year = {2017}, month = {2017 09 01}, pages = {3066-3079}, abstract = {

Identifying key cellular events that facilitate stem cell function and tissue organization is crucial for understanding the process of regeneration. Planarians are powerful model system to study regeneration and stem cell (neoblast) function. Here, using planaria, we show that the initial events of regeneration, such as epithelialization and epidermal organization are critically regulated by a novel cytoplasmic poly A-binding protein, SMED-PABPC2. Knockdown leads to defects in epidermal lineage specification, disorganization of epidermis and ECM, and deregulated wound healing, resulting in the selective failure of neoblast proliferation near the wound region. Polysome profiling suggests that epidermal lineage transcripts, including , are translationally regulated by SMED-PABPC2 Together, our results uncover a novel role for SMED-PABPC2 in the maintenance of epidermal and ECM integrity, critical for wound healing and subsequent processes for regeneration.

}, keywords = {Animals, Cell Lineage, Cell Proliferation, Cytoplasm, Epidermis, Epithelium, Extracellular Matrix, Gene Knockdown Techniques, Homeostasis, Models, Biological, Planarians, Poly(A)-Binding Protein I, Regeneration, RNA, Messenger, Wound Healing}, issn = {1477-9129}, doi = {10.1242/dev.152942}, author = {Bansal, Dhiru and Kulkarni, Jahnavi and Nadahalli, Kavana and Lakshmanan, Vairavan and Krishna, Srikar and Sasidharan, Vidyanand and Geo, Jini and Dilipkumar, Shilpa and Pasricha, Renu and Gulyani, Akash and Raghavan, Srikala and Palakodeti, Dasaradhi} } @article {1163, title = {Hierarchies in light sensing and dynamic interactions between ocular and extraocular sensory networks in a flatworm.}, journal = {Sci Adv}, volume = {3}, year = {2017}, month = {2017 Jul}, pages = {e1603025}, abstract = {

Light sensing has independently evolved multiple times under diverse selective pressures but has been examined only in a handful among the millions of light-responsive organisms. Unsurprisingly, mechanistic insights into how differential light processing can cause distinct behavioral outputs are limited. We show how an organism can achieve complex light processing with a simple "eye" while also having independent but mutually interacting light sensing networks. Although planarian flatworms lack wavelength-specific eye photoreceptors, a 25 nm change in light wavelength is sufficient to completely switch their phototactic behavior. Quantitative photoassays, eye-brain confocal imaging, and RNA interference/knockdown studies reveal that flatworms are able to compare small differences in the amounts of light absorbed at the eyes through a single eye opsin and convert them into binary behavioral outputs. Because planarians can fully regenerate, eye-brain injury-regeneration studies showed that this acute light intensity sensing and processing are layered on simple light detection. Unlike intact worms, partially regenerated animals with eyes can sense light but cannot sense finer gradients. Planarians also show a "reflex-like," eye-independent (extraocular/whole-body) response to low ultraviolet A light, apart from the "processive" eye-brain-mediated (ocular) response. Competition experiments between ocular and extraocular sensory systems reveal dynamic interchanging hierarchies. In intact worms, cerebral ocular response can override the reflex-like extraocular response. However, injury-regeneration again offers a time window wherein both responses coexist, but the dominance of the ocular response is reversed. Overall, we demonstrate acute light intensity-based behavioral switching and two evolutionarily distinct but interacting light sensing networks in a regenerating organism.

}, issn = {2375-2548}, doi = {10.1126/sciadv.1603025}, author = {Shettigar, Nishan and Joshi, Asawari and Dalmeida, Rimple and Gopalkrishna, Rohini and Chakravarthy, Anirudh and Patnaik, Siddharth and Mathew, Manoj and Palakodeti, Dasaradhi and Gulyani, Akash} } @article {1204, title = {Horse Liver Alcohol Dehydrogenase: Zinc Coordination and Catalysis.}, journal = {Biochemistry}, volume = {56}, year = {2017}, month = {2017 07 18}, pages = {3632-3646}, abstract = {

During catalysis by liver alcohol dehydrogenase (ADH), a water bound to the catalytic zinc is replaced by the oxygen of the substrates. The mechanism might involve a pentacoordinated zinc or a double-displacement reaction with participation by a nearby glutamate residue, as suggested by studies of human ADH3, yeast ADH1, and some other tetrameric ADHs. Zinc coordination and participation of water in the enzyme mechanism were investigated by X-ray crystallography. The apoenzyme and its complex with adenosine 5{\textquoteright}-diphosphoribose have an open protein conformation with the catalytic zinc in one position, tetracoordinated by Cys-46, His-67, Cys-174, and a water molecule. The bidentate chelators 2,2{\textquoteright}-bipyridine and 1,10-phenanthroline displace the water and form a pentacoordinated zinc. The enzyme-NADH complex has a closed conformation similar to that of ternary complexes with coenzyme and substrate analogues; the coordination of the catalytic zinc is similar to that found in the apoenzyme, except that a minor, alternative position for the catalytic zinc is \~{}1.3 {\r A} from the major position and closer to Glu-68, which could form the alternative coordination to the catalytic zinc. Complexes with NADH and N-1-methylhexylformamide or N-benzylformamide (or with NAD and fluoro alcohols) have the classical tetracoordinated zinc, and no water is bound to the zinc or the nicotinamide rings. The major forms of the enzyme in the mechanism have a tetracoordinated zinc, where the carboxylate group of Glu-68 could participate in the exchange of water and substrates on the zinc. Hydride transfer in the Michaelis complexes does not involve a nearby water.

}, keywords = {2,2{\textquoteright}-Dipyridyl, Adenosine Diphosphate Ribose, Alcohol Dehydrogenase, Animals, Catalytic Domain, Crystallography, X-Ray, Formamides, Horses, Kinetics, Liver, Models, Molecular, NAD, Phenanthrolines, Protein Binding, Protein Conformation, Water, Zinc}, issn = {1520-4995}, doi = {10.1021/acs.biochem.7b00446}, author = {Plapp, Bryce V and Savarimuthu, Baskar Raj and Ferraro, Daniel J and Rubach, Jon K and Brown, Eric N and Ramaswamy, S} } @article {1199, title = {The miR-124 family of microRNAs is crucial for regeneration of the brain and visual system in the planarian Schmidtea mediterranea}, journal = {Development}, volume = {144}, year = {2017}, pages = {3211{\textendash}3223}, abstract = {

Brain regeneration in planarians is mediated by precise spatiotemporal control of gene expression and is crucial for multiple aspects of neurogenesis. However, the mechanisms underpinning the gene regulation essential for brain regeneration are largely unknown. Here, we investigated the role of the miR-124 family of microRNAs in planarian brain regeneration. The miR-124 family (miR-124) is highly conserved in animals and regulates neurogenesis by facilitating neural differentiation, yet its role in neural wiring and brain organization is not known. We developed a novel method for delivering anti-miRs using liposomes for the functional knockdown of microRNAs. Smed-miR-124 knockdown revealed a key role for these microRNAs in neuronal organization during planarian brain regeneration. Our results also demonstrated an essential role for miR-124 in the generation of eye progenitors. Additionally, miR-124 regulates Smed-slit-1, which encodes an axon guidance protein, either by targeting slit-1 mRNA or, potentially, by modulating the canonical Notch pathway. Together, our results reveal a role for miR-124 in regulating the regeneration of a functional brain and visual system.

}, issn = {0950-1991}, doi = {10.1242/dev.144758}, url = {http://dev.biologists.org/content/144/18/3211}, author = {Sasidharan, Vidyanand and Marepally, Srujan and Elliott, Sarah A. and Baid, Srishti and Lakshmanan, Vairavan and Nayyar, Nishtha and Bansal, Dhiru and S{\'a}nchez Alvarado, Alejandro and Vemula, Praveen Kumar and Palakodeti, Dasaradhi} } @article {1159, title = {Molecular mechanisms and structural features of cardiomyopathy-causing troponin T mutants in the tropomyosin overlap region.}, journal = {Proc Natl Acad Sci U S A}, volume = {114}, year = {2017}, month = {2017 10 17}, pages = {11115-11120}, abstract = {

Point mutations in genes encoding sarcomeric proteins are the leading cause of inherited primary cardiomyopathies. Among them are mutations in the gene that encodes cardiac troponin T (TnT). These mutations are clustered in the tropomyosin (Tm) binding region of TnT, TNT1 (residues 80-180). To understand the mechanistic changes caused by pathogenic mutations in the TNT1 region, six hypertrophic cardiomyopathy (HCM) and two dilated cardiomyopathy (DCM) mutants were studied by biochemical approaches. Binding assays in the absence and presence of actin revealed changes in the affinity of some, but not all, TnT mutants for Tm relative to WT TnT. HCM mutants were hypersensitive and DCM mutants were hyposensitive to Ca in regulated actomyosin ATPase activities. To gain better insight into the disease mechanism, we modeled the structure of TNT1 and its interactions with Tm. The stability predictions made by the model correlated well with the affinity changes observed in vitro of TnT mutants for Tm. The changes in Ca sensitivity showed a strong correlation with the changes in binding affinity. We suggest the primary reason by which these mutations between residues 92 and 144 cause cardiomyopathy is by changing the affinity of TnT for Tm within the TNT1 region.

}, issn = {1091-6490}, doi = {10.1073/pnas.1710354114}, author = {Gangadharan, Binnu and Sunitha, Margaret S and Mukherjee, Souhrid and Chowdhury, Ritu Roy and Haque, Farah and Sekar, Narendrakumar and Sowdhamini, Ramanathan and Spudich, James A and Mercer, John A} } @article {1165, title = {Morphology transition in helical tubules of a supramolecular gel driven by metal ions.}, journal = {Chem Commun (Camb)}, volume = {53}, year = {2017}, month = {2017 Jan 26}, pages = {1538-1541}, abstract = {

Our aim to access a particular chemical functionality on helical tubules has been achieved by the rational molecular design and synthesis of glucono-appended cardanol derivatives. For the first time, we report a chiral molecular packing with α-helical tubules, and chiral symmetry-breaking upon exposure to Cu that generated the final ordered structure via an in situ morphological transition without undergoing any phase change.

}, issn = {1364-548X}, doi = {10.1039/c6cc09120b}, author = {Lalitha, Krishnamoorthy and Sridharan, Vellaisamy and Maheswari, C Uma and Vemula, Praveen Kumar and Nagarajan, Subbiah} } @article {1201, title = {Scaling the effect of hydrophobic chain length on gene transfer properties of di-alkyl{,} di-hydroxy ethylammonium chloride based cationic amphiphiles}, journal = {RSC Adv.}, volume = {7}, year = {2017}, pages = {25398-25405}, abstract = {

The success of gene therapy critically depends on the availability of efficient transfection vectors. Cationic lipids are the most widely studied non-viral vectors. The molecular architecture of the cationic lipid determines its transfection efficiency. Variations in alkyl chain lengths of lipids influence self-assembly and liposomal fusion with the cell membrane. These factors determine the transfection ability of the lipid. Thus{,} to probe the effect of asymmetry in hydrophobic chains on transfection efficiency{,} we designed and synthesized a series of cationic lipids by systematically varying one of the two alkyl chains linked to the quaternary nitrogen centre from C18 to C10 and keeping the other alkyl C18 chain constant (Lip1818-Lip1810). Transfection studies in multiple cultured mammalian cells (CHO{,} B16F10 and HeLa) revealed that the lipids with C18:C14 and C18:C12 alkyl chains (Lip1814 \& Lip1812) showed 20-30\% higher transfection efficacies than their counterparts at 2 : 1 and 4 : 1 lipid to pDNA charge ratios. Cryo-transmission electron images showed unilamellar vesicle structures for the liposomes of lipids. Mechanistic studies involving Small Angle X-ray Scattering (SAXS) revealed that asymmetry in the hydrophobic region has a significant impact on liposomal fusion with the plasma membrane model. Collectively{,} these findings demonstrate that chain length asymmetry in the hydrophobic region of cationic lipids has an important role in their liposome-DNA interactions at optimal 2 : 1 and 4 : 1 lipid to pDNA charge ratios{,} which in turn modulates their gene transfer properties.

}, doi = {10.1039/C7RA02271A}, url = {http://dx.doi.org/10.1039/C7RA02271A}, author = {Hiwale, Ankita A. and Voshavar, Chandrashekhar and Dharmalingam, Priya and Dhayani, Ashish and Mukthavaram, Rajesh and Nadella, Rasajna and Sunnapu, Omprakash and Gandhi, Sivaraman and Naidu, V. G. M. and Chaudhuri, Arabinda and Marepally, Srujan and Vemula, Praveen Kumar} } @article {1203, title = {Structural and functional studies of ferredoxin and oxygenase components of 3-nitrotoluene dioxygenase from Diaphorobacter sp. strain DS2.}, journal = {PLoS One}, volume = {12}, year = {2017}, month = {2017}, pages = {e0176398}, abstract = {

3-nitrotoluene dioxygenase (3NTDO) from Diaphorobacter sp. strain DS2 catalyses the conversion of 3-nitrotoluene (3NT) into a mixture of 3- and 4-methylcatechols with release of nitrite. We report here, X-ray crystal structures of oxygenase and ferredoxin components of 3NTDO at 2.9 {\r A} and 2.4 {\r A}, respectively. The residues responsible for nitrite release in 3NTDO were further probed by four single and two double mutations in the catalytic site of α-subunit of the dioxygenase. Modification of Val 350 to Phe, Ile 204 to Ala, and Asn258 to Val by site directed mutagenesis resulted in inactive enzymes revealing the importance of these residues in catalysis. Docking studies of meta nitrotoluene to the active site of 3NTDO suggested possible orientations of binding that favor the formation of 3-methylcatechol (3MC) over 4-methylcatechol energetically. The electron transfer pathway from ferredoxin subunit to the active site of the oxygenase subunit is also proposed.

}, keywords = {Catalytic Domain, Comamonadaceae, Crystallography, X-Ray, Ferredoxins, Molecular Docking Simulation, Mutation, Oxygenases, Substrate Specificity, Toluene}, issn = {1932-6203}, doi = {10.1371/journal.pone.0176398}, author = {Kumari, Archana and Singh, Deepak and Ramaswamy, S and Ramanathan, Gurunath} } @article {1200, title = {Targeted delivery of microbial metabolite, urolithin A protects from chemically (DSS or TNBS) induced colitis in pre-clinical models}, journal = {The Journal of Immunology}, volume = {198}, year = {2017}, pages = {65.6{\textendash}65.6}, abstract = {

Epidemiological data suggests that consumption of diets rich in phytochemicals are protective in inflammatory bowel diseases (IBDs). However, the beneficial effects are not uniform among individuals and attributed to variations in gut microbiota, and altered capacity to generate certain metabolites. Urolithin A (UroA) (3,8-dihydroxybenzo[c]chromen-6-one) is a microbial metabolite, derived from ellagic acid and ellagitannins, major poly phenolic components in berries and pomegranates. Here, we examined therapeutic applications of UroA and mechanisms of action in IBDs. Our studies suggested that UroA significantly reduced LPS induced inflammatory mediators (e.g., IL-6, TNF-alpha and IL-12) as well as ROS production in mouse bone marrow-derived macrophages (BMDMs), dendritic and THP1 cells. Most importantly, UroA also reduced LPS induced systemic inflammation in mouse models. Next, we examined therapeutic applications of UroA in dextran sodium sulphate (DSS)-induced, 2,4,6-Trinitrobenzenesulfonic acid (TNBS)-induced colitis models. Treatment with UroA significantly reduced both acute and chronic DSS-induced colitis as well as TNBS induced colitis in mouse models. Most importantly, delivery of UroA utilizing inflammation targeting oral nano-particles (single treatment regimen) effectively mitigated the colitis in both models. In summary, these results highlight even presence of single microbial metabolite at right location at right time will have significant beneficial effects to protect from adverse inflammatory activities.

}, issn = {0022-1767}, url = {http://www.jimmunol.org/content/198/1_Supplement/65.6}, author = {Singh, Rajbir and Hegde, Bindu and Von Baby, Becca and Sadeep, C and Kotla, Niranjan and Chandrasekar, Bhargavi and Marepally, Srujan and Bodduluri, Haribabu and Vemula, Praveen K and Jala, Venkatakrishna R} } @article {1167, title = {Blue protein with red fluorescence.}, journal = {Proc Natl Acad Sci U S A}, volume = {113}, year = {2016}, month = {2016 10 11}, pages = {11513-11518}, abstract = {

The walleye (Sander vitreus) is a golden yellow fish that inhabits the Northern American lakes. The recent sightings of the blue walleye and the correlation of its sighting to possible increased UV radiation have been proposed earlier. The underlying molecular basis of its adaptation to increased UV radiation is the presence of a protein (Sandercyanin)-ligand complex in the mucus of walleyes. Degradation of heme by UV radiation results in the formation of Biliverdin IXα (BLA), the chromophore bound to Sandercyanin. We show that Sandercyanin is a monomeric protein that forms stable homotetramers on addition of BLA to the protein. A structure of the Sandercyanin-BLA complex, purified from the fish mucus, reveals a glycosylated protein with a lipocalin fold. This protein-ligand complex absorbs light in the UV region (λ of 375 nm) and upon excitation at this wavelength emits in the red region (λ of 675 nm). Unlike all other known biliverdin-bound fluorescent proteins, the chromophore is noncovalently bound to the protein. We provide here a molecular rationale for the observed spectral properties of Sandercyanin.

}, keywords = {Biliverdine, Crystallography, X-Ray, Fluorescence, Models, Molecular, Proteins, Recombinant Proteins}, issn = {1091-6490}, doi = {10.1073/pnas.1525622113}, author = {Ghosh, Swagatha and Yu, Chi-Li and Ferraro, Daniel J and Sudha, Sai and Pal, Samir Kumar and Schaefer, Wayne F and Gibson, David T and Ramaswamy, S} } @article {1170, title = {Genome-Wide Analysis of Polyadenylation Events in Schmidtea mediterranea.}, journal = {G3 (Bethesda)}, volume = {6}, year = {2016}, month = {2016 10 13}, pages = {3035-3048}, abstract = {

In eukaryotes, 3{\textquoteright} untranslated regions (UTRs) play important roles in regulating posttranscriptional gene expression. The 3{\textquoteright}UTR is defined by regulated cleavage/polyadenylation of the pre-mRNA. The advent of next-generation sequencing technology has now enabled us to identify these events on a genome-wide scale. In this study, we used poly(A)-position profiling by sequencing (3P-Seq) to capture all poly(A) sites across the genome of the freshwater planarian, Schmidtea mediterranea, an ideal model system for exploring the process of regeneration and stem cell function. We identified the 3{\textquoteright}UTRs for \~{}14,000 transcripts and thus improved the existing gene annotations. We found 97 transcripts, which are polyadenylated within an internal exon, resulting in the shrinking of the ORF and loss of a predicted protein domain. Around 40\% of the transcripts in planaria were alternatively polyadenylated (ApA), resulting either in an altered 3{\textquoteright}UTR or a change in coding sequence. We identified specific ApA transcript isoforms that were subjected to miRNA mediated gene regulation using degradome sequencing. In this study, we also confirmed a tissue-specific expression pattern for alternate polyadenylated transcripts. The insights from this study highlight the potential role of ApA in regulating the gene expression essential for planarian regeneration.

}, keywords = {3{\textquoteright} Untranslated Regions, Animals, Computational Biology, Genome, Helminth, Genome-Wide Association Study, High-Throughput Nucleotide Sequencing, MicroRNAs, Molecular Sequence Annotation, Platyhelminths, Poly A, Polyadenylation, Reproducibility of Results, RNA Interference, RNA Processing, Post-Transcriptional, RNA, Messenger}, issn = {2160-1836}, doi = {10.1534/g3.116.031120}, author = {Lakshmanan, Vairavan and Bansal, Dhiru and Kulkarni, Jahnavi and Poduval, Deepak and Krishna, Srikar and Sasidharan, Vidyanand and Anand, Praveen and Seshasayee, Aswin and Palakodeti, Dasaradhi} } @article {617, title = {Gut Microbiota Conversion of Dietary Ellagic Acid into Bioactive Phytoceutical Urolithin A Inhibits Heme Peroxidases.}, journal = {PLoS One}, volume = {11}, year = {2016}, month = {2016}, pages = {e0156811}, abstract = {

Numerous studies signify that diets rich in phytochemicals offer many beneficial functions specifically during pathologic conditions, yet their effects are often not uniform due to inter-individual variation. The host indigenous gut microbiota and their modifications of dietary phytochemicals have emerged as factors that greatly influence the efficacy of phytoceutical-based intervention. Here, we investigated the biological activities of one such active microbial metabolite, Urolithin A (UA or 3,8-dihydroxybenzo[c]chromen-6-one), which is derived from the ellagic acid (EA). Our study demonstrates that UA potently inhibits heme peroxidases i.e. myeloperoxidase (MPO) and lactoperoxidase (LPO) when compared to the parent compound EA. In addition, chrome azurol S (CAS) assay suggests that EA, but not UA, is capable of binding to Fe3+, due to its catechol-like structure, although its modest heme peroxidase inhibitory activity is abrogated upon Fe3+-binding. Interestingly, UA-mediated MPO and LPO inhibition can be prevented by innate immune protein human NGAL or its murine ortholog lipocalin 2 (Lcn2), implying the complex nature of host innate immunity-microbiota interactions. Spectral analysis indicates that UA inhibits heme peroxidase-catalyzed reaction by reverting the peroxidase back to its inactive native state. In support of these in vitro results, UA significantly reduced phorbol myristate acetate (PMA)-induced superoxide generation in neutrophils, however, EA failed to block the superoxide generation. Treatment with UA significantly reduced PMA-induced mouse ear edema and MPO activity compared to EA treated mice. Collectively, our results demonstrate that microbiota-mediated conversion of EA to UA is advantageous to both host and microbiota i.e. UA-mediated inhibition of pro-oxidant enzymes reduce tissue inflammation, mitigate non-specific killing of gut bacteria, and abrogate iron-binding property of EA, thus providing a competitive edge to the microbiota in acquiring limiting nutrient iron and thrive in the gut.

}, issn = {1932-6203}, doi = {10.1371/journal.pone.0156811}, author = {Saha, Piu and Yeoh, Beng San and Singh, Rajbir and Chandrasekar, Bhargavi and Vemula, Praveen Kumar and Haribabu, Bodduluri and Vijay-Kumar, Matam and Jala, Venkatakrishna R} } @article {1172, title = {Low Oxygen Tension Enhances Expression of Myogenic Genes When Human Myoblasts Are Activated from G0 Arrest.}, journal = {PLoS One}, volume = {11}, year = {2016}, month = {2016}, pages = {e0158860}, abstract = {

OBJECTIVES: Most cell culture studies have been performed at atmospheric oxygen tension of 21\%, however the physiological oxygen tension is much lower and is a factor that may affect skeletal muscle myoblasts. In this study we have compared activation of G0 arrested myoblasts in 21\% O2 and in 1\% O2 in order to see how oxygen tension affects activation and proliferation of human myoblasts.

MATERIALS AND METHODS: Human myoblasts were isolated from skeletal muscle tissue and G0 arrested in vitro followed by reactivation at 21\% O2 and 1\% O2. The effect was assesses by Real-time RT-PCR, immunocytochemistry and western blot.

RESULTS AND CONCLUSIONS: We found an increase in proliferation rate of myoblasts when activated at a low oxygen tension (1\% O2) compared to 21\% O2. In addition, the gene expression studies showed up regulation of the myogenesis related genes PAX3, PAX7, MYOD, MYOG (myogenin), MET, NCAM, DES (desmin), MEF2A, MEF2C and CDH15 (M-cadherin), however, the fraction of DES and MYOD positive cells was not increased by low oxygen tension, indicating that 1\% O2 may not have a functional effect on the myogenic response. Furthermore, the expression of genes involved in the TGFβ, Notch and Wnt signaling pathways were also up regulated in low oxygen tension. The differences in gene expression were most pronounced at day one after activation from G0-arrest, thus the initial activation of myoblasts seemed most sensitive to changes in oxygen tension. Protein expression of HES1 and β-catenin indicated that notch signaling may be induced in 21\% O2, while the canonical Wnt signaling may be induced in 1\% O2 during activation and proliferation of myoblasts.

}, keywords = {Adolescent, Cell Cycle Checkpoints, Cell Hypoxia, Cell Proliferation, Cell Separation, Cells, Cultured, Down-Regulation, Female, Gene Expression Regulation, Humans, Ki-67 Antigen, Male, Muscle Development, Muscle Proteins, Myoblasts, Oxygen, Receptors, Notch, Resting Phase, Cell Cycle, Transforming Growth Factor beta, Wnt Signaling Pathway, Young Adult}, issn = {1932-6203}, doi = {10.1371/journal.pone.0158860}, author = {Sellathurai, Jeeva and Nielsen, Joachim and Hejb{\o}l, Eva Kildall and J{\o}rgensen, Louise Helskov and Dhawan, Jyotsna and Nielsen, Michael Friberg Bruun and Schr{\o}der, Henrik Daa} } @article {615, title = {Notch1 regulated autophagy controls survival and suppressor activity of activated murine T-regulatory cells.}, journal = {Elife}, volume = {5}, year = {2016}, month = {2016}, abstract = {

Cell survival is one of several processes regulated by the Notch pathway in mammalian cells. Here we report functional outcomes of non-nuclear Notch signaling to activate autophagy, a conserved cellular response to nutrient stress, regulating survival in murine natural T-regulatory cells (Tregs), an immune subset controlling tolerance and inflammation. Induction of autophagy required ligand-dependent, Notch intracellular domain (NIC) activity, which controlled mitochondrial organization and survival of activated Tregs. Consistently, NIC immune-precipitated Beclin and Atg14, constituents of the autophagy initiation complex. Further, ectopic expression of an effector of autophagy (Atg3) or recombinant NIC tagged to a nuclear export signal (NIC-NES), restored autophagy and suppressor function in Notch1(-/-) Tregs. Furthermore, Notch1 deficiency in the Treg lineage resulted in immune hyperactivity, implicating Notch activity in Treg homeostasis. Notch1 integration with autophagy, revealed in these experiments, holds implications for Notch regulated cell-fate decisions governing differentiation.

}, issn = {2050-084X}, doi = {10.7554/eLife.14023}, author = {Marcel, Nimi and Sarin, Apurva} } @article {621, title = {Structure of a heterogeneous, glycosylated, lipid-bound, {\i}t in vivo-grown protein crystal at atomic resolution from the viviparous cockroach {\i}t Diploptera punctata}, journal = {IUCrJ}, volume = {3}, year = {2016}, month = {Jul}, pages = {282{\textendash}293}, abstract = {

Macromolecular crystals for X-ray diffraction studies are typically grown {\i}t in vitro} from pure and homogeneous samples; however, there are examples of protein crystals that have been identified {\i}t in vivo}. Recent developments in micro-crystallography techniques and the advent of X-ray free-electron lasers have allowed the determination of several protein structures from crystals grown {\i}t in cellulo}. Here, an atomic resolution (1.2{\AA}) crystal structure is reported of heterogeneous milk proteins grown inside a living organism in their functional niche. These {\i}t in vivo}-grown crystals were isolated from the midgut of an embryo within the only known viviparous cockroach, {\i}t Diploptera punctata}. The milk proteins crystallized in space group {\i}t P}1, and a structure was determined by anomalous dispersion from the native S atoms. The data revealed glycosylated proteins that adopt a lipocalin fold, bind lipids and organize to form a tightly packed crystalline lattice. A single crystal is estimated to contain more than three times the energy of an equivalent mass of dairy milk. This unique storage form of nourishment for developing embryos allows access to a constant supply of complete nutrients. Notably, the crystalline cockroach-milk proteins are highly heterogeneous with respect to amino-acid sequence, glycosylation and bound fatty-acid composition. These data present a unique example of protein heterogeneity within a single {\i}t in vivo}-grown crystal of a natural protein in its native environment at atomic resolution.

}, keywords = {glycosylation, protein heterogeneity, sulfur-SAD, viviparity in cockroach}, doi = {10.1107/S2052252516008903}, url = {http://dx.doi.org/10.1107/S2052252516008903}, author = {Banerjee, Sanchari and Coussens, Nathan P. and Gallat, Fran{\c c}ois-Xavier and Sathyanarayanan, Nitish and Srikanth, Jandhyam and Yagi, Koichiro J. and Gray, James S. S. and Tobe, Stephen S. and Stay, Barbara and Chavas, Leonard M. G. and Ramaswamy, Subramanian} } @article {399, title = {Mechanistic heterogeneity in contractile properties of α-tropomyosin (TPM1) mutants associated with inherited cardiomyopathies.}, journal = {J Biol Chem}, volume = {290}, year = {2015}, month = {2015 Mar 13}, pages = {7003-15}, abstract = {

The most frequent known causes of primary cardiomyopathies are mutations in the genes encoding sarcomeric proteins. Among those are 30 single-residue mutations in TPM1, the gene encoding α-tropomyosin. We examined seven mutant tropomyosins, E62Q, D84N, I172T, L185R, S215L, D230N, and M281T, that were chosen based on their clinical severity and locations along the molecule. The goal of our study was to determine how the biochemical characteristics of each of these mutant proteins are altered, which in turn could provide a structural rationale for treatment of the cardiomyopathies they produce. Measurements of Ca(2+) sensitivity of human β-cardiac myosin ATPase activity are consistent with the hypothesis that hypertrophic cardiomyopathies are hypersensitive to Ca(2+) activation, and dilated cardiomyopathies are hyposensitive. We also report correlations between ATPase activity at maximum Ca(2+) concentrations and conformational changes in TnC measured using a fluorescent probe, which provide evidence that different substitutions perturb the structure of the regulatory complex in different ways. Moreover, we observed changes in protein stability and protein-protein interactions in these mutants. Our results suggest multiple mechanistic pathways to hypertrophic and dilated cardiomyopathies. Finally, we examined a computationally designed mutant, E181K, that is hypersensitive, confirming predictions derived from in silico structural analysis.

}, keywords = {Actins, Adenosine Triphosphatases, Calcium, Cardiomyopathies, Humans, Models, Molecular, Myosins, Point Mutation, Protein Stability, Tropomyosin}, issn = {1083-351X}, doi = {10.1074/jbc.M114.596676}, author = {Gupte, Tejas M and Haque, Farah and Gangadharan, Binnu and Sunitha, Margaret S and Mukherjee, Souhrid and Anandhan, Swetha and Rani, Deepa Selvi and Mukundan, Namita and Jambekar, Amruta and Thangaraj, Kumarasamy and Sowdhamini, Ramanathan and Sommese, Ruth F and Nag, Suman and Spudich, James A and Mercer, John A} } @article {2114, title = {Metal substitutions incarbonic anhydrase: a halide ion probe study.}, journal = {Biochem Biophys Res Commun}, volume = {66}, year = {1975}, month = {1975 Oct 27}, pages = {1281-6}, keywords = {Animals, Binding Sites, Cadmium, Carbonic Anhydrases, Cattle, Humans, Hydrogen-Ion Concentration, Magnetic Resonance Spectroscopy, Mercury, Protein Binding, Protein Conformation, Zinc}, issn = {0006-291X}, doi = {10.1016/0006-291x(75)90498-2}, author = {Smith, R J and Bryant, R G} }