Inorganic polyphosphate (polyP) is primarily synthesized by Polyphosphate Kinase-1 (PPK-1) and regulates numerous cellular processes, including energy metabolism, stress adaptation, drug tolerance, and microbial pathogenesis. Here, we report that polyP interacts with acyl CoA carboxylases, enzymes involved in lipid biosynthesis in . We show that deletion of in results in transcriptional and metabolic reprogramming. In comparison to the parental strain, the Δ mutant strain had reduced levels of virulence-associated lipids such as PDIMs and TDM. We also observed that polyP deficiency in is associated with enhanced phagosome-lysosome fusion in infected macrophages and attenuated growth in mice. Host RNA-seq analysis revealed decreased levels of transcripts encoding for proteins involved in either type I interferon signaling or formation of foamy macrophages in the lungs of Δ mutant-infected mice relative to parental strain-infected animals. Using target-based screening and molecular docking, we have identified raloxifene hydrochloride as a broad-spectrum PPK-1 inhibitor. We show that raloxifene hydrochloride significantly enhanced the activity of isoniazid, bedaquiline, and pretomanid against in macrophages. Additionally, raloxifene inhibited the growth of in mice. This is an in-depth study that provides mechanistic insights into the regulation of mycobacterial pathogenesis by polyP deficiency.