PPP1CA drives osteosarcoma tumorigenesis by promoting PKM2 dephosphorylation and tetramerization.

Osteosarcoma (OS) is a highly aggressive malignant tumor with no effective targeted therapies at present. Aerobic glycolysis represents a hallmark metabolic feature of OS, wherein the glycolytic rate-limiting enzyme pyruvate kinase M2 isozyme (PKM2) exhibits high enzymatic activity in OS cells. However, the regulatory mechanisms underlying PKM2 hyperactivation in OS remain poorly understood. PKM2 expression was measured by agarose gel electrophoresis and quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR). Proteomic analysis, coimmunoprecipitation (co-IP), and proximity ligation assay (PLA) were used to identify PKM2-interacting proteins. Protein cross-linking, blue native PAGE (BN-PAGE), and extracellular acidification rate (ECAR)/oxygen consumption rate (OCR) assays were performed to determine the effect of PPP1CA on PKM2 tetramer formation and glycolytic capacity. Functional assays were conducted in vitro and in vivo to assess the biological consequences of PPP1CA-mediated PKM2 activation on proliferation, migration, cell cycle, apoptosis, and in vivo tumor growth. Mechanistically, PPP1CA specifically binds to PKM2 and dephosphorylates it, promoting the conversion of PKM2 from dimers to tetramers. PPP1CA-mediated PKM2 tetramer activation enhanced glycolytic capacity and drove malignant behaviors including proliferation, migration, cell cycle progression, dysregulated apoptosis, and in vivo tumor growth. In clinical specimens, PPP1CA was aberrantly upregulated in OS tissues and inversely correlated with both PKM2 phosphorylation level and patient prognosis. Collectively, these results demonstrate that PPP1CA promotes PKM2 tetramerization through dephosphorylation, thereby enhancing glycolytic capacity and driving malignant progression in OS, highlighting PPP1CA as a potential therapeutic target. Implications: This study reveals that PPP1CA drives osteosarcoma progression by promoting PKM2 tetramerization and glycolysis.