USP16 promotes osteosarcoma progression by stabilizing ESRRA to enhance mitochondrial oxidative phosphorylation.

Osteosarcoma is a highly aggressive primary bone malignancy, yet the underlying mechanisms driving its metabolic reprogramming remain incompletely understood. In this study, we identified ubiquitin-specific protease 16 (USP16) as a critical oncogenic driver in osteosarcoma. Bioinformatics analysis showed that USP16 was markedly upregulated in osteosarcoma tissues and predicted poor patient prognosis. Functional assays revealed that USP16 depletion significantly suppressed osteosarcoma cell proliferation and colony formation in vitro, as well as subcutaneous xenograft tumor growth in vivo, accompanied by the downregulation of cell cycle regulators. Conversely, USP16 overexpression promoted cell proliferation. Seahorse extracellular flux analysis demonstrated that USP16 depletion drastically impaired mitochondrial oxygen consumption rate (OCR) and oxidative phosphorylation (OXPHOS) parameters. Mechanistically, USP16 sustained mitochondrial respiration by deubiquitinating and stabilizing estrogen-related receptor alpha (ESRRA). Crucially, ectopic expression of ESRRA in USP16-knockout cells effectively rescued the suppressed cell proliferation and mitochondrial respiratory defects, whereas pharmacological inhibition of OXPHOS abolished USP16-induced hyperproliferation. Collectively, our findings demonstrate that USP16 drives osteosarcoma progression by orchestrating ESRRA-mediated mitochondrial OXPHOS, highlighting the USP16/ESRRA deubiquitination axis as a promising therapeutic target.