Mitochondrial complex IV subunit COX5B promotes glioblastoma progression by regulating mitochondrial function and suppressing XAF1-dependent apoptosis

Glioblastoma is notoriously invasive and resistant to programmed cell death (PCD). Mitochondria serve as the central hub integrating multiple death signals, thereby orchestrating both apoptotic and non‑apoptotic PCD. Therefore, investigating how mitochondria‑associated PCD operates in GBM becomes particularly critical for exploring potential therapeutic targets. In this study, we discovered that the mitochondria-associated programmed cell death (MPCD) gene, cytochrome c oxidase subunit 5B (COX5B)—a nuclear-encoded regulator of mitochondrial complex IV, promotes the growth of GBM by regulating mitochondrial function and inhibiting XAF1-related apoptosis. By integrating single-cell and bulk transcriptomics, we identified glioma-enriched MPCD genes and developed a machine-learning prognostic model with robust predictive performance. COX5B emerged as a central hub, upregulated in GBM, with high expression correlating with poor survival. Utilizing a multi-disciplinary approach combining immunohistochemistry (IHC), immunofluorescence (IF), quantitative PCR, western blotting, transmission electron microscopy (TEM) and flow cytometry, we demonstrate that COX5B knockdown impaired mitochondrial integrity, triggering cytosolic release of cytochrome c and mtDNA, elevating ROS, collapsing membrane potential, and causing cristae loss and organelle swelling. These mitochondrial insults activated intrinsic apoptosis via Bax/caspase 9/3 and Bcl 2 downregulation, while cytosolic mtDNA induced the JAK-STAT1/XAF1 signaling axis. XAF1 co-silencing partially rescued proliferation, migration, and apoptosis defects. In vivo, COX5B depletion suppressed xenograft growth and promoted apoptosis, effects partially reversed by XAF1 co-depletion. Our findings uncovers a novel COX5B-STAT1-XAF1 cascade linking mitochondrial dysfunction, cytochrome c/mtDNA release, and transcriptional apoptosis control, highlighting COX5B as a promising therapeutic target in GBM.