Pyruvate kinase M2 isoform (PKM2) catalyzes the last step of glycolysis and plays an important role in tumor cell proliferation. Recent studies have reported that PKM2 also regulates apoptosis. However, the mechanisms underlying such a role of PKM2 remain elusive. Here we show that PKM2 translocates to mitochondria under oxidative stress. In the mitochondria, PKM2 interacts with and phosphorylates Bcl2 at threonine (T) 69. This phosphorylation prevents the binding of Cul3-based E3 ligase to Bcl2 and subsequent degradation of Bcl2. A chaperone protein, HSP90α1, is required for this function of PKM2. HSP90α1's ATPase activity launches a conformational change of PKM2 and facilitates interaction between PKM2 and Bcl2. Replacement of wild-type Bcl2 with phosphorylation-deficient Bcl2 T69A mutant sensitizes glioma cells to oxidative stress-induced apoptosis and impairs brain tumor formation in an orthotopic xenograft model. Notably, a peptide that is composed of the amino acid residues from 389 to 405 of PKM2, through which PKM2 binds to Bcl2, disrupts PKM2-Bcl2 interaction, promotes Bcl2 degradation and impairs brain tumor growth. In addition, levels of Bcl2 T69 phosphorylation, conformation-altered PKM2 and Bcl2 protein correlate with one another in specimens of human glioblastoma patients. Moreover, levels of Bcl2 T69 phosphorylation and conformation-altered PKM2 correlate with both grades and prognosis of glioma malignancy. Our findings uncover a novel mechanism through which mitochondrial PKM2 phosphorylates Bcl2 and inhibits apoptosis directly, highlight the essential role of PKM2 in ROS adaptation of cancer cells, and implicate HSP90-PKM2-Bcl2 axis as a potential target for therapeutic intervention in glioblastoma.

Background Non-small cell lung cancer (NSCLC) cells derived intracellular and extracellular programmed cell death ligand 1 (PD-L1) promoted cancer progression and drug resistance, and facilitated tumor immune evasion. However, the detailed molecular mechanisms are still largely unknown. In the present study, we aimed to explore the role of circular RNA circ-CPA4/let-7 miRNA/PD-L1 axis in the regulation of NSCLC progression, drug resistance and tumor immune microenvironment. Methods Real-Time qPCR and Western Blot analysis were conducted to examine gene expressions at transcriptional and translated levels, respectively. The regulatory mechanisms of circ-CPA4, let-7 miRNA and PD-L1 were validated by dual-luciferase reporter gene system and RNA pull-down assay. Cell growth and apoptosis were determined by CCK-8 assay, colony formation assay and Annexin V-FITC/PI double staining assay. Cell mobility was evaluated by transwell assay. Results Circ-CPA4 and PD-L1 were high-expressed, while let-7 miRNA was low-expressed in NSCLC cells and cancer tissues compared to the human bronchial epithelial (HBE) cells and their paired clinical normal adjacent tissues, respectively. Besides, knock-down of circ-CPA4 inhibited cell growth, mobility and epithelial-mesenchymal transition (EMT), and promoted cell death in NSCLC cells by downregulating PD-L1 through serving as a RNA sponge for let-7 miRNA. In addition, the NSCLC cells derived PD-L1-containing exosomes promoted cell stemness and increased resistance of NSCLC cells to cisplatin. Notably, by co-culturing the NSCLC cells with CD8 + T cells isolated from human peripheral blood mononuclear cells (hPBMCs) in a transwell co-culturing system, we found that NSCLC cells inactivated CD8 + T cells in a secreted PD-L1-dependent manner. Further results suggested that circ-CPA4 also positively regulated exosomal PD-L1, and the NSCLC cells with circ-CPA4 ablation re-activated CD8 + T cells in the co-culturing system. Conclusion Taken together, circ-CPA4 regulated cell growth, mobility, stemness and drug resistance in NSCLC cells and inactivated CD8 + T cells in the tumor immune microenvironment through let-7 miRNA/PD-L1 axis.