Synthesis of cytochrome c oxidase 2: a p53-dependent metabolic regulator that promotes respiratory function and protects glioma and colon cancer cells from hypoxia-induced cell death

C Wanka, D P Brucker, O Bähr, M Ronellenfitsch, M Weller, J P Steinbach, J Rieger
2012
P53 has an important role in the processing of starvation signals. P53-dependent molecular mediators of the Warburg effect reduce glucose consumption and promote mitochondrial function. We therefore hypothesized that the retention of wild-type p53 characteristic of primary glioblastomas limits metabolic demands induced by deregulated signal transduction in the presence of hypoxia and nutrient depletion. Here we report that short hairpin RNA-mediated gene suppression of wild-type p53 or ectopic
more » ... ype p53 or ectopic expression of mutant temperature-sensitive dominant-negative p53(V135A) increased glucose consumption and lactate production, decreased oxygen consumption and enhanced hypoxia-induced cell death in p53 wild-type human glioblastoma cells. Similarly, genetic knockout of p53 in HCT116 colon carcinoma cells resulted in reduced respiration and hypersensitivity towards hypoxia-induced cell death. Further, wild-type p53 gene silencing reduced the expression of synthesis of cytochrome c oxidase 2 (SCO2), an effector necessary for respiratory chain function. An SCO2 transgene reverted the metabolic phenotype and restored resistance towards hypoxia in p53-depleted and p53 mutant glioma cells in a rotenone-sensitive manner, demonstrating that this effect was dependent on intact oxidative phosphorylation. Supplementation with methyl-pyruvate, a mitochondrial substrate, rescued p53 wild-type but not p53 mutant cells from hypoxic cell death, demonstrating a p53-mediated selective aptitude to metabolize mitochondrial substrates. Further, SCO2 gene silencing in p53 wild-type glioma cells sensitized these cells towards hypoxia. Finally, lentiviral gene suppression of SCO2 significantly enhanced tumor necrosis in a subcutaneous HCT116 xenograft tumor model, compatible with impaired energy metabolism in these cells. These findings demonstrate that glioma and colon cancer cells with p53 wild-type status can skew the Warburg effect and thereby reduce their vulnerability towards tumor hypoxia in an SCO2-dependent manner. Targeting SCO2 may therefore represent a valuable strategy to enhance sensitivity towards hypoxia and may complement strategies targeting glucose metabolism. Rieger, J (2012). Synthesis of cytochrome c oxidase 2: a p53-dependent metabolic regulator that promotes respiratory function and protects glioma and colon cancer cells from hypoxia-induced cell death. Oncogene, 31(33):3764-3776. Abbreviations: AMPKα, AMP-activated protein kinase α; ACC, actetyl coA carboxylase; COX, cytochrome c oxidase; EGFR, epidermal growth factor receptor; 4-EBP1, 4E binding protein 1; GLUT1, glucose transporter 1; hygro, hygromycin; luc, luciferase; p53ts, temperature-sensitive p53; PI, propidium iodide; PI3K, phosphatidylinositol-3 kinase; PKB/Akt, protein kinase B; phospho, phosphorylated; puro, puromycin; ROS, reactive oxygen species; SCO2, Synthesis 2 of Cytochrome c Oxidase 2; shRNA, short hairpin RNA; siRNA, small interfering RNA; TIGAR, tp53 inducer and regulator of glycolysis, wt, wildtype. Abstract p53 has an important role in the processing of starvation signals. p53-dependent molecular mediators of the Warburg effect reduce glucose consumption and promote mitochondrial function. We therefore hypothesized that the retention of wild-type p53 characteristic of primary glioblastomas limits metabolic demands induced by deregulated signal transduction in the presence of hypoxia and nutrient depletion. Here we report that shRNA-mediated gene suppression of wild-type p53 or ectopic expression of mutant temperature-sensitive dominantnegative p53 V135A increased glucose consumption and lactate production, decreased oxygen consumption and enhanced hypoxia-induced cell death in p53 wild-type human glioblastoma cells. Similarly, genetic knock-out of p53 in HCT116 colon carcinoma cells resulted in reduced respiration and hypersensitivity towards hypoxia-induced cell death. Further, wildtype p53 gene silencing reduced the expression of Synthesis of Cytochrome c Oxidase 2 (SCO2), an effector necessary for respiratory chain function. A SCO2 transgene reverted the metabolic phenotype and restored resistance towards hypoxia in p53-depleted and in p53 mutant glioma cells in a rotenone-sensitive fashion, demonstrating that this effect was dependent on intact oxidative phosphorylation. Supplementation with methyl-pyruvate, a mitochondrial substrate, rescued p53 wild-type but not p53 mutant cells from hypoxic cell death, demonstrating a p53-mediated selective aptitude to metabolize mitochondrial substrates. Further, SCO2 gene silencing in p53 wild-type glioma cells sensitized these cells towards hypoxia. Finally, lentiviral gene suppression of SCO2 significantly enhanced tumor necrosis in a subcutaneous HCT 116 xenograft tumor model, compatible with impaired energy metabolism in these cells. These findings demonstrate that glioma and colon cancer cells with p53 wild-type status can skew the Warburg effect and thereby reduce their 3 vulnerability towards tumor hypoxia in a SCO2-dependent fashion. Targeting SCO2 may therefore represent a valuable strategy to enhance sensitivity towards hypoxia and may complement strategies targeting glucose metabolism.
doi:10.5167/uzh-64674 fatcat:27ya62jzzzb5nesagaylmjnyau