CDK1-Mediated SIRT3 Activation Enhances Mitochondrial Function and Tumor Radioresistance.

Tumor adaptive resistance to therapeutic radiation remains a barrier for further improvement of local cancer control. SIRT3, a member of the sirtuin family of NAD(+)-dependent protein deacetylases in mitochondria, promotes metabolic homeostasis through regulation of mitochondrial protein deacetylation and plays a key role in prevention of cell aging. Here, we demonstrate that SIRT3 expression is induced in an array of radiation-treated human tumor cells and their corresponding xenograft tumors, including colon cancer HCT-116, glioblastoma U87, and breast cancer MDA-MB231 cells. SIRT3 transcriptional activation is due to SIRT3 promoter activation controlled by the stress transcription factor NF-κB. Posttranscriptionally, SIRT3 enzymatic activity is further enhanced via Thr150/Ser159 phosphorylation by cyclin B1-CDK1, which is also induced by radiation and relocated to mitochondria together with SIRT3. Cells expressing Thr150Ala/Ser159Ala-mutant SIRT3 show a reduction in mitochondrial protein lysine deacetylation, Δψm, MnSOD activity, and mitochondrial ATP generation. The clonogenicity of Thr150Ala/Ser159Ala-mutant transfectants is lower and significantly decreased under radiation. Tumors harboring Thr150Ala/Ser159Ala-mutant SIRT3 show inhibited growth and increased sensitivity to in vivo local irradiation. These results demonstrate that enhanced SIRT3 transcription and posttranslational modifications in mitochondria contribute to adaptive radioresistance in tumor cells. CDK1-mediated SIRT3 phosphorylation is a potential effective target to sensitize tumor cells to radiotherapy.

CDK4-mediated MnSOD activation and mitochondrial homeostasis in radioadaptive protection.

Mammalian cells are able to sense environmental oxidative and genotoxic conditions such as the environmental low-dose ionizing radiation (LDIR) present naturally on the earth's surface. The stressed cells then can induce a so-called radioadaptive response with an enhanced cellular homeostasis and repair capacity against subsequent similar genotoxic conditions such as a high dose radiation. Manganese superoxide dismutase (MnSOD), a primary mitochondrial antioxidant in mammals, has long been known to play a crucial role in radioadaptive protection by detoxifying O2(•-) generated by mitochondrial oxidative phosphorylation. In contrast to the well-studied mechanisms of SOD2 gene regulation, the mechanisms underlying posttranslational regulation of MnSOD for radioprotection remain to be defined. Herein, we demonstrate that cyclin D1/cyclin-dependent kinase 4 (CDK4) serves as the messenger to deliver the stress signal to mitochondria to boost mitochondrial homeostasis in human skin keratinocytes under LDIR-adaptive radioprotection. Cyclin D1/CDK4 relocates to mitochondria at the same time as MnSOD enzymatic activation peaks without significant changes in total MnSOD protein level. The mitochondrial-localized CDK4 directly phosphorylates MnSOD at serine-106 (S106), causing enhanced MnSOD enzymatic activity and mitochondrial respiration. Expression of mitochondria-targeted dominant negative CDK4 or the MnSOD-S106 mutant reverses LDIR-induced mitochondrial enhancement and adaptive protection. The CDK4-mediated MnSOD activation and mitochondrial metabolism boost are also detected in skin tissues of mice receiving in vivo whole-body LDIR. These results demonstrate a unique CDK4-mediated mitochondrial communication that allows cells to sense environmental genotoxic stress and boost mitochondrial homeostasis by enhancing phosphorylation and activation of MnSOD.