Induction Of Autophagy Reveals A Cytoprotective Mechanism By Which Dna Polymerase Gamma (Pol-Gamma)Prevents Uvb-Induced Skin Cancer

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PADNA polymerase γ (polγ), a major mitochondrial DNA repair enzyme, is essential for replication and repair of the mitochondrial genome. It is well-established that mitochondrial genomic stability is important for healthy cell survival; however, whether and how polγ plays a role in autophagic response remain unknown. Our previous studies demonstrated that polγ is inactivated upon exposure to ultraviolet B (UVB) irradiation by a peroxynitrite-mediated mechanism. In this study, we identified tyrosine 964, which is near the active site of polγ, as the target of nitration-induced inactivation by the use of LC/MS/MS mass spectroscopy coupled to immunoprecipitation, Western blotting and a 3` to 5` exonuclease activity assay. Treatment of mouse skin epithelial cells with UVB radiation validated the presence of polγ nitration and reduction of its enzymatic activity in vivo. To determine the effect of UVB in autophagic responses, mouse skin epithelial (JB6) cells were transfected with GFP-labeled LC-3 cDNA and exposed to UVB radiation, or wild-type C57BL/6 mice were exposed to UVB radiation and examined for LC-3 cleavage products. UVB radiation of JB6 cells and mouse skin increased LC-3 cleavage products and beclin levels, suggesting increased autophagic response to UVB treatment both in vitro and in vivo. To elucidate the role of Polγ in autophagic responses, the expression of polγ in JB6 cells was inhibited by siRNA for polγ. Consistent with the role of polγ in preventing UVB-induced autophagy, knockdown of polγ led to an increase LC-3 cleavage products. Knockdown of polγ induced prosurvival autophagic responses via the mTOR-signaling-complex-1(mTORC1) and complex-2(mTORC2). Interestingly, the mTORC2 complex plays a prominent role in the prosurvival autophagy responses based on the bromodeoxyuridine uptake assay, following knockdown and overexpression approaches. Importantly, suppression of polγ alone leads to transformation and invasion ofJB6 cells, which are increased following UVB treatment. The results reveal a unique role of Polγ in autophagy and support the role of Polγ in the suppression of prosurvival autophagic response to UVB- mediated carcinogenesis.Citation Format: Sanjit K. Dhar, Vasu Bakthavachalu, Tadahide Izumi, Jing Chen, Haining Zhu, Daret K. St. Clair. Induction of autophagy reveals a cytoprotective mechanism by which DNA polymerase gamma (pol-γ) prevents UVB-induced skin cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2904. doi:10.1158/1538-7445.AM2015-2904