DNA repair protein DNA-PK protects PC12 cells from oxidative stress-induced apoptosis involving AKT phosphorylation

Background Emerging evidence suggest that DNA-PK complex plays a role in the cellular response to oxidative stress, in addition to its function of double strand break (DSB) repair. In this study we evaluated whether DNA-PK participates in oxidative stress response and whether this role is independent of its function in DNA repair. Methods and results We used a model of H2O2-induced DNA damage in PC12 cells (rat pheochromocytoma), a well-known neuronal tumor cell line. We found that H2O2 treatment of PC12 cells induces an increase in DNA-PK protein complex levels, along with an elevation of DNA damage, measured both by the formation of gamma Eta 2 Alpha X foci, detected by immunofluorescence, and gamma H2AX levels detected by western blot analysis. After 24 h of cell recovery, gamma Eta 2 Alpha X foci are repaired both in the absence and presence of DNA-PK kinase inhibitor NU7026, while an increase of apoptotic cells is observed when DNA-PK activity is inhibited, as revealed by counting pycnotic nuclei and confirmed by FACS analysis. Our results suggest a role of DNA-PK as an anti-apoptotic factor in proliferating PC12 cells under oxidative stress conditions. The anti-apoptotic role of DNA-PK is associated with AKT phosphorylation in Ser473. On the contrary, in differentiated PC12 cells, were the main pathway to repair DSBs is DNA-PK-mediated, the inhibition of DNA-PK activity causes an accumulation of DNA damage. Conclusions Taken together, our results show that DNA-PK can protect cells from oxidative stress induced-apoptosis independently from its function of DSB repair enzyme.

Automated summary

The study reveals that DNA-PK complex may play a role in cellular response to oxidative stress independently from its function in DNA repair. It acts as an anti-apoptotic factor in proliferating PC12 cells under oxidative stress conditions, but inhibition of DNA-PK activity in differentiated PC12 cells leads to accumulation of DNA damage.