Elongator Complex Play A Key Role In DNA Damage Induced By ⁶⁰Co γ Ray And Can Specifically Enhance The DNA Repair Ability In Hepatocellular Carcinoma Cells

Objective:To explore the relationship between Elongator complex and DNA damage induced by 60Co γ ray,and the molecular mechanism of Elongator with respect to DNA damage and repair in hepatocellular carcinoma cells.Methods:1. The gene expression of endogenous elp3 and elp4 in tumor cell lines was detected by RT-PCR.2. With OTM as an indicator, after the irradiation of 60Co γ ray,the change of DNA damage in different groups was determined by comet assay.3. With γ-H2AX as a symbol, the extent of DNA damage and the protein expression in different groups were assessed by immunofluorescence(IF) and western blot.4. Cell apoptosis rate in each cell line was measured by flow cytometry.5. Real-time PCR was used to test the expression of the gene related to DNA damage and repair.6. Western blot was applied to confirm the protein expression with regard to DNA damage and repair.Results:1. RT-PCR showed that relative mRNA level of endogenous elp3 and elp4 has no significant change in tumor cell lines.2. Elongator can reduce the extent of DNA damage in HepG2 cells and SK-Hep-1 cells detected by comet assay,and the results indicated Elongator was required in DNA damage induced by 60Co γ ray.3. Elongator can decrease the extent of DNA damage measured by IF and western blot,and these results were consistent with the conclusion of comet assay, namely Elongator can specifically enhance the DNA repair ability.4. Flow cytometry showed that Elongator can lower cell apoptosis rate in each cell line,and the result explained Elongator can maintain genome stability.5. Real-time PCR and western blot showed that the expression of RAD50,Mrell,Nbs1,DNAPK-cs and XRCC6 was related to Elongator in DNA damage and repair.Conclusion:1. In vitro, after the irradiation of 60Co y ray,Elongator complex can reduce the extent of DNA damage and can decrease cell apoptosis rate in each cell line. Elongator can maintain genome stability and play an important role in DNA damage and repair.2. The role of Elongator complex may act through some genes related to DNA damage and repair such as RAD50、Mre11、 Nbs1、DNAPK-cs、XRCC6.Therefore, we may arrive at a conclusion that Elongator will promote cell survival by specifically enhancing its DNA repair ability in hepatocellular carcinoma cells.