miR-516b-5p靶向ATM调控鼻咽癌细胞HONE1DNA修复的机制研究

目的 探讨鼻咽癌细胞HONE1中抑制DNA修复的潜在机制.方法 使用ETP处理鼻咽癌细胞HONE1造成DNA损伤后,通过高通量测序检测鼻咽癌细胞HONE1中miRNA的表达谱.过表达差异miRNA后,通过基于I-PpoI的可诱导DSB系统检测DNA双链断裂处(DNA double-strand breaks,DSB)的...

Study on the Mechanism of miR-516b-5p Targeting ATM to Regulate DNA Repair in Nasopharyngeal Carcinoma Cells HONE1

Objective To explore the potential mechanism of inhibiting DNA repair in nasopharyngeal carcinoma cells HONE1.Methods After ETP treatment of nasopharyngeal carcinoma cells HONE1 caused DNA damage,the expression profile of miRNA in nasopharyngeal carcinoma cells HONE1 was detected by high-throughput sequencing.After over-expression of differential miRNA,the DNA-PKcs content of DNA double-strand breaks(DNA double-strand breaks,DSB)was detected by the inducible DSB system based on I-PpoI.Analyze potential targets of miRNA online through miRDB.Detect miRNA targeted mRNA by luciferase reporter system.Results hsa-miR-9983-3p,hsa-miR-4731-5p,hsa-miR-3675-5p,hsa-miR-516b-5p,hsa-miR-4437,hsa-miR-7114-5p after ETP treatment,The expression level of hsa-miR-637,hsa-miR-5008-5p increased at least 8 times.miR-516b-5p mimics caused a decrease in DSB DNA-PKcs content(P<0.05),while miR-516b-5p inhibitor caused an increase in DSB DNA-PKcs content(P<0.05).After miR-516b-5p was overexpressed in nasopharyngeal carcinoma cell HONE1,the expression level of ATM mRNA and protein expression decreased,and the phosphorylation level of DNA-PKcs decreased.The expression of mir-516b-5p was inhibited,the expression of ATM mRNA decreased.The expression level and protein expression level increased,and the phosphorylation level of DNA-PKcs increased.At the same time,miR-516b-5p targets the 3 terminal non-coding region of ATM.After overexpression of ATM,the DNA-PKcs content of DSB increased(P<0.05),while after knocking down ATM,the DNA-PKcs content of DSB decreased(P<0.05).Conclusion miR-516b-5p degrades ATM mRNA and inhibits ATM translation by targeting the 3 non-coding region of ATM mRNA.Then the phosphorylation level of DNA-PKcs decreases,and the binding of DNA-PKcs to DSB decreases,and finally Inhibited DNA repair of nasopharyngeal carcinoma cells HONE1.