Ku 在电离辐射诱导的DSB修复通路选择中的作用及机制研究

DNA double strand break (DSB) induced by ionizing radiation is one of the most harmful kinds of DNA damage. DSB can be repaired by Homologous Recombination (HR) and Non Homologous End Joining (NHEJ). Since there are multiple DSB repair processes, a cell must properly choose which pathway to employ for each specific DSB. Although, much work has been performed to identify and characterize factors which are required for repair by both DSB repair pathways, the underlying molecular mechanism is still far from clear. Therefore, in the present study, we hypothesize that the modification of Ku protein plays an important role in its dissociation from DSB end and in the DSB repair pathway choice. First we will analyze whether the phosphorylation status of Ku effect its dissociation from DSB end and modulate the DSB repair way choice, next the kinase for Ku phosphorylation will be explored. To elucidate the underlying mechanism, the Ku mutations will be generated and the phosphorylation sites will be identified. Finally the protein interacts with Ku will be explored. The above study will help us elucidate the role of Ku in DSB repair pathway choice and enrich the theory of DSB repair pathway choice.

电离辐射（Ionizing Radiation）导致的DNA双链断裂（Double Strand Break, DSB）是最严重的DNA损伤类型之一。DSB主要通过同源重组（Homologous Recombination, HR）和非同源末端连接（Non Homologous End Joining, NHEJ）两条修复通路进行。研究发现有多条信号通路和多种蛋白参与DSB修复通路的选择，但其分子机制还远未明确。根据前期的实验结果，我们认为Ku蛋白的修饰在DSB修复通路选择中发挥重要作用。本研究将围绕这一问题，首先分析Ku的磷酸化是否影响其从DSB末端解聚及DSB修复通路选择；其次明确介导Ku磷酸化的激酶并构建Ku的突变株鉴定其磷酸化位点，最后鉴定与Ku发生相互作用的蛋白明确Ku磷酸化的生物学意义。通过以上研究，将明确Ku在DSB修复通路选择中的作用，进一步丰富DSB修复通路选择理论。

电离辐射（Ionizing Radiation）导致的DNA双链断裂（Double Strand Break, DSB）是最严重的DNA损伤类型之一。DSB主要通过同源重组（Homologous Recombination, HR）和非同源末端连接（Non Homologous End Joining, NHEJ）两条修复通路进行。研究发现有多条信号通路和多种蛋白参与DSB修复通路的选择，但其分子机制还远未明确。本研究首先通过激光微辐射系统明确了Ku的磷酸化影响其从DSB末端解聚，利用RPA和Rad51焦点分析Ku的磷酸化影响DSB修复通路选择；其次通过体外酶活性实验明确DNA-PKcs为介导Ku磷酸化的激酶；最后通过构建Ku的突变株鉴定其磷酸化位点可能为Ser305。通过以上研究，明确了Ku在DSB修复通路选择中的作用，进一步丰富DSB修复通路选择理论。