细胞周期激酶Cdk1和周期蛋白在DNA双链断裂修复中的作用

DNA double-stranded break(DSB) poses a partiular threat to genome integrity and it is closely related to human cancers. Cell cycle strictly controls the repair of DSBs yet the underlying mechanisms remain unclear. Our preliminary data show that: 1)in yeast, the cell cycle-driving kinase Cdk1 targets the chromatin remodeling factor Fun30, promoting homologous recombination and resistance to DNA damaging agents; 2) the kinase Cdk1 and multiple cyclins are recruited to DSBs, suggesting that they might directly participate in signaling or repair involving unknown functions. In light of the conservative mechanisms,in current study, we propose to investigate how DSB repair is regulated by cell cycle using the model species Sacchromyces cerevisiae. Our goal is to: 1) define Cdk1 targeting sites in Fun30 and investigate how this modification is modulated; 2) reveal the roles and molecular details of Fun30 phosphorylation in DSB repair; 3)determine the molecular basis for association of Cdk1 and cyclins to DSBs; and 4)identify novel functions of cyclins in DNA damage response and repair. In this study, we aim to examine the roles of Fun30 phosphorylation by Cdk1 as well as the functions of chromatin-associated cyclins in DSB repair. Our study will provide deep insights into the molecular details of cell cycle-regulated DSB repair and likely have implications in understanding how human genome stability is preserved.

DNA双链断裂(DSB)对基因组稳定性构成严重威胁，与人类癌症密切相关。细胞周期严格调控DNA双链断裂修复，但其机制尚不清楚。 我们前期研究发现：1）酵母细胞周期激酶Cdk1磷酸化染色质重塑蛋白Fun30，促进同源重组修复，增强对DNA损伤药物抗性；2）激酶Cdk1和多种细胞周期蛋白结合在断裂部位,暗示它们可能直接参与修复，具有未知功能。鉴于DNA修复机制的保守性，本课题以模式生物酵母为材料，研究细胞周期如何调控DSB修复，以期阐明：1）Cdk1磷酸化靶蛋白Fun30的位点及调控机制；2）Fun30磷酸化在双链断裂修复中的作用与机制；3）激酶Cdk1和细胞周期蛋白结合到损伤部位的调控及分子基础；4)细胞周期蛋白在DSB修复中的新作用。本研究旨在探索Fun30磷酸化以及细胞周期蛋白在DSB修复中的作用，从而对细胞周期调控DSB修复分子机制有全新的认识，为研究人类基因组稳定性调控提供理论借鉴。

DNA双链断裂(DSB)对基因组稳定性构成严重威胁，与人类癌症密切相关。细胞周期严格调控DNA双链断裂修复，但其机制尚不完全清楚。我们前期研究发现：1）酵母细胞周期激酶Cdk1磷酸化染色质重塑蛋白Fun30，促进同源重组修复，增强对DNA损伤药物抗性；2）激酶Cdk1和多种细胞周期蛋白结合在断裂部位,暗示它们可能直接参与修复。本课题以模式生物酵母为材料，探究了细胞周期激酶调控Fun30的生物学意义，发现：1）Cdk1在S和G2/M期磷酸化靶蛋白Fun30的Ser28位点，促进同源重组及对DNA损伤药物的抗性；2）激酶Cdk1和细胞周期蛋白结合到损伤部位并受到检查点信号调控；3）Fun30磷酸化的作用机理主要是促进其在染色质上迁移。4）泛素E3连接酶Bre1介导的H2B泛素化通过刺激组蛋白从ssDNA上解离从而促进Rad51结合及同源重组修复。本研究揭示了细胞周期在染色质水平调控DSB修复的新机制以及组蛋白泛素化在DSB修复中的作用，为研究人类基因组稳定性调控提供理论借鉴。