Studies of the Rad60-Smc5-Smc6 DNA Repair Complex

多条DNA修复途径已经进化，以在不断接二连三的遗传毒性物质面前保持基因组的完整性。各种DNA修复机制的缺陷会导致癌症的强烈易感性。除了DNA修复途径外，复制检查点也起着重要作用。作为对S期胁迫的响应，复制检查点通过协调细胞周期停滞、分叉稳定和DNA修复等细胞反应来维持基因组的稳定。复制检查点所需的人类蛋白质突变会增加癌症风险。这项提议旨在探索DNA修复和复制检查点之间的重要接口，人们对此知之甚少。这些研究将使用裂解酵母，裂殖酵母已被证明在分析人类保守的DNA修复和检查点途径方面具有非常宝贵的价值。DNA修复蛋白Rad60与复制检查点激酶CDSL相互作用。RAD60是DNA修复和基因组完整性所必需的Smc5-Smc6复合体的一部分。Rad60受CDSL依赖的调控，因此，第一个目标是解偶联CDSL-Rad60通讯并研究其对基因组稳定性的影响。第二个目标集中在了解Rad60的功能，这反过来应该揭示CDSL维持基因组的重要机制。第三个目的是探索Smc5-Smc6复合体的功能，Rad60是其中的一部分。Smc5-Smc6复合体可能在染色体结构中发挥作用，从而促进DNA修复。人们对染色质结构在DNA修复中的作用知之甚少，因此这些研究将有助于阐明这种联系。这项研究的结果可能会对上述蛋白质的人类同源物进行类似的研究，从而更好地了解复制检查点和人类DNA修复之间的接口。

Multiple DNA repair pathways have evolved to maintain genomic integrity in the face of a constant barrage of genotoxic agents. Defects in various DNA repair mechanisms result in a strong predisposition to cancer. In addition to DNA repair pathways, the replication checkpoint plays an important role. In response to S-phase insults, the replication checkpoint maintains genomic stability by coordinating cellular responses such as; cell-cycle arrest, fork stabilization and DNA repair. Cancer risk is elevated by mutations in human proteins that are required for the replication checkpoint. This proposal is aimed at exploring the important interface between DNA repair and the replication checkpoint, about which little is known. These studies will employ the fission yeast, Schizosaccharomyces pombe, which has proven invaluable in the analysis of DNA repair and checkpoint pathways that are conserved in humans. The DNA repair protein Rad60 interacts with the replication checkpoint kinase, Cdsl. Rad60 is part of the essential Smc5-Smc6 complex that is required for DNA repair and genomic integrity. Rad60 is subject to Cdsl-dependent regulation and therefore, the first aim is to uncouple Cdsl-Rad60 communication and study the effects on genomic stability. The second aim focusses on understanding the function of Rad60, which in turn should reveal an important mechanism through which Cdsl maintains the genome. The third aim is to explore the function of the Smc5-Smc6 complex, of which Rad60 is a part. The Smc5-Smc6 complex likely plays a role in chromosome structure and thereby facilitates DNA repair. Little is known about the role of chromatin structure in DNA repair and therefore these studies will help elucidate this connection. Results obtained in this study will likely prime analogous studies of the human homologues of the above proteins, resulting in a better understanding of the interface between the replication checkpoint and DNA repair in humans.