Studies of the DNA Replication Checkpoint

描述（由申请人提供）：复制真核生物基因组的挑战性任务由于中断DNA复制的内在和外在因素而变得更加困难。为了应对这些困难，细胞有一个DNA复制监控系统，可以感知停滞的复制分叉并指导各种反应。其中一种反应是S-M检查点，当DNA合成(S)正在进行时，它会延迟有丝分裂(M)的开始。其他反应控制DNA复制、修复和重组。这些响应统称为复制检查点。本应用程序建议使用分裂酵母Schizosaccharomyces pombe来研究复制检查点。裂变酵母在发现和分析检查点机制方面发挥了开创性的作用，这些机制在包括人类在内的所有真核生物中都是保守的。这些研究将集中在蛋白激酶Cds1上，这是检查点的主要效应物。其中一个目标是了解Cds1是如何被调控的。这些研究将集中在Mrc1上，Mrc1是一种将复制检查点信号从传感器激酶Rad3传递到Cds1的蛋白质。第二个目标是了解如何稳定停滞的复制分叉。在这个过程中涉及的蛋白质我将被识别和分析。第三个目标是表征Cds1靶点。这些研究将集中在Mus81和Eme1上，它们是Holliday结分解酶的组成部分，可以切割x结构的DNA。所有这些基因都已证实或推测在人类中有同源物。已知其中一些同源物对维持基因组完整性和预防癌症很重要，但它们如何完成这一任务在很大程度上是未知的。预计对裂变酵母复制检查点的研究将为理解人类复制检查点的控制提供一个有价值的框架。

DESCRIPTION (provided by applicant): The challenging task of replicating a eukaryotic genome is made more difficult by intrinsic and extrinsic agents that interrupt DNA replication. To cope with these difficulties, cells have a DNA replication monitoring system that senses stalled replication forks and direct various responses. One of these responses is the S-M checkpoint that delays the onset of mitosis (M) while DNA synthesis (S) is underway. Other responses control DNA replication, repair and recombination. These responses are collectively termed the replication checkpoint. This application proposes to use the fission yeast Schizosaccharomyces pombe to investigate the replication checkpoint. Fission yeast has played a pioneering role in the discovery and analysis of checkpoint mechanisms that are conserved amongst all eukaryotes, including humans. The studies will focus on he protein kinase Cds1, a major effector of the checkpoint. One aim is to understand how Cds1 is regulated. These studies will concentrate on Mrc1, a protein then channels the replication checkpoint signal from the sensor kinase Rad3 to Cds1. The second aim is to understand how stalled replication forks are stabilized. Proteins involved in this process I will be identified and analyzed. The third aim is to characterize Cds1 targets. These studies will focus on Mus81 and Eme1, components of a Holliday junction resolvase that cleaves X-structured DNA. All of these genes have confirmed or presumptive homologs in humans. Some of these homologs are known to be important for maintaining genome integrity and preventing cancer, but how they accomplish this task is largely unknown. It is anticipated that studies of the replication checkpoint in fission yeast will provide a valuable framework for understanding replication checkpoint control in humans.