Cell Cycle Checkpoint Control in Response to DNA Damage

描述（由申请人提供）：细胞周期检查点确保基因组从一个复制细胞周期到下一个复制细胞周期的完整性。在基因组遭受灾难性损伤的情况下，多细胞真核生物的细胞可以发生凋亡，可能是为了将它们从细胞群中清除，并降低遗传不稳定细胞繁殖的风险。另外，细胞可能通过经历细胞周期的短暂停止来对DNA损伤作出反应，这与它们暴露于DNA损伤剂中的生存能力有关。这种反应需要DNA损伤检查点通路；如果受到突变或药物治疗的损害，细胞将带着受损的DNA进入有丝分裂并死亡。裂变酵母已经成为鉴定和表征DNA损伤检查点组分的极有价值的系统。事实上，现在已知的许多在哺乳动物细胞中的检查点通路中起作用的蛋白质，仅仅是基于它们与酵母中基因和功能上鉴定的蛋白质的序列同源性来鉴定的。因此，很明显，利用经典遗传学的力量来鉴定酵母中的蛋白质是一种有效的、富有成效的方法，可以识别和深入了解哺乳动物对应蛋白的功能。本提案中描述的实验将继续研究真核细胞中检查点的关键调节因子Chk1蛋白激酶。此外，我们将重点研究一种新的裂变酵母蛋白，Msc1。Msc1与哺乳动物RbBP2具有相同的结构域，RbBP2是一种通过结合肿瘤抑制蛋白Rb和plus -1的能力而被发现的蛋白质，plus -1是乳腺癌细胞中上调的基因产物。克隆Msc1是因为它可以弥补Chk1功能的缺失。Msc1蛋白对染色体的稳定性很重要。本实验旨在剖析Msc1蛋白在裂变酵母中的作用，为理解人类同源物的功能奠定基础。当细胞的DNA受到损伤时，它们会延迟细胞周期的进程以修复损伤。破坏DNA的药物被用于癌症治疗，因为它们倾向于优先杀死快速分裂的细胞，这是许多类型癌症的共同特征。延迟细胞周期进程的途径被称为检查点途径，了解这些途径的工作原理至关重要，既可以了解DNA损伤如何导致细胞死亡，也可以确定增强化疗药物杀伤的方法。裂变酵母已经成为鉴定和表征DNA损伤检查点组分的极有价值的系统。我们的研究利用这种生物的优良遗传学来阐明在面对DNA损伤时对控制细胞分裂普遍重要的机制。

DESCRIPTION (provided by applicant): Cell cycle checkpoints ensure the integrity of the genome from one replicative cell cycle to the next. In the event of catastrophic damage to the genome, cells of multicellular eukaryotes can undergo apoptosis, presumably to eliminate them from the cell population and reduce the risk of propagating genetically unstable cells. Alternatively, cells may respond to DNA damage by undergoing a transient arrest of the cell cycle, which correlates with their ability to survive exposure to DNA damaging agents. This response requires the DNA damage checkpoint pathway; if compromised by mutation or drug treatment, cells will enter mitosis with damaged DNA and die. The fission yeast has been an extremely valuable system for identifying and characterizing components of the DNA damage checkpoint. Indeed, many proteins that are now known to function in the checkpoint pathway in mammalian cells were identified solely based on their sequence homology to proteins that were identified genetically and functionally in yeast. Thus, it is clear that the identification of proteins in yeast using the power of classical genetics is a valid and productive means of identifying and gaining insight into the function of mammalian counterparts. Experiments described in this proposal will continue to investigate the protein kinase Chk1, a key regulator of the checkpoint in eukaryotic cells. In addition, we will focus on a novel fission yeast protein, Msc1. Msc1 shares structural domain homology with mammalian RbBP2, a protein identified by virtue of its ability to bind the tumor suppressor protein Rb and with PLU-1, the product of a gene that is up regulated in breast cancer cells. Msc1 was cloned because it can compensate for the loss of function Chk1. The Msc1 protein is important for chromosome stability. Experiments described in this proposal aim to dissect the role of the Msc1 protein in fission yeast, to lay the groundwork for understanding the functions of the human homologues. When cells experience damage to their DNA, they delay cell cycle progression in order to repair the damage. Agents that damage DNA are used in therapy of cancer because they tend to preferentially kill cells that are rapidly dividing, a feature common to many types of cancer. Pathways that delay cell cycle progression are called checkpoint pathways and it is critical to understand how these pathways work, both to appreciate how DNA damage brings about cell death and to identify ways in which to enhance killing by chemotherapeutic agents. The fission yeast has been an extremely valuable system for identifying and characterizing components of the DNA damage checkpoint. Our studies make use of the excellent genetics of this organism to elucidate mechanisms that are universally important for governing cell division in the face of DNA damage.