Cell Cycle Checkpoint Control in Response to DNA Damage

描述（由申请方提供）：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。在基因组发生灾难性损伤的情况下，多细胞真核生物的细胞可以经历凋亡，大概是为了将它们从细胞群中消除并降低繁殖遗传不稳定细胞的风险。或者，细胞可以通过经历细胞周期的短暂停滞来响应DNA损伤，这与它们暴露于DNA损伤剂而存活的能力相关。这种反应需要DNA损伤检查点途径;如果受到突变或药物治疗的影响，细胞将进入DNA受损的有丝分裂并死亡。裂殖酵母是一个非常有价值的系统，用于鉴定和表征DNA损伤检查点的组分。事实上，现在已知在哺乳动物细胞中的检查点途径中起作用的许多蛋白质仅基于它们与在酵母中遗传和功能上鉴定的蛋白质的序列同源性来鉴定。因此，很明显，使用经典遗传学的力量鉴定酵母中的蛋白质是鉴定和深入了解哺乳动物对应物功能的有效和富有成效的手段。本提案中描述的实验将继续研究蛋白激酶Chk 1，这是真核细胞中检查点的关键调节因子。此外，我们将重点关注一种新的裂变酵母蛋白，Msc 1。MSC 1与哺乳动物RbBP 2和PLU-1共享结构域同源性，RbBP 2是一种通过结合肿瘤抑制蛋白Rb的能力而鉴定的蛋白质，PLU-1是在乳腺癌细胞中上调的基因的产物。克隆MSC 1是因为它可以补偿Chk 1功能的丧失。Msc 1蛋白对染色体稳定性很重要。本提案中描述的实验旨在剖析Msc 1蛋白在裂殖酵母中的作用，为理解人类同源物的功能奠定基础。当细胞经历DNA损伤时，它们会延迟细胞周期进程以修复损伤。损伤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，了解这些途径如何工作是至关重要的，既要了解DNA损伤如何导致细胞死亡，又要确定增强化疗药物杀伤的方法。裂殖酵母是一个非常有价值的系统，用于鉴定和表征DNA损伤检查点的组分。我们的研究利用这种生物体的优良遗传学来阐明在DNA损伤面前管理细胞分裂的普遍重要机制。

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.