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从一个复制细胞周期到下一个复制细胞周期的基因组完整性。如果基因组发生灾难性损伤，多细胞真核生物的细胞可能会发生凋亡，大概是为了将它们从细胞群中消除，并降低遗传不稳定细胞繁殖的风险。或者，细胞可能通过细胞周期的短暂停滞来对 DNA 损伤做出反应，这与它们在暴露于 DNA 损伤剂时的存活能力相关。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，细胞将进入有丝分裂，DNA 受损并死亡。裂殖酵母是识别和表征 DNA 损伤检查点成分的极其有价值的系统。事实上，目前已知在哺乳动物细胞检查点通路中发挥作用的许多蛋白质仅根据其与酵母中遗传和功能鉴定的蛋白质的序列同源性来鉴定。因此，很明显，利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且富有成效的方法。该提案中描述的实验将继续研究蛋白激酶 Chk1，它是真核细胞检查点的关键调节因子。此外，我们将重点关注一种新型裂殖酵母蛋白 Msc1。 Msc1 与哺乳动物 RbBP2（一种通过结合肿瘤抑制蛋白 Rb 的能力而被鉴定的蛋白质）和 PLU-1（乳腺癌细胞中上调基因的产物）具有同源结构域。克隆Msc1是因为它可以弥补Chk1功能的缺失。 Msc1 蛋白对于染色体稳定性很重要。本提案中描述的实验旨在剖析 Msc1 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。当细胞的 DNA 受到损伤时，它们会延迟细胞周期进程以修复损伤。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。延迟细胞周期进展的途径称为检查点途径，了解这些途径如何发挥作用至关重要，这既可以了解 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。裂殖酵母是识别和表征 DNA 损伤检查点成分的极其有价值的系统。我们的研究利用这种生物体优良的遗传学来阐明在面临 DNA 损伤时控制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.