Mitotic regulation by checkpoints in budding yeast

摘要 染色体分离错误是非整倍体的主要来源和推动因素 肿瘤的发展。主轴组件检查点(SAC)是 防止非整倍体，是基因组稳定性的重要调节因素。SAC是精心策划的 由动粒和一个重大悬而未决的问题，在这项建议中解决，是确定 动粒如何衡量缺乏紧张感和缺乏占有率，以及如何 动粒将这些信息传递给细胞周期机器。我们将完成高考 用Lost-of-Of-For作动粒内囊张力支的分辨率映射 功能突变体。我们已经开发出一种针对MAD3的磷酸特异性抗体，这是第一个 SAC张力分支的生化标记，我们建议用它来解剖这一点 SAC监管的重要组成部分。我们提出了一种新的角色模型 张力检查站里的动毛虫。我们将确定含有MAD3的蛋白质的亚群 是磷酸化的，其中MAD3在细胞中被磷酸化，哪些动粒蛋白 需要用来传输张力信号。我们已经分离出了针对特定基因的新突变体 SAC的入住处。我们将剖析SAC在DNA损伤中的作用 需要SAC蛋白Mec1和Tel1磷酸化的途径，代表了一种新的途径 有丝分裂调控途径。总之，这项提案中的实验将提供重要的 洞察SAC信号如何启动并整合到细胞周期中。

Abstract Errors in chromosome segregation are the major source of aneuploidy and a driving force in the development of tumors. The spindle assembly checkpoint (SAC) is one of the mechanisms that prevents aneuploidy and is an important regulator of genomic stability. The SAC is orchestrated by the kinetochore and a major unresolved problem, addressed in this proposal, is to determine how the kinetochore measures the lack of tension and lack of occupancy and how the kinetochore transfers that information to the cell cycle machinery. We will complete high resolution mapping of the tension branch of the SAC within the kinetochore using loss-of- function mutants. We have developed a phospho-specific antibody to Mad3 that is the first biochemical marker for the tension branch of the SAC and we propose to use it to dissect this important component of SAC regulation. We propose a new model for the role of the kinetochore in the tension checkpoint. We will determine if subsets of Mad3-containing proteins are phosphorylated, where Mad3 is phosphorylated in the cell and which kinetochore proteins are needed to transmit the tension signal. We have isolated novel mutants that are specific to the occupancy branch of the SAC. We will dissect the role for the SAC in the DNA damage pathway which requires Mec1 and Tel1 phosphorylation of SAC proteins and represents a novel pathway of mitotic regulation. Together, the experiments in this proposal will provide important insights into how SAC signaling is initiated and integrated into the cell cycle.