Regulation of Chromosome Segregation

描述（由申请人提供）：细胞分裂的完美执行对所有生物的产生和生存至关重要。在每个细胞周期中，染色体必须准确地划分到子细胞中，以防止基因组不稳定和非整倍体，这是所有肿瘤和许多出生缺陷的标志。我们的目标是阐明确保染色体准确分离的机制，从而有助于理解人类疾病的基础。我们正在研究出芽酵母的染色体分离，因为它适合遗传学和生化分析，而且染色体分离的机制基本上是保守的。染色体分离需要重复染色体的着丝点（姐妹染色单体）的双取向，这样它们就可以与来自相反纺锤极的微管（mt）结合。虽然每一对姐妹着丝点都有生物定向的mt -着丝点连接是必不可少的，但建立、维持和纠正双定向错误的机制仍然在很大程度上是未知的。唯一已知的对着丝粒双向定向至关重要的蛋白质是保守的IpH （Aurora B）蛋白激酶，其定位和活性受sim5 （INCENP）蛋白调控，并受到蛋白磷酸酶i的催化亚基Glc7的反对。我们已经确定了其他可能调节双向定向的基因，因为它们在IpM功能受损时变得至关重要。因此，我们将描述这些基因在双性取向中的作用，并确定它们用于调节双性取向的机制。虽然IpH和Glc7对着丝点的双向定向至关重要，但这些酶的分子靶点很少被发现。因此，我们开发了一种纯化着丝点的方法，这将使我们能够识别与这些酶和双取向相关的翻译后修饰。最后，我们将采用互补的遗传和生化方法来了解IpH和Glc7的活动如何在空间和时间上协调，以确保检测和纠正双取向缺陷。综上所述，这些研究将有助于更好地理解所有真核生物的染色体分离和基因组稳定性的维持。

DESCRIPTION (provided by applicant): The flawless execution of cell division is essential to the generation and survival of all organisms. During every cell cycle, chromosomes must be accurately partitioned to daughter cells to prevent genomic instability and aneuploidy, a hallmark of all tumors and many birth defects. Our goal is to elucidate the mechanisms that ensure accurate chromosome segregation and therefore contribute to understanding the basis of human disease. We are studying chromosome segregation in budding yeast because it is amenable to both genetic and biochemical analyses and the mechanism of chromosome segregation is fundamentally conserved. Chromosome segregation requires that the kinetochores of duplicated chromosomes (sister chromatids) biorient such that they bind to microtubule (MTs) arising from opposite spindle poles. Although it is essential that every pair of sister kinetochores make bioriented MT-kinetochore attachments, the mechanisms that establish, maintain, and correct errors in biorientation are still largely unknown. The only protein known to be essential for kinetochore biorientation is the conserved IpH (Aurora B) protein kinase whose localization and activity is regulated by the SIM 5 (INCENP) protein and opposed by Glc7, the catalytic subunit of protein phosphatase I. We have identified additional genes that likely regulate biorientation because they become essential when IpM function is impaired. We will therefore characterize the role of these genes in biorientation and identify the mechanisms that they use to regulate biorientation. Although IpH and Glc7 are critical for kinetochore biorientation, few molecular targets of these enzymes have been identified. We have therefore developed a method to purify kinetochores that will allow us to identify post-translational modifications associated with these enzymes and biorientation. Finally, we will take complementary genetic and biochemical approaches to understand how the activities of IpH and Glc7 are spatially and temporally coordinated to ensure that biorientation defects are detected and corrected. Taken together, these studies should lead to a better understanding of chromosome segregation and the maintenance of genomic stability in all eukaryotes.