Protection of Centromeric Cohesion by Bub1 and Sgo1

描述（由申请人提供）：生物体的遗传稳定性取决于姐妹染色单体在有丝分裂过程中准确地分裂成两个子细胞，这反过来要求姐妹染色单体之间保持物理联系（内聚），直到它们与有丝分裂纺锤体相连接。从染色体上去除内聚蛋白（维持姐妹染色单体内聚的蛋白质复合体）会导致姐妹染色单体分离。在脊椎动物细胞中，内聚蛋白的清除分两个步骤进行。在前期，沿染色体臂的大部分黏结蛋白通过polo样激酶(Plk1)/Aurora b依赖的黏结蛋白磷酸化被去除。在中期，残余的着丝粒内聚蛋白池被分离酶分裂，使姐妹染色单体分离。我实验室的长期目标是了解哺乳动物细胞中染色体分离的分子机制。在本提案中，我们将解决这一领域的一个有趣的难题：着丝粒内聚蛋白是如何在前期屏蔽Plk1 /Aurora B的作用的？我们最近提供的证据表明纺锤体检查点激酶Bub1和Sgo1着丝粒蛋白和蛋白磷酸酶2A （PP2A）靶向着丝粒，在那里它们抵消Plk1和其他有丝分裂激酶对黏结蛋白的磷酸化。在具体目标1和2中，我们将进一步描述Bub1、Sgo1和PP2A协同保护着丝粒内聚的机制。另一方面，我们的结果也指出Sgo1的作用不依赖于pp2a。事实上，在过去的一年里，我们已经有了两个新颖而相关的发现。Sgo1直接与称为ILF2-ILF3（白细胞介素增强子结合因子2和3）的rna结合蛋白复合物相互作用。Sgo1在体外与RNA结合。目的3和4计划进行实验，以确定这些发现的体内相关性。着丝粒姐妹染色单体内聚的过早丧失导致子细胞中染色体错误分离和染色体数量异常（非整倍性），这有助于癌症的形成和出生缺陷。这项研究将揭示染色体分离的机制，进而可能导致更好地理解和预防人类癌症和出生缺陷（如唐氏综合症）中的染色体不稳定性和非整倍体。

DESCRIPTION (provided by applicant): The genetic stability of an organism depends on the accurate partition of sister chromatids into two daughter cells during mitosis, which in turn requires the maintenance of the physical linkage (cohesion) between sister chromatids until their bipolar attachment to the mitotic spindle. Removal of cohesin (the protein complex that maintains sister chromatid cohesion) from chromosomes then leads to sister chromatid separation. In vertebrate cells, removal of cohesin occurs in two steps. At prophase, most of cohesin along the chromosome arms is removed through Polo-like kinase (Plk1)/Aurora B-dependent phosphorylation of cohesin. At metaphase, the residual centromeric pool of cohesin is cleaved by separase to allow sister chromatid separation. The long-term goal of my lab is to understand the molecular mechanism of chromosome segregation in mammalian cells. In this proposal, we will address an interesting puzzle in this area: how is the centromeric cohesin shielded from the actions of Plk1 /Aurora B in prophase? We have recently provided evidence to suggest that the spindle checkpoint kinase Bub1 targets and the Sgo1 centromeric protein and protein phosphatase 2A (PP2A) to centromeres where they counteract the phosphorylation of cohesin by Plk1 and other mitotic kinases. In Specific Aims 1 and 2, we will further delineate the mechanisms by which Bub1, Sgo1, and PP2A collaborate to protect centromeric cohesion. On the other hand, our results also point to a PP2A-independent role of Sgo1. Indeed, we have made two novel and related findings in the past year. Sgo1 directly interacts with an RNA-binding protein complex called ILF2-ILF3 (interleukin enhancer binding factors 2 and 3). Sgo1 itself binds to RNA in vitro. Experiments are planned in Aims 3 and 4 to establish the in vivo relevance of these findings. Premature loss of centromeric sister chromatid cohesion leads to chromosome missegregation and abnormal numbers of chromosomes in daughter cells (aneuploidy), which contributes to cancer formation and birth defects. The proposed research will shed light on the mechanism of chromosome segregation and may in turn lead to better understanding and prevention of chromosomal instability and aneuploidy in human cancers and birth defects, such as Down Syndrome.