Regulation and function of the Clp 1p protein phosphatase

描述(申请人提供)：细胞周期蛋白依赖性激酶(CDK)的适时激活和失活调节大多数细胞周期转变。每个细胞周期步骤与其他步骤的精确协调对于细胞正确地将完整的基因组传递到每个子细胞是至关重要的。如果做不到这一点，可能会导致多倍体、基因组不稳定和癌症。逆转CDKs作用的磷酸酶的功能还不是很清楚。高度保守的CDC14家族磷酸酶起逆转CDK磷酸化的作用。我们对CDC14磷酸酶的大部分知识都来自酵母。关于这个磷酸酶家族的关键悬而未决的问题是它们是如何调节的，它们的靶点是什么，以及磷酸酶如何调节这些靶点以促进有丝分裂和胞质分裂的关键步骤？我们一直在研究裂解酵母CDC14同系物，也就是众所周知的CLP1。我们以前的工作表明，CLP1的活性与其他关键的细胞周期调节因子的活性密切相关。此外，我们还发现，在有丝分裂结束时，CLP1是促进适当的染色体分离、胞质分裂和CDK1失活所必需的。这些研究已经确定了参与这些重要步骤的CLP1的潜在靶点。在这项提案中，我们将利用这些结果来从分子角度定义CLP1是如何被调控的，以及它如何促进染色体分离、胞质分裂和CDK1失活。CLP1参与了细胞周期进程的这些不同步骤，使其成为控制关键细胞周期事件和促进基因组稳定性的中央调节因子。我们的具体目标是：1)检验该模型，即正确的染色体分离需要在Survivin和INCENP的CDK1磷酸化和CLP1去磷酸化之间保持仔细调控的平衡。2)检验Sid2激酶和14-3-3蛋白Rad24如何在细胞质分裂完成之前维持高活性的CLP1的假说。3)验证CLP1通过对CDC15的作用促进肌动球蛋白环稳定性的假说。4)确定CLP1和后期促进复合体在晚期有丝分裂中如何促进CDC25失活。

DESCRIPTION (provided by applicant): Timely activation and inactivation of Cyclin dependent kinases (CDKs) regulate most cell cycle transitions. Precise coordination of each cell cycle step with the others is essential for cells to correctly transmit an intact genome to each daughter cell. Failure to do so can lead to polyploidy, genomic instability and cancer. The function of phosphatases which reverse the action of CDKs is less well understood. The highly conserved Cdc14-family phosphatases act to reverse Cdk phosphorylation events. Most of our knowledge about Cdc14 phosphatases has come from yeast. Key unanswered questions about this family of phosphatases are how they are regulated, what are their targets, and how does the phosphatase regulate these targets to promote key steps of mitosis and cytokinesis? We have been studying the fission yeast Cdc14 homolog, known as Clp1. Our work previous work showed that Clp1 activity is closely integrated with the activities of other key cell cycle regulators. In addition, we found that Clp1 is required to promote proper chromosome segregation, cytokinesis, and inactivation of Cdk1 at the end of mitosis. These studies have identified potential targets for Clp1 involved in each of these important steps. In this proposal we will exploit these results to define in molecular terms how Clp1 is regulated, and how it acts to promote chromosome segregation, cytokinesis, and Cdk1 inactivation. The involvement of Clp1 in these various steps in cell cycle progression makes Clp1 a central regulator for controlling key cell cycle events and promoting genomic stability. Our specific aims are: 1) To test the model that proper chromosome segregation requires a carefully regulated equilibrium between Cdk1 phosphorylation and Clp1 dephosphorylation of Survivin and INCENP. 2) To test a hypothesis for how the Sid2 kinase and the 14-3-3 protein, Rad24, maintain elevated Clp1 activity until cytokinesis is complete. 3) To test a hypothesis that Clp1 promotes actomyosin ring stability through effects on Cdc15. 4) To determine how Clp1 and the anaphase promoting complex promote Cdc25 inactivation in late mitosis.