Anaphase Promoting Complex-mediated Proteolysis

有丝分裂过程中的一个关键事件是泛素依赖性的降解， 细胞周期蛋白和其他关键蛋白的后期促进复合物（ARC;也称为细胞周期小体）。 基质含有降解基序，如破坏盒和KEN盒， 它们的泛素化以及它们与ARC激活剂Cdc 20和Cdh 1的结合。两个主题都是必需的 用于有效降解，但任一单独足以结合Cdhlp。我们提出了一个途径 用于APC-Cdhip-底物复合物的组装，其中无APC的Cdhlp首先结合底物。 Cdh 1 p与底物破坏盒的结合刺激Cdh 1 p与ARC的结合， 推测是通过Cdhlp的构象变化。 目前的研究旨在加深我们对APC介导的蛋白水解在出芽过程中的理解 酵母特别地，我们希望识别新颖的ARC基板，以了解如何 APC-Cdh 1 p/Cdc 20 p-底物复合物的组装和拆卸，并了解 纺锤体组装检查点利用KEN盒关闭APC介导的蛋白水解， 染色体不能正确地附着在有丝分裂的纺锤体上。 为了实现这些目标，我提出以下具体目标： 1)识别新型ARC基板。 2)评估APC-Cdhip-底物组装途径的通用性。 3)为了确定D-Box接合如何刺激Cdh 1 p与ARC结合： 4)确定保守Cdhlp基序和磷酸化在APC-Cdh 1 p-底物中的作用 组装件. 5)确定APC-Cdh 1 p-底物复合物的降解是否为活性过程。 6)确定主轴组件检查点中MadSp KEN盒的功能。 关键蛋白质的降解对细胞分裂至关重要。因此，理解这种退化是 对于理解正常细胞增殖，以及这个过程在疾病状态下是如何出错的， 尤其是癌症。

A key event in the progression through and exit from mitosis is the ubiquitin-dependent degradation of cyclins and other key proteins by the Anaphase Promoting Complex (ARC; also called the cyclosome).ARC substrates contain degradation motifs such as the Destruction Box and the KEN Box that are required for their ubiquitination and for their binding to the ARC activators, Cdc20 and Cdh1. Both motifs are required for efficient degradation, but either one alone suffices for binding to Cdhlp. We have proposed a pathway for the assembly of the APC-Cdhip-substrate complex in which APC-free Cdhlp first binds a substrate. Engagement of the substrate Destruction Box by Cdh1p stimulates the binding of Cdh1p to the ARC, presumably via a conformation change in Cdhlp. The current studies are aimed at furthering our understanding of APC-mediated protedlysis in budding yeast. In particular, we wish to identify novel ARC substrates, to understand how APC-Cdh1p/Cdc20p-substrate complexes are assembled and disassembled, and to understand how the spindle assembly checkpoint makes use of a KEN box to turn off APC-mediated proteolysis when chromosomes are not properly attached to the mitotic spindle. To achieve these goals, I propose the following Specific Aims: 1) To Identify Novel ARC Substrates. 2) To Assess the Generality of the APC-Cdhip-Substrate Assembly Pathway. 3) To Determine How D-Box Engagement Stimulates Cdh1p Binding to the ARC: 4) To Determine the Roles of Conserved Cdhlp Motifs and of Phosphcrylation in APC-Cdh1p-Substrate Assembly. 5) To Determine Whether Disassembly of the APC-Cdh1p-Substrate Complex is an Active Process. 6) To Determine the Function of the MadSp KEN Box in the Spindle Assembly Checkpoint. Degradation of key proteins is essential for cell division. Understanding this degradation is thus important for understanding normal cell proliferation, and how this process goes awry in disease states, particularly cancers.