Cell Cycle control by cyclin-dependent kinases

超出空间 但前提是。 细胞周期蛋白依赖性蛋白激酶(CDK)参与真核细胞周期的调控。CDK被认为是通过 使细胞中的大量蛋白质底物磷酸化，但这些底物很少被 已确认身份。在拟议的研究中，将使用一种新的生化策略来执行大规模搜索 对于发芽酵母和脊椎动物中的CDK底物。 我们的初步研究已在萌芽酵母中鉴定出约200个CDKL底物，以及 在拟议工作的第一个目标中，将详细分析这些底物的一个子集。其他初步报告 研究表明，与之相关的细胞周期蛋白亚基的特性对 CDK1使许多酵母底物磷酸化，拟议研究的第二个目标将是探索 细胞周期蛋白特异性的分子基础及其在细胞中的生理功能 还将开发方法来系统地鉴定和表征大量的 脊椎动物细胞中的CDK底物。最后，将采用“化学遗传”的方法来构建小鼠。 和小鼠细胞系，其中的CDK1或CDK2可以被高度特异性地抑制，从而允许进行严格的分析 它们在正常细胞生物学和肿瘤中的功能。 这些研究有望为CDK的基本机制提供广泛的新见解 驱动细胞周期事件。这些见解是朝着更好地理解 染色体分离缺陷和其他细胞周期缺陷是癌细胞的特征。

EXCEED THE SPACE PROVIDED. Cyclin-dependent kinases (Cdks) drive the events of the eukaryotic cell cycle. Cdks are thought to act by phosphorylating a large number of protein substrates in the cell, but few of these substrates have been identified. In the proposed studies, a novel biochemical strategy will be used to perform large scale searches for Cdk substrates in budding yeast and vertebrates. Our preliminary studies have led to the identification of about 200 Cdkl substrates in budding yeast, and in the first aim of the proposed work a subset of these substrates will be analyzed in detail. Other preliminary work has revealed that the identity of the associated cyclin subunit has a profound effect on the rate at which Cdkl phosphorylates many yeast substrates, and the second aim of the proposed studies will be to explore the molecular basis of this cyclin specificity and its physiological function in the cell. Methods will also be developed for the systematic identification and characterization of large numbers of Cdk substrates in vertebrate cells. Finally, a 'chemical genetic' approach will be employed to construct mice and mouse cell lines in which Cdkl or Cdk2 can be inhibited with high specificity, allowing rigorous analysis of their function in normal cell biology and in tumors. These studies promise to provide a wide range of new insights into the basic mechanisms by which Cdks drive cell-cycle events. Such insights are an important step toward a better understanding of the chromosome segregation defects and other cell-cycle defects that characterize the cancer cell.