Regulation of the cell cycle by SCF and APC/C ubiquitin ligases

描述（由申请人提供）：泛素-蛋白酶体系统对关键细胞周期调节蛋白的时间协调破坏代表了一种重要的调节机制，以确保特定蛋白质功能在正确的时间、在正确的隔室中以单向方式关闭。细胞周期机制的许多核心组分的蛋白水解由两大类泛素连接酶控制：SCF（Skp 1-Cul 1-F-box蛋白）复合物和后期促进复合物/环体（APC/C）。在人中有68种SCF连接酶，每种SCF连接酶的特征在于不同的F盒蛋白亚基，其通过将底物直接募集到连接酶的其余部分并最终募集到泛素缀合酶来提供特异性。尽管有大量的F-盒蛋白，但只有三种人SCF泛素连接酶（分别含有F-盒蛋白β Trcp、Fbw 7和Skp 2）具有良好建立的底物，其中许多参与细胞周期控制（例如，Cdc 25 A、细胞周期蛋白E、p21、p27、Wee 1）。我们提出了一个项目集中在一个新的层控制的细胞周期网络和它的整合与泛素系统。使用一种新的屏幕，我们已经确定了六个新的推定SCF基板，我们将表征其中之一，即E2 F3，密切参与细胞周期的控制（目的1）的蛋白质的降解的机制，调节和生物学功能。我们将进一步鉴定和表征那些被F-box蛋白Fbw 5靶向破坏以调节细胞周期进程的生物学重要底物（目的2）。最后，我们将研究betaTrcp在控制降解的一种新的基板确定使用生化屏幕：Claspin，一种蛋白质，是DMA复制监视机器的一部分（目的3）。鉴于细胞周期机制的重要功能，细胞周期调节蛋白的降解改变显然是癌细胞典型的无限制增殖的决定因素。随着我们继续解开如何通过泛素系统控制细胞增殖的调节蛋白的预定降解的机制，我们致力于将我们的基础研究结果与恶性转化的理解相结合。预计我们的研究结果将对基础科学和癌症生物学产生影响。

DESCRIPTION (provided by applicant): Temporally coordinated destruction of key cell cycle regulatory proteins by the ubiquitin-proteasome system represents an important regulatory mechanism to ensure that specific protein functions are turned off at the right time, in the right compartment and in a unidirectional fashion. Proteolysis of many core components of the cell cycle machinery is controlled by two major classes of ubiquitin ligases: the SCF (Skp1-Cul1-F-box protein) complexes and the Anaphase Promoting Complex/Cyclosome (APC/C). In humans there are sixty- eight SCF ligases, each characterized by a different F-box protein subunit that provides specificity by directly recruiting the substrate to the rest of the ligase and, ultimately, to the ubiquitin conjugating enzyme. Despite the large number of F-box proteins, only three human SCF ubiquitin ligases (containing the F-box proteins betaTrcp, Fbw7 and Skp2, respectively) have well-established substrates, many of which are involved in cell cycle control (e.g., Cdc25A, cyclin E, Emi1, p21, p27, Wee1). We propose a project focused on a new tier of control of the cell cycle networks and its integration with the ubiquitin system. Using a novel screen, we have identified six novel putative SCF substrates, and we will characterize the mechanism, regulation and biological function of the degradation of one of them, namely E2F3, a protein intimately involved in the control of the cell cycle (Aim 1). We will furthermore identify and characterize those biologically significant substrates that are targeted for destruction by the F-box protein Fbw5 to regulate cell cycle progression (Aim 2). Finally, we will study the role of betaTrcp in controlling the degradation of a novel substrate identified using a biochemical screen: Claspin, a protein that is part of the DMA replication surveillance machinery (Aim 3). Given the crucial function of the cell cycle machinery, altered degradation of cell cycle regulatory proteins is clearly a contributing determinant of the unrestrained proliferation typical of cancer cells. As we continue to unravel the mechanisms of how the scheduled degradation of regulatory proteins by the ubiquitin system controls cellular proliferation, we are committed to the integration of our basic research results with an understanding of malignant transformation. It is anticipated that the results of our studies will have an impact on both basic science and cancer biology.