Novel Substrates of SCF Ubiquitin Ligases in Cell Cycle Control and Cancer

描述(由申请人提供)：泛素系统是细胞过程的主要调节机制，其中速度、特异性和时机至关重要。泛素介导的关键底物的蛋白分解控制细胞周期进程、信号转导途径、分化、凋亡、DNA修复和免疫反应。这一过程是由一个多聚体机器介导的，它由一个调控泛素靶向组件和一个效应器蛋白质降解引擎组成。以泛素为靶标的调节成分本身由几个多聚体元件(例如，SCF泛素连接酶复合体)组成，这些元件贡献了这一过程中固有的许多特异性。在人类中，有68种SCF连接酶，每种连接酶都有不同的F-box蛋白亚基，通过直接将底物招募到连接酶的其余部分，并最终招募到泛素结合酶来提供特异性。值得注意的是，68个人SCF泛素连接酶中只有3个(分别包含F-box蛋白UTrcp、Fbw7和Skp2)具有成熟的底物，其中许多参与细胞周期控制。剩下的65个F-box蛋白被认为是“孤儿”，因为它们的底物仍有待发现。我们最近开发了一种新的免疫纯化策略，该策略可以丰富F-box蛋白的底物，然后进行质谱分析。我们将系统地鉴定具有生物学意义的人类孤儿F-box蛋白底物(特定目标1)。由于我们的研究兴趣，我们将特别关注那些孤立的F-box蛋白，我们的初步结果表明这些蛋白与细胞周期控制和癌症有关。在目标2下，我们将验证在目标1下确定的最具生物学意义的底物。由于SCF连接酶在调节细胞增殖方面发挥关键作用，因此通常是与癌症相关的放松调控的目标，并参与致癌转化。因此，从拟议的研究中获得的信息将与癌症生物学和其他增殖性疾病直接相关。

DESCRIPTION (provided by applicant): The ubiquitin system is a major regulatory mechanism of cellular processes in which speed, specificity, and timing are critical. Ubiquitin-mediated proteolysis of key substrates controls cell cycle progression, signal transduction pathways, differentiation, apoptosis, DNA repair and the immune response. This process is mediated by a multimeric machine, composed of a regulatory ubiquitin-targeting component and an effector protein degradation engine. The regulatory component, which targets ubiquitin to proteins destined for degradation, is itself composed of several multimeric elements (e.g., the SCF ubiquitin ligase complexes) that contribute much of the specificity inherent in the process. 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. Notably, only three out of 68 human SCF ubiquitin ligases (containing the F-box proteins UTrcp, Fbw7 and Skp2, respectively) have well-established substrates, many of which are involved in cell cycle control. The remaining 65 F-box proteins are considered as "orphan" since their substrates still await discovery. We have recently developed a novel immunopurification strategy that enriches for substrates of F-box proteins followed by mass spectrometry analysis. We will systematically identify biologically significant substrates of human orphan F-box proteins (Specific Aim 1). Because of our research interest, we will focus particularly on those orphan F-box proteins that our preliminary results suggest to be involved in cell cycle control and cancer. Under Aim 2, we will validate the biologically most significant substrates identified under Aim 1. Given their critical role in regulating cell proliferation, SCF ligases are often the target of cancer- related deregulation and involved in oncogenic transformation. Therefore, the information gained from the proposed studies will be of direct relevance to cancer biology and other proliferative diseases.