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水解控制细胞周期进程、信号转导途径、分化、凋亡、DNA修复和免疫应答。这一过程是由一个多聚体机器介导的，该机器由一个调节性泛素靶向组分和一个效应蛋白降解引擎组成。将遍在蛋白靶向于注定要降解的蛋白质的调节组分本身由几个多聚体元件组成（例如，SCF遍在蛋白连接酶复合物），其在该过程中贡献了许多固有的特异性。在人类中，存在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。值得注意的是，68种人SCF泛素连接酶中只有3种（分别含有F-box蛋白UTrcp、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。剩余的65种F-box蛋白被认为是“孤儿”，因为它们的底物仍在等待发现。我们最近开发了一种新的免疫纯化策略，富集F-box蛋白的底物，然后进行质谱分析。我们将系统地鉴定人类孤儿F-box蛋白的生物学重要底物（特定目的1）。由于我们的研究兴趣，我们将特别关注那些孤儿F-box蛋白，我们的初步结果表明，这些蛋白参与细胞周期控制和癌症。在目标2下，我们将验证目标1下确定的最具生物学意义的底物。鉴于它们在调节细胞增殖中的关键作用，SCF连接酶通常是癌症相关的失调的靶标并且参与致癌转化。因此，从拟议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.