Biochemical characterization of cell cycle deubiquitinating enzymes

细胞增殖是由一个复杂的生化反应网络来调节的，该网络确保每个细胞分裂步骤都正确地按正确的顺序进行。在这些反应中，细胞调节蛋白被称为泛素-蛋白酶体系统（UPS）的酶系统定时降解。在这个系统中，名为E3泛素连接酶的酶复合物将多个泛素拷贝添加到底物蛋白，标记它以供蛋白酶体快速降解。另一个被鉴定为去遍在蛋白酶（DUB）的酶家族从底物蛋白中去除遍在蛋白，对抗E3连接酶的作用。E3连接酶和DUB活性的平衡决定了底物蛋白的稳定性，并在细胞分裂周期的调节中起着关键作用。这种平衡的失调导致不受控制的细胞增殖和遗传不稳定。 目前缺乏对E3连接酶和DUB网络的全面了解，这些酶参与细胞分裂的调控。使用大规模的功能基因组学方法，我们的实验室最近已经确定了新的DUBs控制的稳定性和表达水平的关键细胞周期调控蛋白。这项研究的目的是阐明这些酶的作用的分子机制，以确定其细胞靶点的库，并确定其对细胞增殖的影响。我们将使用互补的生物化学，遗传学，定量蛋白质组学和细胞生物学方法的组合来解决这些问题。总的来说，这些研究将有助于更好地了解蛋白质降解和UPS在控制细胞分裂及其在人类疾病中的功能障碍中的作用。

Cell proliferation is regulated by a complex network of biochemical reactions that ensure that each cell division step is performed correctly and in proper sequence. Among these reactions is the timed degradation of cellular regulatory proteins by an enzymatic system known as the ubiquitin-proteasome system (UPS). In this system, enzyme complexes named E3 ubiquitin ligases add multiple copies of ubiquitin to a substrate protein, tagging it for rapid degradation by the proteasome. Another family of enzymes identified as deubiquitinating enzymes (DUBs) remove ubiquitin from substrate proteins, opposing the action of E3 ligases. The balance in the activities of E3 ligases and DUBs determine the stability of substrate proteins and plays a key role in the regulation of the cell division cycle. Deregulation of this balance leads to uncontrolled cell proliferation and genetic instability. A comprehensive picture of the network of E3 ligases and DUBs involved in the regulation of cell division is currently lacking. Using large-scale functional genomic approaches, our laboratory has recently identified novel DUBs that control the stability and expression level of key cell cycle regulatory proteins. The objectives of this research proposal are to elucidate the molecular mechanisms underlying the effect of these enzymes, to identify the repertoire of their cellular targets, and to define their impact on cell proliferation. We will use a combination of complementary biochemistry, genetic, quantitative proteomics and cell biology approaches to address these questions. Collectively, these studies will contribute to a better understanding of the role of protein degradation and the UPS in the control of cell division and its malfunction in human diseases.