A proteomics-based approach for the analysis of sumoylation patterns in normal and apoptotic human cells

相扑是一种蛋白质修饰，其定义是蛋白质相扑与目标蛋白质的共价和可逆的结合。这条途径是哺乳动物细胞生存所必需的，这证明了它的关键作用。在自然条件下，只发现少量的总和蛋白被修饰，这使得识别它们变得困难。尽管如此，已经有数十种细胞和病毒蛋白被证明是被相扑修饰的，而且很可能还有更多的蛋白质还有待确认。虽然对单个蛋白质的苏莫化结果已经有了很好的描述，但对苏莫化(作为一种途径)在特定细胞功能中的重要性却知之甚少。这项拟议的研究计划将使用蛋白质组学工具来纯化特定途径中涉及的人类SUMoyl化蛋白质。相扑靶标将被确定，然后我们将使用遗传工具来研究相扑对这些单个蛋白质的影响。这种蛋白质组学/功能双重方法将首次应用于研究苏木糖化与细胞凋亡(细胞程序性死亡)以及细胞周期进程之间的关系。具体地说，我们将确定在不同的细胞周期阶段和在诱导细胞凋亡后被总和或去甲基化的蛋白质。然后我们将详细分析这些蛋白质的相扑作用。我们期望这个项目将发现新的相扑蛋白靶点，并展示这种修饰在正常细胞和凋亡细胞中的作用。这项工作还可能为旨在调节细胞周期的癌症治疗确定新的靶点。从长远来看，我们将应用这一方法来研究在其他情况下，如病毒感染的相扑作用的重要性。

Sumoylation, a protein modification, is defined by the covalent and reversible attachment of the protein SUMO to the target protein. This pathway is required for mammalian cell survival, arguing for its critical role.  Only small amounts of sumoylated proteins are found modified in natural conditions, making their identification difficult. Nonetheless, dozens of cellular and viral proteins haven been shown to be modified by SUMO, and it is likely that many more remain to be identified. Although the outcome of sumoylation has been well characterized for individual proteins, the importance of sumoylation (as a pathway) in specific cellular functions is poorly understood. The proposed research program will use proteomic tools for the purification of human sumoylated proteins involved in specific pathways. SUMO targets will be identified, then we will use genetic tools to investigate the impact of sumoylation for these individual proteins. This proteomics/functional dual approach will be first applied to investigate the relationships between sumoylation and apoptosis (programmed cell death) as well as cell cycle progression. Specifically, we will identify proteins that are either sumoylated or desumoylated at various cell cycle stages and following apoptosis induction. Then we will analyze in details the role of sumoylation for these proteins. We expect that this project will identify new protein targets of SUMO and show the role of this modification in the normal and apoptotic cell. This work might also identify new targets for cancer therapies aimed at regulating the cell cycle. On the long term, we will apply this approach to investigate the importance of sumoylation in other contexts such as viral infections.