Molecular Basis of RING E3 Ligase Pirh2 and HECT E3 Ligase Itch Interaction.

蛋白质是人体中最重要的分子之一。调节蛋白质的稳定性和周转是细胞中的一项关键任务。大多数蛋白质通过泛素（Ub）-蛋白酶体途径（UPP）降解。UPP影响多种细胞过程和底物，并且UPP中的缺陷可导致人类疾病的发病机制。UPP在生物学中的核心作用得到了2004年诺贝尔化学奖的认可。泛素介导的蛋白质降解是一个三步过程，涉及三种酶：E1（Ub激活酶），E2（Ub缀合酶）和E3（Ub蛋白连接酶）。最近发现一类新的泛素化酶E4（一种Ub链组装因子）通过泛素融合降解途径降解某些蛋白质。这些酶一起催化一个或多个泛素部分与蛋白质赖氨酸残基的共价连接。E3酶在底物的识别和泛素化中起核心作用，并分为三个亚组：（i）HECT结构域连接酶，如E6-AP和Itch;（ii）RING结构域连接酶，包括Pirh 2和Mdm 2;和（iii）代表非典型RING结构域的U盒结构域连接酶，如UFD 2和CHIP（Hsp 70相互作用蛋白的羧基末端）。泛素化蛋白随后被26 S蛋白酶体靶向降解。 我们的实验室正在研究体内两种分子Pirh 2和Itch的调节和功能，这两种分子可能能够调节细胞生长。这两种分子属于E3连接酶家族，其促进蛋白质降解。我们以前证明Pirh 2可以负调节瘙痒的活动和功能。我们的研究团队将使用先进的实验室工具来探索人体控制瘙痒水平的系统，以及这种控制与细胞生长的关系。特别是，我们将努力了解分子Pirh 2和Itch是否是这些控制系统的关键部分，以及它们如何受到影响。我们的计划是研究人类细胞中的这两种分子，并了解它们彼此之间的关系以及它们与其他参与细胞生长和正常发育的分子之间的关系。因此，这一提议的结果对细胞生物学具有重要意义。我们希望我们的研究将有助于我们理解RING E3连接酶Pirh 2和HECT E3连接酶Itch之间相互作用的分子机制。

Proteins are one of the most important group of molecules in the human body. Regulation of protein stability and turnover is a key task in the cell. The majority of proteins are degraded by the ubiquitin (Ub)-proteasome pathway (UPP). The UPP affects a wide variety of cellular processes and substrates and defects in the UPP can result in the pathogenesis of human diseases. The central role of the UPP in biology has been recognized with the Nobel Prize for Chemistry in 2004. Ubiquitin-mediated protein degradation is a three-step process involving three enzymes: E1 (Ub-activating enzyme), E2 (Ub-conjugating enzyme), and E3 (Ub protein ligase). A new class of ubiquitination enzyme, E4 (a Ub chain assembly factor), was recently shown to be necessary for the degradation of some proteins via the ubiquitin fusion degradation pathway. Together, these enzymes catalyze the covalent attachment of one or more ubiquitin moieties to protein lysine residues. E3 enzymes play a central role in the recognition and ubiquitination of substrates and fall into three subgroups: (i) HECT domain ligases such as E6-APand Itch; (ii) RING domain ligases, including Pirh2 and Mdm2; and (iii) U-box domain ligases representing a non-canonical RING domain such as UFD2 and CHIP (carboxyl terminus of Hsp70-interacting protein). The ubiquitinated proteins are subsequently targeted for degradation by the 26S proteasome. Our laboratory is studying the regulation and function in the body of two molecules known as Pirh2 and Itch that may be able to regulate cell growth. The two molecules belong to the E3 ligase family, which promote protein degradation. We previously demonstrated that Pirh2 can negatively regulate Itch activities and functions. Our research team will use advanced laboratory tools to explore the body's systems for controlling levels of Itch and how this control relates to cell growth. In particular, we will work towards an understanding of whether the molecules Pirh2 and Itch are critical parts of these control systems and how they can be influenced. Our plan is to study the two molecules in human cells, and to learn their relationship to each other and to other molecules that are involved with cell growth and normal development. Thus, the outcome of this proposal has significant implications for cell biology. We are hopeful that our research will contribute to our understanding of the molecular mechanism of interaction between RING E3 ligase Pirh2 and HECT E3 ligase Itch.