Regulation of WASH-mediated Endosomal Protein Recycling

 描述(由申请人提供)：蛋白质在细胞内的运输是一个高度调控的过程，是细胞生物学的一个基本方面。内体蛋白循环途径是一种重要的细胞运输过程，它通过将溶酶体中的蛋白质循环回高尔基体或质膜，促进溶酶体中降解蛋白的解救。这一基本的细胞过程的主要贡献者是WASH蛋白。WASH是肌动蛋白成核促进因子WASP家族的一部分，它专门促进肌动蛋白在内体上的组装，以促进蛋白质循环。重要的是，洗涤和内体蛋白循环与许多人类疾病有关，包括遗传性散发性截瘫、阿尔茨海默氏症、自闭症和癌症。此外，这一细胞过程是运输致命性毒素所必需的，如霍乱、蓖麻毒素、 并将炭疽病带到他们的行动地点。因此，了解WASH的功能和调控对人类健康具有重要意义。WASH被保持在自动抑制状态，因为它被并入多亚基复合体，称为WASH调节复合体或SHRC。最近，我们描述了泛素化是如何激活WASH的。MAGE-L2-TRIM27 E3泛素连接酶非典型的K63连接泛素化WASH是WASH激活、产生内体F-肌动蛋白和内体蛋白循环的重要事件。利用一些体外生化和细胞方法，该项目旨在通过翻译后修饰来确定调节WASH活性的机制和信号通路。这些研究对于我们理解WASH介导的内体蛋白循环，打破与操纵这一细胞过程相关的人类疾病的关键障碍，以及增进我们对这一基本细胞过程的科学知识具有重要意义。

 DESCRIPTION (provided by applicant): Transport of proteins throughout the cell is a highly regulated process that is a fundamental aspect of cell biology. The endosomal protein recycling pathway is one important cellular transport process which promotes the rescue of proteins destined for degradation in the lysosome by recycling them back to the Golgi or plasma membrane from endosomes. A major contributor to this fundamental cellular process is the WASH protein. WASH is part of the WASP family of actin nucleation promoting factors and specifically promotes actin assembly on endosomes to facilitate protein recycling. Importantly, WASH and endosomal protein recycling have been linked to a number of human diseases, including hereditary sporadic paraplegia, Alzheimer's, autism, and cancer. Furthermore, this cellular process is required for the transport of lethal pathogenic toxins, such as cholera, ricin, and anthrax to their site of action. Therefore, understanding the function and regulation of WASH has important implication on human health. WASH is held in an auto-inhibited state due to its incorporation into a multi-subunit complex, referred to as the WASH regulatory complex or SHRC. Recently, we described how WASH is activated by ubiquitination. Non- canonical K63-linked ubiquitination of WASH by the MAGE-L2-TRIM27 E3 ubiquitin ligase is an essential event in WASH activation, generation of endosomal F-actin, and endosomal protein recycling. Utilizing a number of in vitro biochemical and cellular approaches, this project aims to determine the mechanisms and signaling pathways regulating WASH activity through post-translational modifications. These studies are important for our understanding of WASH-mediated endosomal protein recycling, breaking critical barriers to treating human diseases associated with manipulation of this cellular process, and advancing our scientific knowledge of this fundamental cellular process.