Molecular basis for the recognition of N- and C-degrons

我们体内的每个细胞都含有数以千计的蛋白质，需要精确地控制蛋白质的动态平衡，才能维持平衡和功能的蛋白质组。细胞中蛋白质的丰度取决于蛋白质合成和降解的速度。任何不需要的蛋白质都需要被迅速降解。此外，错误折叠、聚集和异常的蛋白质也需要及时降解。泛素-蛋白酶体系统(UPS)是维持蛋白质动态平衡的关键蛋白质降解途径，UPS相关蛋白的异常与多种人类疾病有关，包括神经退行性变、异常衰老和癌症。在泛素-蛋白酶体系统中，泛素激活(E1)、泛素结合(E2)和泛素连接酶(E3)等一系列酶使目标蛋白被泛素化，泛素化的底物随后被26S蛋白酶体降解成短肽。E3泛素连接酶通过识别底物中存在的降解信号来特异性地靶向底物，例如位于靶蛋白N-末端或C-末端的信号基序。几乎所有的蛋白质都有其独特的N-端或C-端序列背景，它们可以成为降解信号，并被相应的E3连接酶识别，通过不同的N-或C-降解子途径进行降解。如果我们要更深入地了解泛素-蛋白酶体系统，全面了解E3泛素连接酶对特定退化的识别如何驱动选择性蛋白质降解是至关重要的。在这一发现研究计划中，我们的目标是通过研究这些降解途径的底物识别机制，识别每条途径的生理底物，并为这些系统开发特定的小分子抑制剂或化学探针，来系统地表征这些降解途径。具体地说，我们将着重于以下三个目标：1)了解底物序列选择性和ATE1在Arg/N-degron途径中催化蛋白质精氨酸化的机理；2)揭示FEM1C在Arg/C-degron途径中对Arg/C-degron序列的选择性和识别机制；以及3)确定FEM1B具有不同于FEM1C的C末端序列选择性的原因，这将有助于揭示Arg/C-degron途径中不同FEM1成员之间的分工。这一研究计划不仅将有助于更好地阐明退化途径在基本生物过程中的作用，而且还将有助于出现治疗与这些退化途径相关的人类疾病的药物，最终改善加拿大人和世界各地人民的健康。最后但并非最不重要的一点是，这项研究计划将为培养未来蛋白质科学和泛素生物学领域的基础和应用科学家提供坚实的技术平台。

Each cell in our body contains thousands of proteins, and the protein homeostasis need to be precisely controlled in order to maintain a balanced and functional proteome. The abundance of proteins in cells is determined by the rates of protein synthesis and degradation. Any unwanted proteins need to be degraded rapidly. In addition, misfolded, aggregated and abnormal proteins need to be degraded in a timely fashion as well. The ubiquitin-proteasome system (UPS) is a critical protein degradation pathway that maintains protein homeostasis, and aberrations of the UPS associated proteins contribute to various human diseases, including neurodegeneration, abnormal aging and cancers. In the ubiquitin-proteasome system, proteins targeted for degradation are ubiquitinated by a cascade of enzymes: ubiquitin-activating (E1), ubiquitin-conjugating (E2) and ubiquitin-ligase (E3) enzymes, and the ubiquitinated substrates are subsequently degraded into short peptides by the 26S proteasome. An E3 ubiquitin ligase specifically targets its substrates by recognition of degradation signals present in substrates, such as signal motifs located  at the N-termini or C-termini of the target proteins. Virtually all proteins have their unique N- or C-terminal sequence contexts, which can become degradation signals and be recognized by respective E3 ligases for degradation through distinct N- or C-degron pathways. A comprehensive understanding of how the recognition of specific degrons by E3 ubiquitin ligases drives selective protein degradation is critical if we are to attain a deeper understanding of the ubiquitin-proteasome system. In this discovery research program, we aim to characterize these degradation pathways systematically through investigating their substrate recognition mechanisms, identification of physiological substrates for each pathway and develop specific small-molecule inhibitors or chemical probes for these systems. Specifically, we would like to focus on the following three aims: 1) Understand substrate sequence selectivity and catalytic mechanism of protein arginylation by ATE1 in the Arg/N-degron pathway; 2) Delineate the Arg/C-degron sequence selectivity and recognition mechanism by FEM1C in the Arg/C-degron pathway; and 3) Determine why FEM1B has different C-terminal sequence selectivity than FEM1C, which will shed lights on the labor of division among the different FEM1 members in the Arg/C-degron pathway. This research program will not only be valuable to better elucidate the roles of degron pathways in fundamental biological processes, but also contribute to the advent of therapeutic agents against human diseases associated with these degron pathways, ultimately improving the health of Canadians and people around the world. Last but not the least, this research program will provide a solid  platform to train future basic and application scientists in the fields of protein science and ubiquitin biology.