Conserved regulation of proteostasis by post-translational protein AMPylation

摘要 本MIRA R35应用程序的主要目标是了解分子基础和机制， 蛋白质AMP化调节正常生理学和蛋白质聚集背景下的蛋白质稳态- 相关疾病。蛋白质AMP化由专用的AMP化酶赋予，其存在于单个 在大多数后生动物中复制。这些酶越来越多地被认为是控制蛋白质稳态， 当淀粉样蛋白--如β淀粉样蛋白（Aβ）、α-突触核蛋白（α-Syn）和多聚谷氨酰胺重复时， 蛋白质-聚集体。然而，我们对AMPase调节、靶点偏好及其在 在生理上以及疾病状态中仍然是难以捉摸的。缩小这些知识差距是获得 对翻译后蛋白AMP化如何控制蛋白质稳态具有根本性的重要见解，并将 潜在地使得能够开发利用该过程的治疗干预。这个的目标 建议1）剖析FIC-1/FICD介导的蛋白质的机制和生理影响 2）阐明了蛋白质AMP化在细胞质中的机制基础， 原子核我们将使用多方面的，翻译和多生物体的方法来解决遗传学和 生物化学与蛋白质聚集和蛋白质稳态调节中的这些中心问题相关。我们 研究计划利用四个互补的模型系统：线虫C。elegans，纯化蛋白质 用于体外生物化学、人类细胞系和体内小鼠模型，每种模型都具有独特的优势 在健康和疾病的背景下检查蛋白质AMP化。我们将利用我们独特的体内和体内 体外试验和工具，同时继续设计新的方法，以确定酶，目标，信号 参与AMPylation介导的蛋白质稳态调节的级联反应和生物学过程 续费在未来五年，我们的目标是充分发展现有项目，并设计新的工具， 技术，以不断产生具有转化潜力的遗传和机械数据。一种机械的 对这些过程的理解将促进我们对蛋白质折叠和蛋白质稳定途径的认识 这将代表着发现新的干预措施来预防或治疗蛋白质的关键步骤。 聚集相关疾病。

ABSTRACT The primary goal of this MIRA R35 application is to understand the molecular basis and mechanisms by which protein AMPylation regulates proteostasis in normal physiology and in the context of protein aggregation- associated diseases. Protein AMPylation is conferred by dedicated AMPylases, which are present in a single copy in most metazoans. These enzymes are increasingly recognized to control proteostasis, modulating how and when amyloidogenic proteins – such as amyloid-β (Aβ), α-synuclein (α-Syn) and poly-glutamine repeat proteins – aggregate. However, our knowledge of AMPylase regulation, target preference and their roles in physiological as well as in disease states remains elusive. Closing these gaps in knowledge is key for gaining fundamentally important insights into how post-translational protein AMPylation controls proteostasis, and will potentially enable the development of therapeutic interventions that capitalize on this process. The goals of this proposal are to 1) Dissect the mechanistic and physiological impact of FIC-1/FICD-mediated protein AMPylation on proteostasis, and 2) elucidate the mechanistic basis of protein AMPylation in the cytoplasm and nucleus. We will use a multi-faceted, translational and multi-organism approach to address the genetics and biochemistry associated with these central questions in protein aggregation and proteostasis regulation. Our research program is utilizing four complementary model systems: the nematode C. elegans, purified proteins for in vitro biochemistry, human cell lines and in vivo mouse models, each offering its unique set of advantages to examine protein AMPylation in the context of health and disease. We will leverage our unique in vivo and in vitro assays and tools, while continuing to design novel approaches, to define the enzymes, targets, signaling cascades and biological processes involved in AMPylation-mediated proteostasis regulation in unpreceded details. In the next five years, our goal is to fully develop current projects and to devise new tools and techniques to continuously produce genetic and mechanistic data with translational potential. A mechanistic understanding of these processes will advance our knowledge of protein folding and proteostasis pathways which will represent key steps towards the discovery of novel interventions to prevent or treat protein aggregation-associated disorders.