Conserved regulation of proteostasis by post-translational protein AMPylation

摘要 此Mira R35应用程序的主要目标是了解其分子基础和机制 在正常生理和蛋白质聚集的背景下，蛋白质AMP化调节蛋白平衡- 相关疾病。蛋白质的AMPyl化是由专用的AMPylase授予的，这些酶存在于单个 在大多数后生动物中复制。这些酶被越来越多地认识到控制蛋白平衡，调节如何 而当淀粉样蛋白-β(Aβ)，α-突触核蛋白(α-Syn)和聚谷氨酰胺重复)等淀粉样蛋白 蛋白质--聚集体。然而，我们对AMPylase调控、靶向偏好及其在 生理状态和疾病状态仍然难以捉摸。弥合这些知识差距是获得 对翻译后蛋白质AMP化如何控制蛋白平衡以及将 有可能使利用这一进程的治疗干预措施的发展成为可能。这样做的目的是 建议1)剖析FIC-1/FICD介导的蛋白质的机制和生理影响 2)阐明细胞质和细胞内蛋白质AMP化的机制。 原子核。我们将使用多方面的、翻译的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平衡调控中的这些中心问题有关。我们的 研究计划利用四个互补的模型系统：线虫、线虫、纯化蛋白 对于体外生物化学、人类细胞系和体内小鼠模型，每一种都有其独特的一组优势 在健康和疾病的背景下检查蛋白质的AMP化。我们将利用我们独特的体内和体内 体外分析和工具，同时继续设计新的方法，以定义酶，靶标，信号 参与AMP介导的蛋白平衡调节的级联反应和生物过程 细节。在未来五年，我们的目标是充分发展现有的项目，并设计出新的工具和 不断产生具有翻译潜力的遗传和机械数据的技术。机械师 对这些过程的理解将促进我们对蛋白质折叠和蛋白稳态途径的了解 这将是发现预防或治疗蛋白质的新干预措施的关键步骤 聚集性疾病。

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.