Elucidating the mechanism of action of novel ClpP activators in activation of the mitochondrial unfolded protein response.

线粒体未折叠蛋白反应(UPRMT)是一种高度保守的应激途径 当失调与一系列退行性疾病有因果关系时 帕金森症、老年痴呆症、共济失调和衰老。成分鉴定的最新进展 UPRMT的研究激发了人们对这一“可用药”途径的兴趣。UPRMT的核心是 线粒体蛋白水解酶ClpP，是损伤和错误折叠的蛋白质周转所必需的 对细胞压力的反应。我们和其他人最近发现ClpP是一个出人意料的目标 一类新的抗癌化合物--咪喹酮(ONC201)及其类似物 (https://www.the-scientist.com/news-opinion/found--a-cancer-drugs-mechanism-of-action- 65918)。我们发现这些化合物激活了ClpP和UPRMT，这是由 线粒体蛋白质降解，线粒体呼吸链活性受损，以及 增加综合应激反应(ISR)蛋白(CHOP/ATF4)。因此，其主要目标是 本研究旨在探讨UPRMT的基本机制，阐明药物是如何在体内发挥作用的。 ClpP的诱导激活启动了重要的应激信号，调节细胞生长和 新陈代谢。我们提出了三个目标来实现这一点：在目标1中，我们将使用 蛋白质组学方法鉴定线粒体释放的ClpP底物和多肽。 在目标2中，我们将确定ClpP激活如何失调线粒体代谢，以及 通过糖酵解和TCA循环影响葡萄糖和谷氨酰胺的消耗。在《目标3》中我们 将决定ClpP激活如何改变胞质信号事件，即激活 ISR和由此导致的蛋白质合成减少。如果成功，我们的研究将提供 对UPRMT的生物学功能以及ClpP如何调节这一功能的重大新见解 在正常和疾病状态下，通路起着调节细胞应激的作用。

The mitochondrial Unfolded Protein Response (UPRMT) is a highly conserved stress pathway that when dysregulated is causally connected to a host of degenerative diseases including Parkinsons, Alzheimers, Freiderichs Ataxia and aging. The recent identification of components in the UPRMT has stimulated interest in this as a “druggable” pathway. Central to the UPRMT is the mitochondrial protease ClpP, required for the turnover of damaged and misfolded proteins in response to cellular stresses. We and others recently identified ClpP as an unexpected target for a novel class of anti-cancer compounds known as imipridones (ONC201) and related analogs (https://www.the-scientist.com/news-opinion/found--a-cancer-drugs-mechanism-of-action- 65918). We showed that these compounds activated ClpP and the UPRMT as determined by the degradation of mitochondrial proteins, impaired mitochondrial respiratory chain activity, and increased integrated stress response (ISR) proteins (CHOP/ATF4). Thus, the main objective of this research is to investigate the basic mechanisms of the UPRMT and elucidate how drug- induced activation of ClpP initiates important stress signals that regulate cell growth and metabolism. We propose three aims to accomplish this: in Aim 1 we will use comprehensive proteomics approaches to identify ClpP substrates and peptides released from the mitochondria. In Aim 2 we will determine how ClpP activation dysregulates mitochondrial metabolism, and affects the consumption of glucose and glutamine by glycolysis and the TCA cycle. In Aim 3 we will determine how ClpP activation alters cytosolic signaling events, namely the activation of the ISR and the resulting reduction in protein synthesis. If successful, our studies will provide significant new insight into the biological functions of the UPRMT and how ClpP regulates this pathway functions to modulate cell stress in normal and disease states.