Systems-to-structure approaches for defining mitochondrial protein function

项目摘要 线粒体是代谢和信号传导的中心，其功能对除少数真核细胞外的所有细胞都是必需的 类型尽管线粒体是细胞生物学的标志性发电站，但它们的许多方面仍然存在 这一事实导致我们几乎完全无法解决线粒体功能障碍 治疗上这种功能障碍与一系列罕见的先天性代谢缺陷有关， 越来越多的常见疾病-包括帕金森氏症，阿尔茨海默氏症，各种癌症，以及2型糖尿病 糖尿病-通常通过不同的方式。例如，异常的线粒体生物发生可能无法正确地设置 细胞线粒体含量;失调的信号传导过程可能无法校准线粒体活性， 改变细胞的需要;和蛋白质故障可以使核心生物能量过程无效。一个主要 理解并最终解决这些过程的瓶颈是，驱动它们的蛋白质具有 往往无法识别。与此同时，数百种已知的线粒体蛋白质的功能， 这些作用没有得到界定，或者充其量也只是知之甚少。2008年，我领导了一项综合努力， 哺乳动物线粒体蛋白质组的综合纲要-称为MitoCarta-， 许多已知的哺乳动物线粒体蛋白，并暴露了这一知识的主要差距：惊人的~300 约1100种蛋白质中没有注释功能，包括约50种现在与人类直接相关的蛋白质。 疾病因此，我的研究计划的高层次目标是实现一个更全面的 通过系统地建立孤儿的功能来理解线粒体生物学 线粒体蛋白及其在疾病相关过程中的作用。我们首先设计小说， 多维分析旨在建立这些蛋白质之间的新联系， 路径和过程。这些包括定制的、高通量的蛋白质-蛋白质相互作用筛选，大规模的 酵母和人类细胞基因敲除的基于规模质谱的分析，以及计算 接近。然后，我们采用机械和结构的方法来定义选择蛋白质的功能， 生化深度，包括古老的和非典型的激酶和脂质结合蛋白，使线粒体 辅酶Q生物合成途径和其他必需的代谢过程。最后，我们将探讨如何后- 转录和翻译后调节因子的作用是建立一个定制的线粒体基础结构 能够满足不断变化的细胞需求。总的来说，通过有目的地阐明未探索的领域， 线粒体生物学，我们正在迅速达到一个更完整的了解这些细胞器做什么， 它们的蛋白质成分是如何实现其无数功能的。这些努力有望帮助建立一个深刻的， 对线粒体生物化学的机械理解，这将激发新的治疗策略， 大量的人类疾病都源于线粒体功能障碍。 !

PROJECT SUMMARY Mitochondria are centers of metabolism and signaling whose function is essential to all but a few eukaryotic cell types. Despite their position as the iconic powerhouses of cellular biology, many aspects of mitochondria remain remarkably obscure—a fact that contributes to our near complete inability to address mitochondrial dysfunction therapeutically. Such dysfunction is associated with a spectrum of rare inborn errors of metabolism and an increasing number of common diseases—including Parkinson’s, Alzheimer’s, various cancers, and type 2 diabetes—often through distinct means. For instance, aberrant mitochondrial biogenesis can fail to properly set cellular mitochondrial content; dysregulated signaling processes can fail to calibrate mitochondrial activity to changing cellular needs; and malfunctioning proteins can render core bioenergetic processes ineffectual. A major bottleneck to understanding—and ultimately addressing—these processes is that the proteins driving them have often not been identified. Concurrently, the functions of hundreds of known mitochondrial proteins that may fulfill these roles are undefined, or at best are poorly understood. In 2008, I led an integrative effort to generate a comprehensive compendium of the mammalian mitochondrial proteome—termed MitoCarta—that doubled the number of known mammalian mitochondrial proteins and exposed this major gap in knowledge: A striking ~300 of the ~1100 proteins had no annotated function, including ~50 that are now directly associated with human disease. Thus, the high-level goal of my research program is to achieve a more comprehensive understanding of mitochondrial biology by systematically establishing the functions of orphan mitochondrial proteins and their roles within disease-related processes. We do so by first devising novel, multi-dimensional analyses designed to make new connections between these proteins and established pathways and processes. These include customized, high-throughput protein-protein interaction screens, large- scale mass spectrometry-based profiling of yeast and human cell gene knockouts, and computational approaches. We then employ mechanistic and structural approaches to define the functions of select proteins at biochemical depth, including ancient and atypical kinases and lipid binding proteins that enable the mitochondrial coenzyme Q biosynthesis pathway and other essential metabolic processes. Finally, we investigate how post- transcriptional and post-translational regulators operate to establish a customized mitochondrial infrastructure capable of meeting changing cellular needs. Overall, by purposefully elucidating the unexplored areas of mitochondrial biology, we are rapidly arriving at a more complete understanding of what these organelles do and how their protein componentry enables their myriad functions. These efforts promise to help establish a deep, mechanistic understanding of mitochondrial biochemistry that will motivate novel therapeutic strategies for the vast array of human disorders rooted in mitochondrial dysfunction. !