Biogenesis of alpha-helical mitochondrial outer membrane proteins in higher eukaryotes

项目总结/摘要 线粒体是内共生起源的重要细胞器，其已经进化到在生物体内发挥关键作用。 真核生物生理学线粒体活性依赖于嵌入线粒体外层的蛋白质 细胞膜（OMM），介导细胞质-细胞质通讯，以及细胞的关键方面， 细胞凋亡和先天免疫反应等功能。α-螺旋蛋白是一个重要的亚类， OMM蛋白。然而，它们如何插入哺乳动物细胞中OMM的脂质双层， 最近不清楚。 我与丽贝卡·沃希斯和乔纳森·韦斯曼的实验室合作， OMM驻留蛋白MTCH 2是一个新的插入酶家族的定义成员，它既是必需的， 足以将α-螺旋蛋白插入OMM中。MTCH 2是溶质的发散成员 载体家族25（SLC 25），并且已经进化为利用用于插入的典型转运蛋白折叠。生物信息 分析显示，MTCH 2在过氧化物酶体中具有同源物，并且在整个动物中具有直向同源物，这表明MTCH 2是一种新的基因。 α-螺旋蛋白插入跨膜和真核生物的共同机制。 基于这一发现，该提案旨在全面了解α-螺旋 蛋白质被正确地靶向OMM并插入到真核生物的脂质双层中。这项工作将 解决了细胞生物学中的一个基本问题，因为OMM蛋白质组已经发展到支持越来越多的 更复杂的功能。α-螺旋蛋白由一个或多个 跨膜结构域（TA），尽管它们的TM在数量和生物物理范围上可以显著变化， 例如疏水性的特性。本提案旨在首先加深对MTCH 2的理解 功能首先，将结合体内和生物化学技术来绘制底物TM的路线 通过MTCH 2进入脂质双分子层，直接测试MTCH 2保守的亲水性沟是否具有 在这个过程中的直接作用。其次，这项工作将确定MTCH 2的结构同源物是否具有 保留了它们在真核生物中的插入酶能力。 通过定义MTCH 2的分子基础获得的生物化学技能和知识将是 结合系统功能基因组学来确定α-螺旋蛋白如何靶向OMM， 以及是否需要除了MTCH 2以外的其他因素来支持这种多样性类别的生物发生。 总的来说，这一建议将为逐步发展的支持机制提供重要的见解。 真核生物此外，对控制α-螺旋OMM蛋白插入的机制的深入了解将有助于 对于开发与线粒体外膜蛋白失调相关的新疗法至关重要， 包括神经退行性疾病，如帕金森氏症和阿尔茨海默氏症。

Project Summary/Abstract Mitochondria are essential organelles of endosymbiotic origin which have evolved to play critical roles in eukaryotic physiology. Mitochondrial activity is dependent on proteins embedded in the outer mitochondrial membrane (OMM), which mediate mitochondrial-cytoplasmic communication, and critical aspects of cellular function such as apoptosis and the innate immune response. a-helical proteins are an important subset of OMM proteins. However, how they get inserted into the lipid bilayer of the OMM in mammalian cells has until recently been unclear. Work by myself, in collaboration with Rebecca Voorhees and Jonathan Weissman’s labs, has identified the OMM resident protein MTCH2, a defining member of a novel family of insertases, as both necessary and sufficient for the insertion of a-helical proteins into the OMM. MTCH2 is a diverged member of the solute carrier family 25 (SLC25), and has evolved to exploit the canonical transporter fold for insertion. Bioinformatic analysis reveals that MTCH2 has a homolog in peroxisomes, and orthologs across holozoa, suggesting a common mechanism for a-helical protein insertion across membranes and eukaryotes. Building on this finding, this proposal aims to develop a comprehensive understanding of how a-helical proteins are correctly targeted to the OMM and inserted into the lipid bilayer across eukaryotes. This work will address a fundamental question in cell biology, as the OMM proteome has evolved to support increasingly more complex functions in higher eukaryotes. a-helical proteins are defined by the presence of one or more transmembrane domains (TAs), though their TMs can vary significantly in number and a range of biophysical characteristics such as hydrophobicity. This proposal aims to first develop a deeper understanding of MTCH2 function. First, in vivo and biochemical techniques will be combined to map the route a substrate TM takes through MTCH2 into the lipid bilayer, directly testing whether MTCH2’s conserved hydrophilic groove has a direct role in this process. Second, this work will establish whether structural homologs of MTCH2 have retained their insertase ability across eukaryotes. The biochemical skills and knowledge acquired by defining the molecular basis for MTCH2 will be combined with systematic functional genomics to establish how a-helical proteins get targeted to the OMM, and whether other factors besides MTCH2 are required to support the biogenesis of this diverse class. Cumulatively, this proposal will provide important insights into the mechanisms that have evolved to support eukaryotic life. Further, intimate knowledge of the machinery that governs a-helical OMM protein insertion will be critical for developing new treatments associated with outer mitochondrial membrane protein dysregulation, including neurodegenerative diseases such as Parkinson’s and Alzheimer’s.