Protein sorting to peroxisomes via mitochondria: Characterization of the molecular mechanism and its impact on formation of organelle contact sites.

通过线粒体对过氧化物酶体进行蛋白质分选：分子机制的表征及其对细胞器接触位点形成的影响。

• 批准号: 455559921
• 负责人: Dr. Johannes Freitag
• 金额: --
• 依托单位: Arbeitsgruppe Genetik
• 依托单位国家: 德国
• 项目类别: Research Grants
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/455559921?language=en
• 关键词: Protein; sorting; peroxisomes; via; mitochondria

The protein phosphatase Ptc5 from Saccharomyces cerevisiae contains an N-terminal targeting signal for mitochondria and a C-terminal targeting signal for peroxisomes (PTS1). We recently reported that Ptc5 is sorted to peroxisomes via mitochondria by a novel targeting mechanism (Stehlik et al, 2020). We now will characterize the mechanism in greater detail(1.). In addition, we will focus on the following hypothesis: translocation of proteins into peroxisomes from mitochondria triggers the formation of organelle contact sites via molecular tug-of-war, which results from simultaneous interaction of the cargo protein with the import machineries of both compartments. Thus, protein translocation per se may organelles (2.).1. Elucidation of the molecular mechanism of protein sorting to peroxisomes via mitochondria1.1 Intra-molecular signals required for the novel translocation mechanism will be characterized.1.2 Genes identified in the high-content screen (e.g. ERMES components) will be further investigated to address the roles of their gene products for Ptc5 translocation. In addition, we will examine components of the mitochondrial import machinery, which are often encoded by essential genes.1.3 Finally, we will set up an in vitro reconstitution system that recapitulates our in vivo data.2. Functional consequences of indirect sorting to peroxisomesIn the second part of the project we will elucidate how sorting via competing targeting signals influences the physical interaction of mitochondria and peroxisomes. We have identified several S. cerevisiae proteins showing this domain structure (Stehlik et al, 2020).2.1 The effect of tug-of-war like sorting on the association of peroxisomes and mitochondria will be addressed by studying contact site formation upon depletion of components of the peroxisomal import machinery.2.2 We will analyze the biological function of proteins with competing targeting signals (Cat2, Tes1 and Pxp2) via mutants that are transported to peroxisomes directly without detour. Their effect on contact site formation and metabolite exchange between peroxisomes and mitochondria will be determined.2.3 We will create a switchable tether, which can be used to induce artificial contacts. Strains containing this tether will be analyzed for peroxisome function and protein translocation from mitochondria to peroxisomes.The planned project will help to understand how peroxisomal import via a mitochondrial detour contributes to the interplay of both organelles. We are convinced that the study is important as peroxisomes and mitochondria share many metabolic pathways, but how they physically interact is less understood. Since mammalian proteins are known that contain competing peroxisomal and mitochondrial targeting signals, our project may result in a better understanding of the role of peroxisome and mitochondria interaction in human physiology.

来自酿酒酵母（Saccharomyces cerevisiae）的蛋白磷酸酶Ptc5含有针对线粒体的N端靶向信号和针对过氧化物酶体（PTS1）的C端靶向信号。我们最近报道，Ptc5通过一种新的靶向机制通过线粒体被分选到过氧化物酶体中（Stehlik et al，2020）。我们现在将更详细地描述该机制（1.）。此外，我们将专注于以下假设：从线粒体到过氧化物酶体的蛋白质易位触发细胞器接触位点的形成，通过分子拔河，这是由于货物蛋白与两个隔间的进口机器同时相互作用的结果。因此，蛋白质易位本身可能是细胞器（2.）。1.阐明蛋白质分选过氧化物酶体的分子机制，通过蛋白质1.1所需的新的易位机制的分子内信号将进行表征。1.2在高含量筛选（例如ERMES组件）中识别的基因将进一步研究，以解决其基因产物的Ptc5易位的作用。此外，我们将研究线粒体输入机制的组成部分，这些组成部分通常由必需基因编码。1.3最后，我们将建立一个体外重建系统，以重现我们的体内数据。间接分选过氧化物酶体的功能后果在该项目的第二部分，我们将阐明如何通过竞争靶向信号分选影响线粒体和过氧化物酶体的物理相互作用。我们发现了几个S。显示这种结构域结构的酿酒蛋白（Stehlik等人，2020）。2.1通过研究过氧化物酶体输入机制的组分耗尽后接触位点的形成，将解决拔河式分选对过氧化物酶体和线粒体缔合的影响。2.2我们将分析具有竞争靶向信号的蛋白质的生物学功能（Cat2，Tes1和Pxp2）通过突变体直接运输到过氧化物酶体而无需绕道。将确定它们对接触位点形成和过氧化物酶体与线粒体之间代谢物交换的影响。2.3我们将创建可切换的系链，其可用于诱导人工接触。将分析含有该系链的菌株的过氧化物酶体功能和蛋白质从线粒体转运到过氧化物酶体。计划中的项目将有助于了解过氧化物酶体通过线粒体绕道进口如何有助于两种细胞器的相互作用。我们相信这项研究是重要的，因为过氧化物酶体和线粒体共享许多代谢途径，但它们如何物理相互作用还不太清楚。由于哺乳动物蛋白质是已知的，含有竞争过氧化物酶体和线粒体靶向信号，我们的项目可能会导致更好地了解过氧化物酶体和线粒体相互作用在人类生理学中的作用。