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Role of lipid transfer proteins and membrane contact sites for intramitochondrial lipid trafficking

脂质转运蛋白和膜接触位点在线粒体内脂质运输中的作用

基本信息

批准号：
268448891
负责人：
Professor Dr. Thomas Langer
金额：
--
依托单位：
Max-Planck-Institut für Biologie des Alterns
依托单位国家：
德国
项目类别：
Research Grants
财政年份：
2015
资助国家：
德国
起止时间：
2014-12-31 至 2020-12-31
项目状态：
已结题

来源：
https://gepris.dfg.de/gepris/projekt/268448891?language=en
关键词：
Role lipid transfer proteins membrane

项目摘要

Lipid transport in cells occurs in form of vesicles or in a non-vesicular manner. Recent evidence suggests that lipid trafficking in mitochondria involves dedicated lipid transfer proteins (LTPs) and is dependent on a close apposition of cellular membranes. We have identified novel LTPs that mediate the transport of phosphatidic acid across the intermembrane space of yeast and human mitochondria to allow cardiolipin biosynthesis in the mitochondrial inner membrane. These LTPs comprise heterodimeric complexes, which are composed of Ups1 (or PRELID1 in human) and Mdm35 (or TRIAP1 in human). The LTPs comprise a highly conserved protein family with several homologues being present in all eukaryotic cells, but the function of other members of the protein family remains enigmatic. The maintenance of the lipid composition of mitochondrial membranes also depends on membrane contact sites between the endoplasmic reticulum (ER) and the mitochondrial outer membrane and between both mitochondrial membranes. However, how these membrane contacts contribute to the trafficking of various lipids and how they cooperate with LTPs is only poorly understood. In the present project, we will define the role of LTPs and membrane contact sites for lipid trafficking in mitochondria and examine the function of different members of the Ups/PRELI family of LTPs. A focus of research will be on the role of Ups2 for the biogenesis of the major membrane constituent phosphatidylethanolamine (PE). We will define the role of Ups2 in PE biogenesis using biochemical and cell biological approaches in vivo. Purified Ups2-Mdm35 complexes will be used to examine a putative function of Ups2 as LTP and, if applicable, determine the lipid specificity of the LTP. These experiments will be complemented by studies on membrane tethering complexes, in order to unravel a function of contacts between both mitochondrial membranes during PE biogenesis. Moreover, we will analyze how phosphatidylglycerol is transported from the ER to the mitochondrial inner membrane for cardiolipin synthesis. We will assess the role of membrane tethering complexes for PG transport and perform a genome-wide genetic screen in yeast to identify novel proteins involved in lipid transport from the ER to the inner membrane. Finally, we will employ photoactivatable and clickable phospholipid derivatives to monitor lipid import into mitochondria and to identify novel lipid-protein interactions that occur during lipid transfer and synthesis in mitochondria. These studies will provide new insights into the mechanisms of lipid trafficking in mitochondria and will address issues of general cell biological relevance related to the role of LTPs and membrane contact sites.

细胞中的脂质运输以囊泡的形式或以非囊泡的方式发生。最近的证据表明，线粒体中的脂质运输涉及专用的脂质转移蛋白（LTP），并且依赖于细胞膜的紧密并置。我们已经鉴定出新型 LTP，可介导磷脂酸穿过酵母和人类线粒体膜间隙的运输，从而允许线粒体内膜中心磷脂的生物合成。这些 LTP 包含异二聚体复合物，由 Ups1（或人中的 PRELID1）和 Mdm35（或人中的 TRIAP1）组成。 LTP 包含一个高度保守的蛋白质家族，所有真核细胞中都存在几种同源物，但该蛋白质家族其他成员的功能仍然是个谜。线粒体膜脂质组成的维持还取决于内质网（ER）和线粒体外膜之间以及线粒体膜之间的膜接触位点。然而，人们对这些膜接触如何促进各种脂质的运输以及它们如何与 LTP 合作知之甚少。在本项目中，我们将定义 LTP 和膜接触位点在线粒体脂质运输中的作用，并检查 LTP Ups/PRELI 家族不同成员的功能。研究的重点将是 Ups2 在主要膜成分磷脂酰乙醇胺 (PE) 生物合成中的作用。我们将使用体内生化和细胞生物学方法来定义 Ups2 在 PE 生物发生中的作用。纯化的 Ups2-Mdm35 复合物将用于检查 Ups2 作为 LTP 的假定功能，并在适用时确定 LTP 的脂质特异性。这些实验将通过对膜束缚复合物的研究来补充，以揭示 PE 生物发生过程中线粒体膜之间的接触功能。此外，我们将分析磷脂酰甘油如何从内质网转运至线粒体内膜以合成心磷脂。我们将评估膜束缚复合物在 PG 转运中的作用，并在酵母中进行全基因组遗传筛选，以鉴定参与从 ER 到内膜的脂质转运的新型蛋白质。最后，我们将采用可光激活和可点击的磷脂衍生物来监测线粒体中的脂质输入，并识别线粒体中脂质转移和合成过程中发生的新型脂质-蛋白质相互作用。这些研究将为线粒体脂质运输机制提供新的见解，并将解决与 LTP 和膜接触位点的作用相关的一般细胞生物学相关问题。