Phospholipid Dynamics in Membrane Assembly

细胞生物学和生物化学中一个尚未解决的基本问题是细胞的分子定义 膜生物发生的磷脂转运机制。这一过程对所有细胞来说都是必不可少的 生长、复制、分化和动态平衡。这项提议的重点是利用酵母的力量 分子遗传学，剖析这一过程，并理解它如何贯穿始终的一般原理 真核生物。我们已经在定义几个基因和基因产物方面取得了进展 参与了运输反应，但我们的知识仍然存在很大差距。这项工作将继续下去 集中在涉及氨基甘油磷脂(磷脂酰丝氨酸， 磷脂酰乙醇胺和磷脂酰胆碱)，因为我们以前开发的工具很大 便于测量膜间转移事件，还提供了遗传筛查和 对在这些运输过程中有缺陷的菌株的选择。在第一个具体目标中，我们将进一步 确定蛋白质泛素化在调节内质之间的物理联系中的作用 网状结构和线粒体，并作为多蛋白质组装的成核部位 参与脂类运动的复合体。在第二个具体目标中，我们将把我们的发现从 以前的遗传学和蛋白质-蛋白质相互作用研究，以重建生物之间的联系 和人工合成的供体膜；以及从机械上定义单个蛋白质和脂肪如何 调节特定氨基环磷脂的运输。在第三个具体目标中，我们将继续 应用遗传和生化工具阐明与出口有关的基因和基因产品 线粒体和高尔基体中的磷脂酰乙醇胺。在第四个具体目标中，我们将使用 最近开发的磷脂酰丝氨酸脱羧酶的条件等位基因在小鼠中表达，以定义 这种酶在哺乳动物系统中线粒体生物发生中的作用。基因和基因的结合 我们提出的生化方法将提供新的机制和分子信息 磷脂在真核细胞中的转运过程，为研究磷脂的调节提供了新的见解 膜生物发生。这项基础性工作与人类疾病有关的领域最为相关 控制细胞生长和线粒体功能障碍，如癌症和心脏病。

A fundamental unsolved problem of cell biology and biochemistry is the molecular definition of the mechanisms of phospholipid transport for membrane biogenesis. This process is essential for all cell growth, replication, differentiation and homeostasis. The focus of this proposal is to use the power of yeast molecular genetics to dissect the process and understand the general principles of how it occurs throughout eukaryotic organisms. We have made progress in defining several of the genes and gene products that participate in the transport reactions, but large gaps in our knowledge still remain. The work will continue to concentrate on the transport reactions involving the aminoglycerophospholipids (phosphatidylserine, phosphatidylethanolamine and phosphatidylcholine), since the tools we have previously developed greatly facilitate measuring inter-membrane transfer events, and also provide the basis of genetic screens and selections for strains that are defective in these transport processes. In the first Specific Aim we will further define the role of protein ubiquitination in regulating physical associations between the endoplasmic reticulum and the mitochondria, and in serving as a nucleation site for the assembly of multi-protein complexes involved in moving the lipids. In the second Specific Aim we will extend our findings from previous genetic, and protein-protein interaction studies, to reconstitute the associations between biological and synthetic donor membranes in vitro; and mechanistically define how individual proteins and lipids regulate the transport of specific aminoglcyerophospholipids. In the third Specific Aim we will continue to apply genetic and biochemical tools to elucidate the genes and gene products involved in the export of phosphatidylethanolamine from mitochondria and the Golgi apparatus. In the fourth Specific Aim we will use recently developed conditional alleles for phosphatidylserine decarboxylaseexpressed in mice, to define the function of this enzyme in mitochondrial biogenesis in mammalian systems. The combined genetic and biochemical approaches we propose will provide new mechanistic and molecular information about phospholipid transport processes in eukaryotic cells and provide new insights into the regulation of membrane biogenesis. This fundamental work is most relevant to areas of human disease concerned with control of cell growth and mitochondrial dysfunction, such as cancer and heart disease.