Mechanisms of membrane homeostasis through protein and lipid transport

项目概要/摘要 真核细胞必须维持质膜的特定蛋白质和脂质组成以及所有的细胞膜。 内部膜结合的细胞器，以正常运作。尽管膜生物发生是 作为生命的关键，建立膜的组成和组织的机制仍然很差 明白我们研究膜的不对称性是如何建立的，这是真核细胞的一个基本特征 质膜由磷脂酰丝氨酸和磷脂酰乙醇胺的富集所限定， 细胞质小叶中，而鞘脂和磷脂酰胆碱通常富集在细胞外小叶中， 双层。细胞外小叶上PS和PE的调节暴露有助于细胞信号传导， 胞质分裂、血液凝固、细胞-细胞融合、凋亡细胞尸体去除和宿主-病毒相互作用。膜 不对称性是由IV型P-型ATP酶（P4-ATP酶）驱动的，其是一个大家族的翻转酶， 脂质从细胞外小叶转移到膜双层的胞质小叶。P4-ATP酶亚家族是 在真核生物中高度保守，并且这些转运蛋白与病理条件有关 如肥胖相关的2型糖尿病、心血管疾病、肝病、听力损失、免疫缺陷， 和严重的神经系统疾病此外，P4-ATP酶是囊泡介导的细胞凋亡的关键组分。 高尔基复合体和内体系统内的蛋白质运输机制。通过它们在蛋白质中的作用 运输，P4-ATP酶帮助控制质膜的精确蛋白质组成，高尔基复合体 内体和溶酶体。这项研究将确定P4-ATP酶如何识别和 利用结构、生物化学和分子生物学技术， 基因方法。这些结构/功能研究将包括P4-ATP酶活性是如何通过后 翻译修饰和蛋白质-蛋白质相互作用。我们还将探讨细胞的要求， 细胞形态发生中特定底物脂质（如葡萄糖神经酰胺和磷脂酰丝氨酸）的转运， 真菌发病机理、营养信号传导和蛋白质运输。对于后面的研究，我们将探讨P4- ATP酶有助于驱动囊泡介导的蛋白质转运，重点是由COPI和逆转录酶形成的载体。 泛素化在P4-ATP酶和COPI依赖性转运途径中的非典型作用也将被讨论。 定义了总的来说，这些研究应该导致更好地了解P4-ATP酶如何发挥其作用。 重要的功能，并将是非常宝贵的，我们的能力，以了解和最终治疗相关的病理 缺乏P4-ATP酶

Project Summary/Abstract Eukaryotic cells must maintain a specific protein and lipid composition of the plasma membrane and all of the internal membrane-bound organelles in order to function normally. Even though membrane biogenesis is crucial for life, mechanisms for establishing the composition and organization of membranes remain poorly understood. We study how membrane asymmetry is established, a fundamental feature of the eukaryotic cell plasma membrane defined by the enrichment of phosphatidylserine and phosphatidylethanolamine within the cytosolic leaflet, while sphingolipids and phosphatidylcholine are typically enriched in the extracellular leaflet of the bilayer. Regulated exposure of PS and PE on the extracellular leaflet contributes to cell signaling, cytokinesis, blood clotting, cell-cell fusion, apoptotic cell corpse removal and host-viral interactions. Membrane asymmetry is driven by type IV P-type ATPases (P4-ATPases), which are a large family of flippases that pump lipids from the extracellular leaflet to the cytosolic leaflet of the membrane bilayer. The P4-ATPase subfamily is highly conserved among eukaryotes and these transporters have been implicated in pathological conditions such as obesity-linked type 2 diabetes, cardiovascular disease, liver disease, hearing loss, immune deficiency, and severe neurological disease. In addition, P4-ATPases are critical components of the vesicle-mediated protein trafficking machinery within the Golgi complex and endosomal system. Through their role in protein trafficking, P4-ATPases help control the precise protein composition of the plasma membrane, Golgi complex endosomes and lysosomes. The proposed studies will determine how the P4-ATPases recognize and transport their lipid substrates to establish membrane asymmetry using structural, biochemical and molecular genetic approaches. These structure/function studies will include how P4-ATPase activity is regulated by post- translational modification and protein-protein interactions. We will also probe the cellular requirements for transport of specific substrate lipids, like glucosylceramide and phosphatidylserine, on cell morphogenesis, fungal pathogenesis, nutrient signaling, and protein trafficking. For the latter studies, we will probe how P4- ATPases help drive vesicle-mediated protein transport with a focus on carriers formed by COPI and retromer. Atypical roles for ubiquitination in the P4-ATPase- and COPI-dependent transport pathways will also be defined. In total, these studies should lead to a much better understanding of how P4-ATPases exert their essential function, and will be invaluable to our ability to understand and ultimately treat pathologies associated with P4-ATPase deficiency.