MOLECULAR STUDIES OF SELECTIVE PROTEIN TRANSPORT

选择性蛋白质运输的分子研究

• 批准号: 6151044
• 负责人: Gregory S Payne
• 金额: $ 29.11万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-02-01 至 2001-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6151044
• 关键词: Golgi apparatus; Saccharomyces cerevisiae; biological signal transduction; clathrin; endocytosis; gene mutation; intracellular transport; membrane proteins; molecular cloning; nucleic acid sequence; pheromone; protein structure function; protein transport; receptor; yeast two hybrid system

DESCRIPTION: The compartmental organization of eukaryotic cells relies on the accurate distribution of proteins to different compartments and on retention of proteins within each compartment. The ability to sort proteins with different destinations is a key feature of the transport machinery which governs protein distribution and retention. The molecular basis of this selectivity is the focus of this proposal. Studies of yeast strains carrying mutations in the clathrin heavy chain subunit revealed that clathrin participates in selective protein transport processes at the plasma membrane and the Golgi complex: endocytosis of the mating pheromone receptors, sorting of vacuolar precursors from the Golgi complex to vacuoles, and a previously unrecognized role in localization of membrane proteins to a late Golgi compartment. Additional proteins have been identified which function in Golgi membrane protein localization and a collection of mutants with defects in this process have been isolated. These studies form the foundation for a combination of genetic, cell biology and biochemical approaches to investigate three aspects of selective protein transport: 1) the mechanism of Golgi membrane protein localization; 2) the role played by clathrin-associated proteins in clathrin-dependent protein transport processes; 3) the mechanism of receptor internalization during receptor-mediated endocytosis of mating peptide pheromones. Mutants (tcs) with defects in Golgi membrane protein localization will be used to clone wild-type versions of genes whose protein products participate in the localization process. The localization of TCS-encoded proteins will be determined in wild-type and mutant strains to explore interrelationships between the Tcs proteins and proteins involved in clathrin-coated vesicle formation. To define the function of clathrin coats in Golgi membrane protein localization, gene disruptions will be generated to eliminate all subunits of a clathrin-associated protein (AP-l) complex which is postulated to play a central role in the formation of clathrin-coated vesicles at the Golgi complex. Genetic strategies will be employed to identify functionally redundant AP subunits and other proteins which interact with AP-l. Finally, a novel class of endocytosis targeting signals will be used as probes to isolate proteins responsible for selective packaging of receptors into nascent endocytic vesicles. Together, these approaches will identify previously unrecognized components of the selective protein transport machinery and address the specific roles played by each identified participant.

描述：真核细胞的隔室组织依赖 蛋白质的准确分布到不同的隔室以及 在每个隔间内保留蛋白质。 分类蛋白质的能力 与不同的目的地是运输机械的关键特征 控制蛋白质分布和保留率。 分子基础 这种选择性是该提议的重点。 对网状蛋白重链中携带突变的酵母菌菌株的研究 亚基揭示了网格蛋白参与选择性蛋白的转运 质膜和高尔基体络合物的过程：内吞作用 交配信息素受体，从高尔基体分类液泡前体 复杂到液泡，以及以前在本地化中的作用 膜蛋白到晚期高尔基体。 其他蛋白质具有 已确定在高尔基膜蛋白定位中哪个功能和A 在此过程中，具有缺陷的突变体的集合已被隔离。 这些研究构成了遗传，细胞生物学结合的基础 以及研究选择性蛋白的三个方面的生化方法 运输：1）高尔基膜蛋白定位的机理； 2） 网格蛋白相关蛋白在网格蛋白依赖性蛋白中扮演的作用 运输过程； 3）受体内在化的机制 受体介导的交配肽信息素的内吞作用。 高尔基膜蛋白定位中缺陷的突变体（TC）将是 用于克隆蛋白质产品的基因的野生型版本 在本地化过程中。 TCS编码蛋白的定位将 在野生型和突变菌株中确定以探索相互关系 TCS蛋白和参与网格蛋白涂层囊泡的蛋白质之间 形成。 定义高尔基膜中网格蛋白外套的功能 蛋白质定位，将产生基因破坏以消除所有 网格蛋白相关蛋白（AP-L）复合物的亚基，该蛋白质被假设 在形成网格蛋白涂层囊泡的核心作用 高尔基综合体。 将采用遗传策略来识别功能 冗余AP亚基和其他与AP-L相互作用的蛋白质。 最后， 一类新型的内吞作用信号将用作探针 分离蛋白质，负责选择性包装受体 新生的内吞囊泡。 这些方法将共同确定 选择性蛋白转运的先前未识别的成分 机械和解决每个确定的特定角色 参与者。