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Trafficking pathways to the cell surface in yeast

酵母细胞表面的运输途径

基本信息

批准号：
7925130
负责人：
AMY Y CHANG
金额：
$ 11.51万
依托单位：
UNIVERSITY OF MICHIGAN AT ANN ARBOR
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-09-30 至 2011-03-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): We propose to study two major mechanisms that regulate the structure and function of the plasma membrane: ER quality control and lipid rafts. We have used mutants of PMA1 encoding the yeast plasma membrane H+ATPase as tools to understand normal membrane biogenesis as well as the molecular mechanisms of regulation at the ER and at the plasma membrane. The dominant negative Pma1-D378N mutant is recognized and targeted for destruction by the ER-associated degradation (ERAD) pathway, whereas the temperature-sensitive Pma1-10 mutant fails to associate with lipid rafts and is removed from the plasma membrane for vacuolar degradation. Each pma1 mutant has been used as the basis for genetic selection leading to identification of Eps1, a novel component of the ERAD pathway, and Yvh1, a dual-specificity phosphatase, which may play a role in quality control at the cell surface. This proposal integrates biochemical, cell biological and genetic approaches to study the molecular mechanisms of these novel proteins. Specific Aim I addresses a proposed role for Eps1, a member of the protein disulfide isomerase family, in substrate recognition during ERAD. In addition, the ERAD substrate, Pma1-D378N, will be analyzed in detail because it represents a major resource for studying ERAD. Specific Aim II uses the pma1-10 mutant and its suppressor yvh1 to study the relationship between protein phosphorylation, ubiquitination, association with lipid rafts, and protein stability. Our studies have important implications for understanding human diseases caused by misfolding and ERAD of important molecules, including cystic fibrosis. Our results on lipid rafts will have significance for understanding the immune response (during which signaling occurs in rafts), AIzheimer's disease (in which protein processing is raft-based), and lipid storage diseases (in which raft lipids are abnormally accumulated in lysosomes).

描述（由申请人提供）：我们建议研究调节质膜结构和功能的两种主要机制：ER质量控制和脂筏。我们已经使用的突变体的PMA 1编码的酵母质膜H+ ATP酶的工具，以了解正常的膜生物合成以及在ER和质膜的分子调控机制。显性负性Pma 1-D378 N突变体被ER相关降解（ERAD）途径识别并靶向破坏，而温度敏感性Pma 1 -10突变体不能与脂筏结合，并从质膜上除去进行液泡降解。每个pma 1突变体已被用作遗传选择的基础，导致ERAD途径的一个新的组成部分Eps 1和Yvh 1，双特异性磷酸酶，这可能在细胞表面的质量控制中发挥作用的鉴定。该建议整合了生物化学，细胞生物学和遗传学方法来研究这些新蛋白质的分子机制。具体目标我地址的Eps 1，蛋白质二硫键异构酶家族的成员，在ERAD过程中的底物识别的作用。此外，ERAD底物Pma 1-D378 N将被详细分析，因为它代表了研究ERAD的主要资源。Specific Aim II使用pma 1 -10突变体及其抑制因子yvh 1来研究蛋白磷酸化、泛素化、与脂筏的结合以及蛋白稳定性之间的关系。我们的研究对于理解由重要分子的错误折叠和ERAD引起的人类疾病具有重要意义，包括囊性纤维化。我们对脂筏的研究结果将对理解免疫反应（在此期间信号发生在筏），阿尔茨海默病（其中蛋白质加工是筏为基础的）和脂质储存疾病（其中筏脂质异常积累在溶酶体中）具有重要意义。