YEAST LATE GOLGI MEMBRANE PROTEIN RETENTION

酵母晚期高尔基体膜蛋白保留

• 批准号: 2192806
• 负责人: Steven F Nothwehr
• 金额: $ 9.56万
• 依托单位: UNIVERSITY OF MISSOURI-COLUMBIA
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-09-30 至 2000-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2192806
• 关键词: Golgi apparatus; Saccharomyces cerevisiae; affinity chromatography; aminopeptidase; antibody; eukaryote; gel mobility shift assay; gene complementation; gene expression; genetic mapping; intracellular transport; laboratory rabbit; membrane proteins; molecular cloning; nucleic acid sequence; phenotype; protein signal sequence; protein structure function; protein transport; western blottings

The research outlined in this proposal is directed towards the general question of how membrane proteins are properly sorted and localized to specific eukaryotic cellular organelles. The biogenesis and maintenance of organelles, a poorly understood process dependent on correct localization and retention of membrane proteins, is critical for proper cell function. In addition, several human diseases (e.g. cystic fibrosis and familial hypercholesterolaemia) arise due to defects in the sorting of membrane proteins. While many membrane protein sorting events have been identified in eukaryotic cells, little is known of the machinery responsible for recognizing and localizing membrane proteins to the correct cellular compartment. This proposal describes a multi-faceted approach to characterize the mechanism by which the Golgi apparatus retains its resident Golgi membrane proteins. This analysis has been conducted in the yeast, S. cerevisiae, because this eukaryotic organism is easily amenable to genetics. The late Golgi enzyme, dipeptidyl aminopeptidase (DPAP) A, has been shown to contain a retention signal it its cytoplasmic domain which is both necessary and sufficient for its localization in the Golgi apparatus. In order to identify the cellular machinery that recognizes this signal and retains DPAP A in the Golgi, a genetic screen has been developed to identify mutants which fail to properly retain their Golgi membrane proteins. The grd (Golgi retention defective) mutants identified thus far fall into 10 complementation groups. The elucidation of the role of the individual grd gene products in the retention of Golgi membrane proteins promises to illuminate the Golgi retention mechanism and in particular may provide the basis for identification of the DPAP A retention signal receptor. Independent genetic and biochemical approaches will also be pursued that more directly focus on identification of the retention signal receptor. Experiments to determine whether or not retention occurs via retrieval from a post-Golgi compartment will also be performed. The results of these studies should help illuminate the mechanism of yeast Golgi membrane protein retention and add significantly our overall knowledge of membrane protein sorting.

该提案中概述的研究针对一般 膜蛋白如何正确分类和本地化的问题 特定的真核细胞细胞器。生物发生和维护 细胞器，这是一个依赖正确本地化的过程知识的过程 膜蛋白的保留对于适当的细胞功能至关重要。 此外，几种人类疾病（例如囊性纤维化和家族性 高胆固醇血症是由于膜分类的缺陷而引起的 蛋白质。虽然已经确定了许多膜蛋白分类事件 在真核细胞中，对负责的机械知之甚少 将膜蛋白识别和定位到正确的细胞 车厢。 该建议描述了一种多方面的方法来表征 高尔基仪器保留其常驻高尔基膜的机制 蛋白质。该分析是在酵母菌酿酒酵母中进行的 因为这种真核生物很容易受到遗传学。已故 Golgi酶，二肽基氨基肽酶（DPAP）A已显示为 包含保留信号，其细胞质结构域是 必要且足够在高尔基体设备中定位。在 为了识别识别此信号的蜂窝机械和 保留DPAP A中的Golgi，已经开发了一个遗传屏幕 识别无法正确保留高尔基膜的突变体 蛋白质。到目前为止确定的GRD（高尔基保留缺陷）突变体 分为10个互补组。阐明了 单个GRD基因产物在高尔基膜蛋白的保留中 有望照亮高尔基人保留机制，尤其是可能 为识别DPAP保留信号提供了基础 受体。独立的遗传和生化方法也将是 追求更直接地专注于识别保留信号 受体。实验以确定保留是否通过 还将从后高尔基室检索。这 这些研究的结果应有助于阐明酵母的机制 高尔基膜蛋白保留率，并显着增加我们的总体 了解膜蛋白分类。