SORTING OF PROTEINS IN REGULATED SECRETORY CELLS

调节分泌细胞中的蛋白质排序

• 批准号: 3287617
• 负责人: HSIAO-PING H MOORE
• 金额: $ 12.1万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 1985
• 资助国家: 美国
• 起止时间: 1985-07-01 至 1988-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3287617
• 关键词: Golgi apparatus; adrenocorticotropic hormone; affinity chromatography; antibody formation; cell cell interaction; cell type; complementary DNA; genetic manipulation; histochemistry /cytochemistry; hormone regulation /control mechanism; immunoelectron microscopy; immunofluorescence technique; immunoprecipitation; intracellular membranes; molecular biology; molecular cloning; monoclonal antibody; mutagens; neoplastic cell culture for noncancer research; nucleic acid sequence; proinsulin; proteolysis; radioimmunoassay; secretion; somatotropin; vesicle /vacuole

The general aim of this proposal is to understand how proteins are targeted to their correct intracellular compartment after synthesis. Specifically, the tumor cell line, AtT-20, derived from the mouse anterior pituitary, will be used to study assembly of the secretory granules which are involved in regulated secretion of peptide hormones. Recent evidence suggests that cells with regulated secretory function (i.e. neural, endocrine and exocrine cells) sort their secretory and plasma membrane proteins into distinct secretory pathways for externalization. Thus these cells have a regulated pathway in which specialized secretory products are stored in dense core secretory granules. Release of these products is regulated by secretagogues. In contrast, the cells utilize a second, constitutive, pathway for transporting other plasma membrane and secreted proteins to the cell surface. Externalization from this pathway is not regulated, and does not involve storage in the secretory granules. It is the aim of this proposal to understand how proteins are sorted in these cells such that only a subset of proteins are transported to the secretory granules. First, sorting signals will be sought on proteins targeted to the regulated pathway. This will be achieved by in vitro mutagenesis of DNA sequences encoding secretory granule proteins, followed by expressing the altered DNAs in AtT-20 cells to examine their effects on targeting. We will seek to determine if a common sorting signal is shared by secretory granule proteins from the different cell types which are capable of regulated secretion. Secondly, the temporal and the spatial relationships of sorting to other posttranslational processes will be examined by DNA transfection and immunoelectron microscopy. Finally, we will attempt to identify the cellular machinery involved in sorting by affinity chromatography and monoclonal antibody techniques.

这项提议的总体目标是了解蛋白质是如何被靶向的