SORTING OF PROTEINS IN REGULATED SECRETORY CELLS

调节分泌细胞中的蛋白质排序

基本信息

批准号：
3287622
负责人：
HSIAO-PING H MOORE
金额：
$ 18.44万
依托单位：
UNIVERSITY OF CALIFORNIA BERKELEY
依托单位国家：
美国
项目类别：
财政年份：
1985
资助国家：
美国
起止时间：
1985-07-01 至 1993-06-30
项目状态：
已结题

项目摘要

The general aim of this proposal is to understand how proteins are targeted to their correct intracellular compartments after synthesis. Specifically, we will use the pituitary cell line, AtT- 2O, to study molecular events underlying the assembly of secretory granules. Recent evidence suggests that neuroendocrine cells possess a constitutive and a regulated pathway for secretion, and that proteins destined for the cell surface are selectively sorted into different secretory vesicles for externalization. How proteins select the proper organelles, and how the machinery of these two types of secretory vesicles differs from each other are not understood. In this proposal, we will try to identify the sorting signals that direct peptide hormones into regulated granules by in vitro mutagenesis and DNA transfection. We will also purify the sorting compartment and characterize them biochemically. To elucidate the cellular sorting mechanisms, we will isolate specific sorting proteins from the sorting compartment and study their interactions with the transported proteins. We will ask if different sorting proteins are used by different types of cells with regulated secretory functions, or whether these cells share a common sorting machinery. To identify other components involved in sorting, we will set up an in vitro system to assay sorting in permeabilized cells. This system will be used to assess the role of clathrin in regulated secretion. Finally, we will devise systems to dissect functional components of the granule that are involved in the temporal and spatial regulation of secretory activities.

该提议的总体目的是了解蛋白质的蛋白质针对其正确的细胞内隔室合成。具体而言，我们将使用垂体细胞系，Att- 2o，研究分泌组装的基础的分子事件颗粒。最近的证据表明神经内分泌细胞拥有构成和受管制的途径，并具有注定针对细胞表面的蛋白质选择性排序进入不同的分泌囊泡以进行外在化。如何蛋白质选择适当的细胞器，以及如何机械这两种类型的分泌囊泡彼此不同不理解。在此建议中，我们将尝试确定将将肽激素引导到受调节的信号中通过体外诱变和DNA转染的颗粒。我们将还净化分类舱并表征它们生化。为了阐明细胞分选机制，我们将从分类室分离特定的分类蛋白并研究他们与运输蛋白的相互作用。我们会询问是否使用不同类型的不同排序蛋白具有调节分泌功能的细胞，或这些细胞是否共享一种通用的分类机械。识别其他组件参与分类，我们将建立一个体外系统来分析在透化细胞中分类。该系统将用于评估网格蛋白在调节分泌中的作用。最后，我们会的设计系统以剖析颗粒的功能成分参与分泌的时间和空间调节活动。