SYNAPTIC FUNCTION IN THE NEMATODE C. ELEGANS

描述：（来自申请人的摘要）拟议研究的目标是 识别和理解突触所需分子的功能， 功能近年来，我们看到这一领域取得了很大进展， 突触蛋白质已经通过生物化学手段鉴定。但据 目前尚不清楚这些蛋白质中的每一种如何在突触中发挥作用，也不清楚它们的全部功能。 参与这一基本生物学过程的蛋白质已被鉴定出来。 我们已经在小杆线虫中鉴定了突触功能突变体 优雅其中一些突变存在于已知的突触蛋白中。之中 我们建议集中在三种蛋白质上，这些蛋白质被认为在 突触囊泡的内吞作用。我们将描述 这些突变体的细节，从而测试模型，已提出的 这些蛋白质的功能基于生化数据。其他突变 在我们的筛选中发现的是编码新蛋白质的基因。我们将 在分子水平上表征这些蛋白质之一，并确定 这种蛋白质的功能在突触传递的特点突变 表型的详细信息。目标1. NSF。有人建议，NSF可以在 在胞吐作用的融合步骤处，或在胞吐作用的预对接步骤处， 内吞作用之前的融合后步骤。我们将测试NSF的假设， 在后融合步骤中是需要的;具体地， 囊泡蛋白突触泡蛋白的内吞作用。目标2. AP 2的u2亚基。的 网格蛋白衔接子复合物AP 2被认为是内吞 突触囊泡然而，目前尚不清楚突触囊泡循环是否 仅通过网格蛋白介导的过程发生。也有可能 突触使用AP 2的一个专门的中型亚单位，而不是普遍存在的u2 中亚基我们将测试是否需要AP 2的u2中亚基 突触囊泡内吞作用。目标3. Synaptojanin。已经提出 肌醇磷酸酶synaptojanin是内吞 突触囊泡，因为它与内吞机制有关。我们 将确定突触内吞是否需要突触结蛋白， 囊泡成分从质膜。此外，我们的初步 研究表明，它可能是运输囊泡所必需的。 突触的前体我们将测试一个模型， synaptojanin在囊泡运输中的功能。目标4。RIC-7。在这方面的突变 基因干扰胆碱能和GABA能介导的行为。我们将确定 利用超微结构和 生理工具。

DESCRIPTION: (from applicant's abstract) The goal of the proposed research is to identify and understand the function of the molecules required for synaptic function. In recent years we have seen great progress in this field as a number of synaptic proteins have been identified by biochemical means. However, it is not clear how each of these proteins functions in the synapse nor have all of the proteins involved in this fundamental biological process been identified. We have identified synaptic function mutants in the nematode Caenorhabditis elegans. Some of these mutations are in known synaptic proteins. From among these we propose to focus on three proteins which are believed to function in the endocytosis of synaptic vesicles. We will characterize the phenotype of these mutants in detail and thereby test models which have been proposed for the function of these proteins based on biochemical data. Other mutations identified in our screens are in genes encoding novel proteins. We will characterize one of these proteins at a molecular level and determine the function of this protein in synaptic transmission by characterizing the mutant phenotype in detail. Aim 1. NSF. It has been proposed that NSF may function at a pre-docking step of exocytosis, at the fusion step of exocytosis, or at a post-fusion step preceding, endocytosis. We will test the hypothesis that NSF is required at a post-fusion step; specifically, that it is required for the endocytosis of the vesicle protein synaptobrevin. Aim 2. u2 subunit of AP2. The clathrin adaptor complex AP2 is believed to be required for endocytosis of synaptic vesicles. However, it is not clear whether synaptic vesicle recycling occurs solely via a clathrin-mediated process. It is also possible that the synapse uses a specialized medium subunit of AP2 rather than the ubiquitous u2 medium subunit. We will test whether the u2 medium subunit of AP2 is required for synaptic vesicle endocytosis. Aim 3. Synaptojanin. It has been proposed that the inositol phosphatase synaptojanin is required for endocytosis of synaptic vesicles because it is associated with the endocytic machinery. We will determine whether synaptojanin is required for endocytosis of synaptic vesicle components from the plasma membrane. In addition, our preliminary studies indicate that it may be required for the transport of vesicle precursors to the synapse. We will test a model for the mechanism of synaptojanin function in vesicle transport. Aim 4. RIC-7. Mutations in this gene disrupt cholinergic and GABAergic-mediated behaviors. We will determine the function of this novel synaptic protein using ultrastructural and physiological tools.