Role of PICK1 in AMPA Receptor Transport

描述（由申请人提供）：我们将研究PICK1（与C激酶-1相互作用的蛋白）及其调节因子NSF （n -乙基马酰亚胺敏感因子）在AMPA受体（AMPAR）运输中的作用。AMPAR是中枢神经系统兴奋电流的主要来源，AMPAR突触丰度的变化调节着突触强度。PICK1和NSF结合到AMPAR GluR2亚基的c端细胞质结构域，在控制AMPAR突触水平方面具有相反的作用。PICK1通过PDZ结构域与GluR2结合，并在海马锥体神经元的核内体样囊泡中与AMPAR共定位。PICK1的表达降低了AMPAR突触膜的丰度，并且与长期抑制（LTD）的机制有关，这些都表明PICK1在AMPAR内吞作用中起作用。NSF与SNAP蛋白一起以解离SNARE复合物而闻名。在AMPAR中，NSF与GluR2的相互作用对于维持AMPAR电流和突触丰度是必要的。我们最近的工作揭示了NSF的一种新功能。NSF与PICK1-GluR2复合物的SNAPs结合在一起，并从GluR2中置换PICK1。通过取代一个内吞因子，NSF维持突触中的AMPAR。本项目的目的是：1)确定GluR2-PICK1-NSF-SNAP复合物组装和NSF拆卸的结构基础。我们将定义稳定复合物的蛋白质-蛋白质接触，并确定在拆卸过程中snap是否将旋转扭矩从NSF传递到PICK1。我们将确定α -和β - snaps破坏PICK1-GluR2复合物的不同能力的基础。2)确定PICK1在AMPAR贩运中的作用。我们将确定起始膜和终点膜以及G蛋白在转运机制中的作用。3)通过生理刺激确定体内调控，包括PICK1在GluR2/GluR3组成循环中的作用。这些研究将揭示突触调控和记忆形成的基本机制。

DESCRIPTION (provided by applicant): We will study the roles of PICK1 (Protein Interacting with C Kinase-1) and its regulator, NSF (N-ethylmaleimide Sensitive Factor) in AMPA receptor (AMPAR) trafficking. AMPAR are the major source of excitatory currents in the CNS and changes in AMPAR synaptic abundance regulate synaptic strength. PICK1 and NSF bind to the C-terminal, cytoplasmic domain of the AMPAR GluR2 subunit, and have opposing roles in controlling AMPAR synaptic levels. PICK1 binds to GluR2 via a PDZ domain and colocalizes with AMPAR in endosome-like vesicles in hippocampal pyramidal neurons. PICK1 expression reduces AMPAR synaptic membrane abundance and PICK is implicated in the mechanism of long term depression (LTD), all indicating a role for PICK1 in AMPAR endocytosis. NSF, together with the SNAP proteins, is known for dissociating SNARE complexes. In the case of AMPAR, the interaction of NSF with GluR2 is necessary for maintaining AMPAR currents and synaptic abundance. Our recent work has revealed a novel function for NSF. NSF binds together with the SNAPs to the PICK1-GluR2 complex and displaces PICK1 from GluR2. By displacing an endocytosis factor, NSF maintains AMPAR in the synapse. The Aims of this project are: 1) To determine the structural basis for assembly of the GluR2-PICK1-NSF-SNAP complex and its disassembly by NSF. We will define protein-protein contacts that stabilize the complex and determine whether the SNAPs transmit rotational torque from NSF to PICK1 during disassembly. We will determine the basis for the differential ability of alpha- and beta-SNAPs to disrupt PICK1-GluR2 complexes. 2) To determine the role of PICK1 in AMPAR trafficking. We will identify the starting and destination membranes and the roles of G proteins in the transport mechanism. 3) To determine regulation in vivo by physiologic stimuli, including the role of PICK1 in GluR2/GluR3 constitutive recycling. These studies will reveal basic mechanisms relevant to synapse regulation and memory formation.