Assembly Chaperone Complex for Membrane Proteins in the Endoplasmic Reticulum

内质网膜蛋白的组装伴侣复合物

• 批准号: 10302235
• 负责人: Tingwei Mu
• 金额: $ 6.87万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-01 至 2024-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10302235
• 关键词: Affinity Chromatography; Brain; Cell membrane; Cell surface; Complex; Endoplasmic Reticulum; Epilepsy; Equilibrium; Genetic; Hsp47 protein; Human; Ion Channel; Ion Channel Gating; Knowledge; Mass Spectrum Analysis; Membrane; Membrane Potentials; Membrane Proteins; Molecular Chaperones; Mutation; Neuraxis; Neurotransmitters; Organelles; Pathogenicity; Physiology; Process; Protein Subunits; Proteins; Proteomics; Quality Control; Research; Role; Surface; Testing; base; gamma-Aminobutyric Acid; loss of function; mutant; nervous system disorder; neural circuit; protein complex; protein misfolding; proteostasis; receptor; receptor function

Project Description There is a significant knowledge gap in understanding how multi-subunit membrane proteins are inserted, folded, and assembled in the membrane of the endoplasmic reticulum (ER), a cellular organelle for protein quality control with the assistance of molecular chaperones. We use γ- aminobutyric acid type A (GABAA) receptors to answer this question. The GABAA receptors, the primary inhibitory neurotransmitter-gated ion channels in the mammalian central nervous systems, are a pentameric protein complex. Despite extensive research on GABAA receptors physiology on the plasma membrane and their role in controlling the inhibition-excitation balance in neural circuits, little effort has been made to investigate how the protein homeostasis (proteostasis) network regulates the folding and assembly of GABAA proteins in the ER. This brings a significant barrier for the treatment of genetic epilepsies because numerous epilepsy-associated mutations in GABAA receptor subunits cause subunit protein misfolding in the ER and/or disrupt assembly of the pentameric complex, leading to decreased cell surface localization of the receptor complex and imbalanced neural circuits. Here, specifically, our affinity purification-mass spectrometry-based proteomics analysis identified a potential Membrane Protein Assembly Chaperone Complex (MPACC) that interacts with endogenous GABAA receptors, consisting of heat shock protein 47 (Hsp47) in the ER lumen and the ER membrane protein complex (EMC) in the ER membrane. In Aim 1, we will test our hypothesis that Hsp47 positively regulates the assembly of endogenous GABAA receptors in the ER and their functional surface expression. In Aim 2, we will test our hypothesis that the EMC positively regulates the assembly of GABAA receptors in the ER membrane and thus their functional surface expression. In Aim 3, we will manipulate the assembly chaperone complex in the ER to correct the function of pathogenic GABAA receptors.

项目描述