Structural analysis of alcohol-dependent activation of GIRKs

GIRK 酒精依赖性激活的结构分析

• 批准号: 7987821
• 负责人: SENYON CHOE
• 金额: $ 45.24万
• 依托单位: SALK INSTITUTE FOR BIOLOGICAL STUDIES
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-10 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7987821
• 关键词: Alcohol abuse; Alcohol consumption; Alcohol dependence; Alcohol withdrawal syndrome; Alcoholism; Alcohols; Animal Model; Behavior; Binding; Biology; Brain; Cessation of life; Chemicals; Chromosomes, Human, Pair 1; Complex; Cysteine; Development; Drug Addiction; Electrophysiology (science); Ethanol; Ethanol dependence; Exhibits; Foundations; Future; GIRK2 subunit, G protein-coupled inwardly-rectifying potassium channel; GIRK3 subunit, G protein-coupled inwardly-rectifying potassium channel; GTP-Binding Proteins; Genes; Goals; Grant; Health; Heavy Drinking; Human; Institutes; Ion Channel; Kinetics; Knockout Mice; Knowledge; Lead; Ligands; Mediating; Modeling; Modification; Molecular; Mus; Mutagenesis; Mutation; Neurons; Pain; Paper; Pentobarbital; Pharmacologic Substance; Physiological; Potassium; Potassium Channel; Predisposition; Proteins; Publications; Publishing; Quantitative Trait Loci; Receptor Signaling; Research; Research Institute; Research Proposals; Resolution; Role; Self-Administered; Seminal; Signaling Molecule; Structure; Testing; Texas; United States; Universities; Wild Type Mouse; Withdrawal; Work; alcohol behavior; alcohol effect; alcohol research; alcohol response; austin; base; economic cost; experience; genetic regulatory protein; hypnotic; innovation; insight; interdisciplinary approach; interest; inward rectifier potassium channel; mutant; novel; public health relevance; relating to nervous system; research study; response; sedative; structural biology; tool

DESCRIPTION (provided by applicant): Alcohol (ethanol) consumption alters neural activity in the brain by modulating different types of ion channels. An emerging concept in the field is that some of the physiological effects of ethanol are mediated by direct modulation of ion channels in the brain. One of the targets in the brain for ethanol is the G protein-gated inwardly rectifying potassium (GIRK) channel, which is activated by ethanol. Mice lacking GIRK2 channels exhibit diminished ethanol-induced tolerance to pain and self-administer more ethanol than wild-type mice. Moreover, a quantitative trait loci with a large effect on predisposition to sedative withdrawal, such as from ethanol, was narrowed to a region on chromosome 1 in mice that contains Girk3 gene. GIRK3 knockout mice exhibit less severe sedative-hypnotic withdrawal. Though GIRK2 and GIRK2/3 channels are implicated in ethanol-related behaviors, the molecular mechanism underlying this response is not well understood. Recently, we showed with high-resolution structural studies that alcohols bind directly to hydrophobic pockets of inwardly rectifying potassium channels. Mutations in the alcohol-binding pocket of GIRK2 channels significantly alter ethanol activation. We hypothesize that ethanol binds to hydrophobic pockets in GIRK2/3 channels and facilitate a conformational change that is relayed to the channel's gate and opens the channel. In this research proposal, we plan to use an innovative approach of high-resolution crystallographic studies, structure-based mutagenesis and advanced electrophysiological recordings to investigate this hypothesis. Specifically, we will conduct a structure-function analysis of the ethanol-binding pocket in GIRK2/3 channels (1), solve high-resolution structures of GIRK channels complexed with ethanol to reveal conformational changes in the channel protein that occur with ethanol-dependent gating (2), and elaborate mechanistic models for ethanol-dependent activation of GIRK channels, utilizing single-channel recordings and chemical modification of cysteine-substituted channels (3). Completion of these proposed experiments will reveal the structural basis of ethanol modulation of GIRK channels, which will provide insights into the mechanism of ethanol-modulation of other types of ion channels. Understanding the molecular mechanism underlying ethanol modulation of ion channels could lead to development of novel pharmaceutical agents for treating alcohol-dependence, directly benefiting human health. PUBLIC HEALTH RELEVANCE: Ethanol, a major drug of addiction and abuse in the U.S., directly modulates brain ion channels, which control the excitability of brain neurons. The goal of this grant is to investigate the molecular and structural mechanisms underlying ethanol-dependent activation of neuronal potassium channels. Results from these studies could lead to the development of novel pharmaceutical agents that specifically modulate potassium channels or antagonize actions of ethanol.

描述（由申请人提供）：酒精（乙醇）消耗通过调节不同类型的离子通道来改变大脑中的神经活动。该领域的一个新兴概念是，乙醇的某些生理作用是由大脑中离子通道的直接调节介导的。大脑中乙醇的靶标之一是G蛋白门控的内部整流钾（Girk）通道，该通道被乙醇激活。缺乏GIRK2通道的小鼠比野生型小鼠表现出乙醇诱导的对疼痛的耐受性和自助剂的耐受性。此外，对镇静性戒断的易感性影响很大的定量性状基因座（例如从乙醇）被缩小到含有GIRK3基因的小鼠1染色体上的区域。 GIRK3敲除小鼠表现出较低的镇静性催眠戒断。尽管GIRK2和GIRK2/3通道与乙醇相关的行为有关，但该反应的基础机制尚不清楚。最近，我们通过高分辨率结构研究表明，醇直接与内向整流的钾通道的疏水袋结合。 GIRK2通道的酒精结合口袋中的突变会显着改变乙醇激活。我们假设乙醇在GIRK2/3通道中与疏水口袋结合，并促进了构象变化，该构象变化被传达到通道的栅极并打开通道。在这项研究建议中，我们计划使用高分辨率晶体学研究，基于结构的诱变和先进的电生理记录的创新方法来研究这一假设。 Specifically, we will conduct a structure-function analysis of the ethanol-binding pocket in GIRK2/3 channels (1), solve high-resolution structures of GIRK channels complexed with ethanol to reveal conformational changes in the channel protein that occur with ethanol-dependent gating (2), and elaborate mechanistic models for ethanol-dependent activation of GIRK channels, utilizing single-channel recordings and chemical modification of半胱氨酸取代的通道（3）。这些提出的实验的完成将揭示Girk通道的乙醇调节的结构基础，这将为其他类型的离子通道的乙醇调节机理提供见解。了解离子通道的乙醇调节的分子机制可能会导致发展新型药物以治疗酒精依赖性，直接使人类健康受益。 公共卫生相关性：乙醇是美国成瘾和滥用的主要药物，直接调节脑离子通道，该通道控制着脑神经元的兴奋性。该赠款的目的是研究神经元钾通道的乙醇依赖性激活所基于的分子和结构机制。这些研究的结果可能导致新型药物的发展，这些药物专门调节钾通道或对抗乙醇的作用。