Molecular Mechanisms, Modulation, and Synaptic Organization of Kainate Receptors

红藻氨酸受体的分子机制、调节和突触组织

• 批准号: 10274561
• 负责人: Joel Reuben Meyerson
• 金额: $ 36.32万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-06-03 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10274561
• 关键词: AMPA Receptors; Address; Area; Biochemical; Biology; Brain; Cations; Characteristics; Complex; Cryoelectron Microscopy; Data; Development; Drug Targeting; Electrophysiology (science); Epilepsy; Family; Fluorescence; Glutamate Receptor; Glutamates; Goals; Homo; Human; Ion Channel; Kainic Acid Receptors; Kinetics; Knowledge; Mediating; Molecular; Mood Disorders; N-Methyl-D-Aspartate Receptors; Nature; Neurotransmitters; Physiological; Physiology; Positioning Attribute; Preparation; Property; Proteins; Research; Resolution; Role; Shapes; Structure; Synapses; Synaptic Transmission; System; Work; desensitization; gamma-Aminobutyric Acid; insight; member; millisecond; neurotransmitter release; novel therapeutics; pain perception; programs; receptor; single molecule

Molecular Mechanisms, Modulation, and Synaptic Organization of Kainate Receptors Kainate receptors (KARs) are members of the ionotropic glutamate receptor (iGluR) family of cation channels, which also includes AMPA and NMDA receptors. They localize to synapses where they respond to L- glutamate (L-Glu) which is the most abundant excitatory neurotransmitter in the brain. KARs serve a canonical role in synaptic depolarization, and non-canonical roles in modulating L-Glu and GABA release and synapse maturation. They are involved in pain perception, epilepsy and mood disorders which has made them important drug targets. Unfortunately, our understanding of KAR molecular mechanisms is limited without structures for the most abundant KAR complexes in the brain. My research program aims to address this gap by developing molecular mechanisms for KAR gating, modulation, and organization. KARs assemble as tetramers from a pool of five subunits (GluK1-5). After assembly and positioning at the synapse, they respond to synaptic L-Glu by opening their ion channel and, under the sustained presence of L-Glu, desensitize within milliseconds to close their channel. A goal of my lab is to understand how KAR subunits are organized in the receptor, and how the subunits coordinate together to control the ion channel. My recent cryo-electron microscopy (cryo-EM) work revealed the surprising organization, symmetries, and domain interfaces of homo-tetrameric GluK2 (Meyerson et al. 2014 Nature; Meyerson et al. 2016 Nature). However, these results are of limited physiological value because native KARs in the brain are hetero-tetramers composed of GluK2 and GluK5 subunits. The GluK2/K5 hetero-tetramer has strict assembly rules and unique kinetic properties which underpin KAR physiology, but little is known of its subunit organization, or activation and desensitization mechanisms. Our preliminary data show successful expression, purification, and initial structure determination of the GluK2/K5 heteromer to 3.7 Å resolution with cryo-EM. In research Area 1 we aim to determine how GluK2/K5 organizes and functions by using a combination of single molecule fluorescence, electrophysiology, and cryo-EM. KARs do not function in isolation. Synaptic KAR complexes incorporate transmembrane Neto2 auxiliary proteins which shapes their characteristic kinetic profile. KARs also attach to C1ql2 proteins which are required for proper receptor localization. Understanding how KARs interact with these proteins is essential to bridging the gap between molecular mechanism and synapse biology and is a major research goal. Towards this end we have developed new biochemical preparations which enable detailed mechanistic interrogation on these systems. In Area 2 we will determine how KARs interact with and are modulated by C1ql2 and Neto2. I hypothesize that our work in research Areas 1 and 2 will give important insights into KAR physiology.

红藻氨酸受体的分子机制、调节和突触组织 红藻氨酸受体（KAR）是离子型谷氨酸受体（iGluR）家族的成员， 通道，其中还包括AMPA和NMDA受体。它们定位于突触，在那里它们对L- 谷氨酸（L-Glu）是大脑中最丰富的兴奋性神经递质。KAR服务于一个规范 在突触去极化中的作用，以及在调节L-Glu和GABA释放和突触 成熟他们参与疼痛感知，癫痫和情绪障碍，这使得他们 重要的药物靶点不幸的是，我们对KAR分子机制的理解是有限的， 大脑中最丰富的KAR复合物的结构。我的研究计划旨在解决 通过开发KAR门控、调制和组织的分子机制来弥补这一差距。 KAR从五个亚基（GluK 1 -5）的库中组装成四聚体。组装和定位后， 在突触中，它们通过打开它们的离子通道来响应突触L-Glu，并且在持续存在的 L-Glu在几毫秒内降低敏感性关闭通道我实验室的一个目标是了解KAR 亚基在受体中的组织，以及亚基如何协调在一起来控制离子通道。 我最近的低温电子显微镜（cryo-EM）工作揭示了令人惊讶的组织，对称性， 同源四聚体GluK 2的结构域界面（Meyerson et al. 2014 Nature; Meyerson et al. 2016 Nature）。 然而，这些结果具有有限的生理学价值，因为脑中的天然KAR是异源四聚体 由GluK 2和GluK 5亚基组成。GluK 2/K5异源四聚体具有严格的组装规则和独特的 动力学特性是KAR生理学的基础，但对其亚基组织或激活知之甚少 和脱敏机制。我们的初步数据显示成功的表达，纯化和初始化。 GluK 2/K5异聚体的结构测定，用cryo-EM达到3.7 μ m分辨率。研究领域1 我们的目标是确定GluK 2/K5如何组织和功能，通过使用单一的组合， 分子荧光、电生理学和冷冻电镜。 KAR不是孤立地发挥作用的。突触KAR复合物掺入跨膜Neto 2 形成其特征动力学曲线的辅助蛋白。KAR还附着于C1 q12蛋白， 是正确定位受体所必需的。了解KAR如何与这些蛋白质相互作用至关重要 弥合分子机制和突触生物学之间的差距，是一个主要的研究目标。朝向 为此，我们开发了新的生物化学制剂，使详细的机械询问， 这些系统。在区域2中，我们将确定KAR如何与C1 q12相互作用并受C1 q12调节 Neto2。我假设我们在研究领域1和2的工作将对KAR提供重要的见解 physiology.