Elucidating transsynaptic regulation of metabotropic glutamate receptors

阐明代谢型谷氨酸受体的跨突触调节

• 批准号: 10508656
• 负责人: William Grant LUDLAM
• 金额: $ 6.68万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-01-01 至 2025-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10508656
• 关键词: Autoreceptors; Binding; Biological Assay; Brain; Cell Adhesion Molecules; Cells; Cognitive; Complex; Coupled; Cryoelectron Microscopy; Deuterium; Dimerization; Disease; Environment; Epilepsy; Epitopes; Exposure to; Family; Feedback; Fibronectins; Foundations; G-Protein-Coupled Receptors; Glutamates; Homeostasis; Homo; Hormones; Hydrogen; Hyperactivity; In Vitro; Individual; Knock-out; Leucine-Rich Repeat; Mass Spectrum Analysis; Measures; Membrane; Metabotropic Glutamate Receptors; Modeling; Molecular; Motor; Movement Disorders; Mus; Mutation; Neurons; Neurotransmitters; Parkinson Disease; Pathology; Physiological; Physiology; Positioning Attribute; Post-Traumatic Stress Disorders; Proteins; Receptor Signaling; Regulation; Resolution; Role; Seizures; Signal Transduction; Stimulus; Structural Models; Structure; Synapses; Testing; base; crosslink; dimer; experimental study; extracellular; grasp; insight; postsynaptic; presynaptic; receptor; receptor binding; reconstitution; therapy design

Project Summary / Abstract It has recently been discovered that G-protein coupled receptors (GPCR) can be regulated by extracellular interactions. The mechanisms behind this phenomenon remain poorly understood. One such interaction is the trans-synaptic connection between presynaptic group III metabotropic glutamate receptors (mGluR) and postsynaptic cell-adhesion molecules of the extracellular leucine-rich repeat and fibronectin type III domain-containing (ELFN) family. Group III mGluRs sense glutamate release and inhibit further release in a negative feedback mechanism. A great body of evidence points to the essential role of synaptic glutamate homeostasis in a range of cognitive and motor functions with dysregulation leading to movement disorders and epilepsy. Binding and stabilization of mGluRs by trans-synaptic interactions with ELFN proteins is critical for regulating glutamate homeostasis. Loss of ELFN proteins in mice tremendously augments glutamate release and results in seizures and hyperactivity. Understanding how these proteins interact in the synaptic environment will provide valuable insight into how neurons maintain control over glutamate levels in the synapse and has implications for physiology and disease. Importantly, both mGluRs and ELFN proteins have been shown to form homo- and hetero-dimers. This proposal aims to test the hypothesis that apart from positioning and stabilizing group III mGluRs at the synapses, ELFN proteins act to allosterically modulate mGluR activity in part by influencing the dimerization dynamics of mGluR subunits. To test this hypothesis, I will build a structural model of the ELFN-mGluR interaction using crosslinking coupled with mass spectrometry, hydrogen/deuterium exchange, and cryo-electron microscopy to understand what binding determinants govern the ELFN-mGluR binding interaction. Additionally, I will investigate the mechanisms of mGluR allosteric modulation by ELFN proteins using a variety of cell-based signaling assays. Together, these experiments will provide a clear foundation for understanding how the brain maintains glutamate homeostasis inside the synapse.

项目摘要/摘要 最近发现，G蛋白偶联受体(GPCR)可以通过 细胞外相互作用。这一现象背后的机制仍然知之甚少。这样的一个 相互作用是突触前III型代谢型谷氨酸受体之间的跨突触联系 (MGluR)和细胞外富亮氨酸重复序列和纤维连接蛋白类型的突触后细胞黏附分子 III含结构域(ELFN)家族。第三组mGluRs感知谷氨酸释放并抑制进一步释放 负反馈机制。大量证据表明突触谷氨酸的重要作用 一系列认知和运动功能的动态平衡，调节失调导致运动障碍和 癫痫。通过跨突触相互作用与ELFN蛋白结合和稳定mGluRs对 调节谷氨酸动态平衡。小鼠ELFN蛋白的丢失可显著增加谷氨酸的释放 并导致癫痫发作和多动症。了解这些蛋白质如何在突触环境中相互作用 将为神经元如何维持对突触中谷氨酸水平的控制提供有价值的见解 对生理学和疾病的影响。重要的是，mGluRs和ELFN蛋白都已被证明形成 同二聚体和异二聚体。这一提议旨在检验这样一种假设，即除了定位和稳定 第三类mGluR在突触，ELFN蛋白作用于变构调节mGluR活性，部分是通过 影响mGluR亚基的二聚化动力学。为了验证这一假设，我将构建一个结构模型 用交联-质谱联用法研究ELFN-mGluR相互作用 交换和冷冻电子显微镜，以了解什么结合决定因素控制ELFN-mGluR 结合相互作用。此外，我还将研究ELFN对mGluR变构的调节机制 使用各种基于细胞的信号分析的蛋白质。总而言之，这些实验将提供一个明确的 为了解大脑如何在突触内维持谷氨酸平衡奠定了基础。