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 最近已经发现，G蛋白偶联受体（GPCR）可以通过 细胞外相互作用。这种现象背后的机制仍然很少理解。一个这样的 相互作用是突触前III组代替型谷氨酸受体之间的反射突触连接 （MGLUR）和突触后细胞粘附分子的细胞外亮氨酸重复和纤连蛋白类型 III含领域（ELFN）家族。 III组MGLURS有谷氨酸释放并抑制在A中进一步释放 负反馈机制。大量证据表明突触谷氨酸的基本作用 在一系列认知和运动功能中的体内平衡，导致运动障碍和 癫痫。通过反式突触相互作用与ELFN蛋白的结合和稳定MGLURS对 调节谷氨酸稳态。小鼠中ELFN蛋白的损失极大地增强了谷氨酸释放 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 将为神经元如何保持对突触中的谷氨酸水平的控制，并具有宝贵的见解 对生理和疾病的影响。重要的是，已显示MGLURS和ELFN蛋白质形成 同性恋和异二聚体。该建议旨在检验以下假设：除了定位和稳定 在突触下的III组mglurs，ELFN蛋白作用于变构调节Mglur活性的一部分是由 影响mglur亚基的二聚化动力学。为了检验这一假设，我将建立一个结构模型 使用质谱，氢/氘的交联相互作用的Elfn-mglur相互作用 交换和冷冻电子显微镜，以了解哪种结合决定因素控制ELFN-MGLUR 结合相互作用。此外，我将研究ELFN MGLUR变构调制的机制 使用各种基于细胞的信号测定的蛋白质。这些实验将共同提供清晰的 理解大脑如何维持突触内谷氨酸稳态的基础。