NSF/MCB-BSF: Collaborative Research: Towards development of the structural determinants of the Glutamate receptor gating regulation by auxiliary membrane anchored proteins

高级认知功能，包括定量推理、学习和记忆，起源于人脑，是通过突触连接的复杂神经元网络之间通信的结果。这种突触连接的关键分子成分是离子型谷氨酸受体(IGluR)，这是突触后神经细胞膜上的一种蛋白质，它传导电信号来传递来自突触前神经元的信息。这一神经传递过程受到多种与谷氨酸受体结合的辅助蛋白的调节。这个项目将阐明不同的辅助蛋白如何与谷氨酸受体以及彼此相互作用，以启动和调节神经元之间的通信。这项研究增强了我们对大脑功能的分子基础的了解。这项研究将涉及本科生和研究生，在脑研究的前沿提供教育环境和培训机会。还计划了涉及当地高中和大学的外联活动，以促进对STEM领域的多样化参与。人们越来越认识到，大多数蛋白质在动态形成和重新形成的复合体内的细胞中运作。这对信号受体蛋白尤其重要，例如大脑中的离子型受体，它通过突触后膜传导电流，促进神经元之间的快速通信。AMPA受体(AMPAR)--iGluRs的一个亚型--介导哺乳动物大脑中最快的兴奋性神经传递。相应地，AMPAR运输、定位和功能的调节是决定突触强度和可塑性的关键机制，而突触强度和可塑性是高认知大脑功能(如学习和记忆)的基础。最近发现，突触AMPAR是一种与多种辅助蛋白的复合体。本项目旨在研究AMPAR与胞结AMPAR调节蛋白(CKAMPs)的分子相互作用以及它们与跨膜AMPAR调节蛋白(TARP)的相互作用。大量的AMPAR和CKAMP突变体将被用来确定参与受体调节的相互作用结构域和特定残基。将使用包括对接和分子动力学模拟在内的计算结构建模来指导进一步确定蛋白质-蛋白质界面，这将通过电生理学实验进行测试，并生成和分析用于冷冻-EM成功结构确定的候选构建体。这一合作努力结合了互补的方法和持续的信息和试剂交换，可能会提供所需的协同作用，以达到对AMPAR监管的新水平的理解。除了提供对AMPAR门控和调控的基本见解外，这项研究的结果也将成为研究AMPAR在突触生理学中介导的过程的路线图。该奖项由分子和细胞生物科学部、整合组织系统司和生命风险基金规则共同资助。这一美国/以色列合作项目得到了美国国家科学基金会和以色列双国科学基金会的支持。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

High cognitive functions, including quantitative reasoning, learning and memory, originate in the human brain as a result of communication between complex networks of neurons connected through synapses. The key molecular element of this synaptic connection is the ionotropic glutamate receptor (iGluR), a protein in the postsynaptic neuronal membrane that conducts electrical signals to deliver messages from the presynaptic neuron. This process of neurotransmission is modulated by a variety of auxiliary proteins that bind to glutamate receptors. This project will illuminate how different auxiliary proteins interact with the glutamate receptor and with each other to initiate and modulate communication between neurons. This study enhances our knowledge of the molecular underpinnings of brain function. The research will involve undergraduate and graduate students, providing an educational environment and training opportunities at the forefront of brain research. Outreach activities involving local high schools and colleges are planned as well to promote diverse participation in STEM fields.It is increasingly recognized that the majority of proteins operate in cells within dynamically formed and re-formed complexes. This is especially important for signaling receptor proteins, such as ionotropic receptors in the brain that conduct electrical current through postsynaptic membranes to facilitate fast communication between neurons. The AMPA receptors (AMPAR) - a subtype of iGluRs - mediate the fastest excitatory neurotransmission in the mammalian brain. Correspondingly, regulation of AMPAR trafficking, localization and function is the key mechanism determining synaptic strength and plasticity that underlies high cognitive brain functions, such as learning and memory. It has been discovered recently that synaptic AMPARs function as complexes with a multitude of auxiliary proteins. This project aims to characterize molecular interactions of AMPARs with the cys-knot AMPAR modulating proteins (CKAMPs) and their interplay with the transmembrane AMPAR regulatory proteins (TARPs). A large collection of AMPAR and CKAMP mutants will be employed to identify interaction domains and specific residues involved in receptor regulation. Computational structural modeling including docking and molecular dynamics simulations will be employed to guide further determination of the protein-protein interfaces, which will be tested by electrophysiology experiments, and to generate and analyze candidate constructs for successful structure determination by cryo-EM. This collaborative effort, combining complementary approaches and continuous exchange of information and reagents, may provide the synergy needed to reach a new level of understanding of AMPAR regulation. In addition to providing fundamental insights into AMPAR gating and regulation, the results of this study will also serve as a roadmap for investigating AMPAR-mediated processes in synaptic physiology.This award was co-funded by the Division of Molecular and Cellular Biosciences, the Division of Integrative Organismal Systems, and the Rules of Life Venture Fund.This collaborative US/Israel project is supported by the US National Science Foundation and the Israeli Binational Science Foundation.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.