Control of AMPA receptor function by phosphorylation

通过磷酸化控制 AMPA 受体功能

• 批准号: 8213435
• 负责人: Stephen F Traynelis
• 金额: $ 33.23万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-02-01 至 2014-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8213435
• 关键词: AMPA Receptors; Accounting; Address; Agonist; Binding; Biochemical; Brain; C-terminal; Calmodulin; Cations; Cell model; Chemosensitization; Cognition; Communication; Coupling; Data; Elements; Event; Excitatory Synapse; Glutamate Receptor; Glutamates; Goals; Hippocampus (Brain); Hydrogen Bonding; Individual; Infection; Ion Channel; Lead; Learning; Ligands; Long-Term Potentiation; Mediating; Membrane; Memory; Modeling; Movement; Mutagenesis; Neuraxis; Neurons; Peptides; Phosphorylation; Phosphorylation Site; Phosphotransferases; Play; Probability; Property; Protein Kinase; Protein Kinase C; Proteins; Recombinants; Regulation; Role; Scanning; Series; Serine; Signal Pathway; Signal Transduction; Structural Models; Synapses; Synaptic plasticity; System; Testing; Time; Work; base; functional mimics; ionic bond; neurotransmission; novel; polypeptide; postsynaptic; public health relevance; receptor; receptor function; research study; response; stargazin; trafficking

DESCRIPTION (provided by applicant): The AMPA-type glutamate receptors are ligand-gated cation channels that mediate fast excitatory neurotransmission in the brain, and thus are critically involved in all aspects of brain function including cognition, movement, learning, and memory. The function and number of postsynaptic AMPA receptors are dynamically regulated to control the strength of synaptic connections, and this plasticity is a key feature of cellular models of learning and memory. Signals that trigger synaptic plasticity lead to phosphorylation of AMPA receptors by protein kinases, and this phosphorylation controls AMPA receptor trafficking and function. Phosphorylation by protein kinase C (PKC) or Ca2+/calmodulin dependent kinase II (CamKII) of an intracellular serine residue (Ser831) located on the GluR1 subunit enhances AMPA receptor function to increase synaptic strength during expression of long-term potentiation (LTP), one model of synaptic plasticity. Although previous studies observed that CamKII phosphorylation of GluR1 enhances the single channel conductance, no conceptual or structural mechanism has been described for this unique form of ion channel regulation. The goal of the experiments proposed here is to understand functionally, structurally, and conceptually how phosphorylation of GluR1 Ser831 potentiates AMPA receptor function. We will focus on Ser831 in GluR1 because of the unique mechanism of potentiation (increased unitary conductance), and will expand the study to evaluate for the first time how three nearby phosphorylation sites (Ser818, Thr840, Ser845) might functionally interact with phospho-Ser831. Furthermore, we will test whether the effects of phospho-Ser831 reflect either intra-protein interactions between the phospho-Ser831 and intracellular portions of the receptor, or inter-protein interactions between phospho-Ser831 and GluR1 binding partners. Completion of these studies will provide a comprehensive functional and structural understanding of an under-studied feature of synaptic plasticity-phosphorylation mediated changes in postsynaptic AMPA channel function. The proposed experiments address three questions: 1. What is the mechanism by which phosphorylation regulates AMPA receptor function? Single channel currents will be recorded to determine how phosphorylation of Ser831 controls GluR1 function. We will also evaluate the interactions of Ser831 with nearby phosphorylation sites, and validate our conclusions in neurons. 2. What is the structural basis for phospho-serine regulation of AMPA receptor function? We will identify intracellular GluR1 residues as phospho-Ser831 hydrogen bonding partners. We will additionally search for inter-protein interactions involving GluR1 that depend on the phosphorylation of Ser831. 3. Can models of independent subunit gating describe AMPA receptor regulation by phosphorylation? We will analyze the response of patches with one active GluR1 channel (plus stargazin) to the rapid application of a maximally effective concentration of glutamate. These data will be used to develop a novel model of subunit gating that can account for the potentiation of GluR1 channel function by phosphorylation of Ser831. PUBLIC HEALTH RELEVANCE: AMPA receptors mediate communication between neurons in the central nervous system, and thus play an important role in virtually all brain functions. The AMPA receptors are comprised of four different subunits (GluR1-4). Among these, the GluR1 subunit has been shown to play a unique role in activity-dependent synaptic plasticity. GluR1 is subject to C-terminal phosphorlyation by a variety of kinases, and this phosphorylation can influence trafficking to the membrane and AMPA receptor function. In this proposal we examine CamKII phosphorylation of GluR1-Ser831. CamKII activation has been shown to be a critical step in some forms of synaptic plasticity, presumably through phosphorylation of the GluR1 subunit. Phosphorylation of GluR1-Ser831 increases single channel conductance by an unknown mechanism. This proposal describes three series of experiments that will evaluate the underlying functional and structural mechanisms of the effects on receptor function following phosphorylation at GluR1-Ser831. Understanding how AMPA receptor function is sculpted by intracellular signaling pathways is an important step towards understanding the mechanisms of synaptic plasticity, which likely underlie higher order functions such as learning and memory.

描述（由申请人提供）：AMPA型谷氨酸受体是配体门控阳离子通道，可介导大脑中快速兴奋的神经传递，因此与大脑功能的各个方面有关键有关，包括认知，运动，学习和记忆。在动态调节突触后AMPA受体的功能和数量以控制突触连接的强度，并且这种可塑性是蜂窝学习和记忆模型的关键特征。触发突触可塑性导致蛋白激酶对AMPA受体的磷酸化的信号，而这种磷酸化控制了AMPA受体的运输和功能。蛋白激酶C（PKC）或Ca2+/钙调蛋白依赖性激酶II（CAMKII）的磷酸化在GLUR1亚基上的细胞内丝氨酸残基（SER831）的磷酸化增强了AMPA受体功能，从而增强了长期强度表达（LTP）表达的突触强度（一种模型），一种模型，一种模型。尽管先前的研究观察到GLUR1的CAMKII磷酸化增强了单个通道电导，但对于这种独特的离子通道调节形式，尚未描述任何概念或结构机制。此处提出的实验的目的是从功能，结构和概念上理解GLUR1 Ser831的磷酸化如何增强AMPA受体功能。由于具有独特的增强机制（增加单位电导），我们将重点关注SER831，并将扩大研究以首次评估附近的三个附近的磷酸化位点（SER818，THR840，SER845）如何与磷s-ser831相互作用。此外，我们将测试磷酸-SER831的作用是反映受体的磷酸-SER831与细胞内部分之间的蛋白质内相互作用，还是磷酸化 - ser831和Glur1结合伴侣之间的蛋白质间相互作用。这些研究的完成将为突触可塑性 - 磷酸化介导的突触后AMPA通道功能的变化提供全面的功能和结构理解。提出的实验解决了三个问题：1。磷酸化调节AMPA受体功能的机制是什么？将记录单个通道电流，以确定SER831的磷酸化如何控制GLUR1功能。我们还将评估SER831与附近磷酸化位点的相互作用，并验证我们在神经元中的结论。 2。AMPA受体功能的磷酸链调节的结构基础是什么？我们将确定细胞内GLUR1残基为磷酸 - SER831氢键伙伴。我们还将搜索涉及Ser831磷酸化的GLUR1的蛋白间相互作用。 3。独立亚基门控模型可以通过磷酸化描述AMPA受体调节吗？我们将用一个活性GLUR1通道（加星状蛋白）对斑块的响应进行分析，以快速应用最大有效浓度的谷氨酸。这些数据将用于开发一种新型的亚基门控模型，该模型可以通过Ser831的磷酸化来解释GLUR1通道功能的增强。公共卫生相关性：AMPA受体介导了中枢神经系统中神经元之间的沟通，因此在几乎所有大脑功能中都起着重要作用。 AMPA受体由四个不同的亚基组成（GLUR1-4）。其中，GLUR1亚基已显示在活动依赖性突触可塑性中起着独特的作用。 GLUR1受多种激酶受到C末端磷光化的影响，这种磷酸化可以影响对膜和AMPA受体功能的运输。在此提案中，我们检查了Glur1-Ser831的CaMKII磷酸化。 CAMKII激活已被证明是某些形式的突触可塑性的关键步骤，大概是通过GLUR1亚基的磷酸化。 GLUR1-SER831的磷酸化通过未知机制增加了单个通道电导。该建议描述了三个系列实验，这些实验将评估Glur1-Ser831磷酸化后对受体功能的影响的潜在功能和结构机制。了解如何通过细胞内信号通路雕刻AMPA受体功能是了解突触可塑性机制的重要一步，这可能是高级功能（例如学习和记忆）的基础。