Integrin-Arg-SHP2 signaling regulates NMDAR function and neuron morphology

整合素-Arg-SHP2信号调节NMDAR功能和神经元形态

• 批准号: 9128713
• 负责人: Aaron Donald Levy
• 金额: $ 2.83万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-01 至 2017-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9128713
• 关键词: Address; Adolescence; Affect; Alleles; Alzheimer' s Disease; Attenuated; Autistic Disorder; Binding; Biochemical; Biotinylation; Cognitive deficits; Confocal Microscopy; Data; Defect; Dendrites; Dendritic Spines; Diagnosis; Disease; Economic Burden; Electron Microscopy; Electrophysiology (science); Functional disorder; Genetic; Head; Health; Hippocampus (Brain); Integrin alpha3beta1; Integrins; Knock-in; Link; Maintenance; Measures; Mediating; Mental Retardation; Mental disorders; Molecular; Mood Disorders; Morphology; Mus; N-Methyl-D-Aspartate Receptors; N-Methylaspartate; NMDA receptor antagonist; Neurodegenerative Disorders; Neurons; Noonan Syndrome; PTPN11 gene; Pathology; Phosphorylation; Phosphorylation Site; Play; Prosencephalon; Protein Tyrosine Kinase; Regulation; Regulatory Pathway; Role; Schizophrenia; Signal Transduction; Site; Slice; Structure; Surface; Synapses; Techniques; Testing; Therapeutic Intervention; Tyrosine Phosphorylation Site; Vertebral column; addiction; adhesion receptor; autism spectrum disorder; density; developmental disease; excitatory neuron; gain of function; insight; mutant; nervous system disorder; neuron loss; receptor function; research study; social; treatment strategy

DESCRIPTION (provided by applicant): Integrin-Arg-SHP2 signaling regulates NMDAR function and neuron morphology. Dendritic spine stability is disrupted in psychiatric and neurological disorders. Regulation of NMDA receptor (NMDAR) activity by integrin adhesion receptors plays a fundamental role in dendritic spine maturation and stability, but the molecular mechanisms by which integrins regulate NMDARs are unknown. We have shown that loss of integrin α3ß1 signaling through the Abl2/Arg non-receptor tyrosine kinase causes widespread dendritic spine loss in late adolescence due to increased GluN2B subunit-mediated NMDA receptor currents. I provide strong evidence that Arg acts through the tyrosine phosphatase SHP2 to control GluN2B phosphorylation and function. NMDA receptor dysfunction is a hallmark of psychiatric, neurodevelopmental and neurological diseases such as schizophrenia, autism, and Alzheimer's disease. Understanding the mechanism by which integrins regulate NMDAR function is critical to understand how synaptic stability is compromised in these disorders and to develop treatment strategies. In this proposal, I will test the hypothesis that integrin-Arg-SHP2 signaling functionally regulates the NMDA receptor to control dendritic spine stability. My first aim is to determine how SHP2 regulates GluN2B phosphorylation and function. In this aim, I will use substrate-trapping mutants of SHP2 to test whether GluN2B is directly dephosphorylated by SHP2 and to determine which site in GluN2B is targeted by SHP2. GluN2B phosphorylation promotes its surface localization. Therefore, I will also use surface biotinylation of NMDARs in cultured hippocampal neurons to measure how genetic and pharmacological loss- or gain-of-function of Arg and SHP2 influence NMDAR surface expression. My second aim is to elucidate an integrin-NMDA receptor regulatory mechanism. GluN2B phosphorylation increases NMDAR-mediated currents, and my preliminary data show that activation of SHP2 decreases GluN2B phosphorylation. To test how SHP2 regulates NMDAR function, I will measure how loss- or gain-of-function of SHP2 activity affects NMDAR-mediated currents in hippocampal slices. I will also test the hypothesis that SHP2 functions as a mechanistic link between integrin-Arg signaling and the NMDAR by assessing whether an SHP2 gain-of-function allele can suppress elevated NMDAR currents in mice lacking integrin α3ß1 or Arg. My third aim is to understand how SHP2 regulates dendritic spine and synapse structure. Our lab has shown that integrin-Arg signaling attenuates NMDAR activity to stabilize spines, and I hypothesize that SHP2 mediates the effects of Arg signaling on NMDAR function. I will use confocal and electron microscopy to quantitatively measure how loss- and gain-of-function of SHP2 activity in mice affects dendritic spine and synapse structure and stability. I will also test whether an activated SHP2 allele can suppress the spine destabilization resulting from loss of integrin α3ß1 or Arg in mice.

描述（由适用提供）：整合素-ARG-SHP2信号传导调节NMDAR功能和神经元形态。树突状的脊柱稳定性在精神病和神经系统疾病中被禁用。整合素胶粘剂受体对NMDA受体（NMDAR）活性的调节在树突状脊柱成熟和稳定性中起着基本作用，但是整合素调节NMDAR的分子机制尚不清楚。我们已经表明，通过ABL2/ARG非受体酪氨酸激酶通过ABL2/ARG非受体酪氨酸激酶的丧失导致青春期晚期的树突状脊柱损失，因为GLUN2B亚基介导的NMDA受体电流增加。我提供了有力的证据表明，ARG通过酪氨酸磷酸酶SHP2起作用以控制Glun2b磷酸化和功能。 NMDA受体功能障碍是精神病，神经发育和神经系统疾病的标志，例如精神分裂症，自闭症和阿尔茨海默氏病。了解整联蛋白调节NMDAR功能的机制对于了解突触稳定性如何在这些疾病中损害和发展治疗策略至关重要。在此提案中，我将检验以下假设：整联蛋白ARG-SHP2信号在功能上调节NMDA受体以控制树突状脊柱稳定性。我的第一个目的是确定SHP2如何调节GLUN2B磷酸化和功能。在此目标中，我将使用SHP2的底物捕获突变体来测试Glun2b是否直接被SHP2脱磷酸化，并确定GLUN2B中的哪个位点由SHP2靶向。 Glun2b磷酸化促进其表面定位。因此，我还将在培养的海马神经元中使用NMDAR的表面生物素化来测量ARG和SHP2的遗传和药物损失或功能的影响如何影响NMDAR表面表达。我的第二个目的是阐明整合素-NMDA受体调节机制。 Glun2b光术增加了NMDAR介导的电流，而我的初步数据表明，SHP2的激活降低了Glun2b光术。为了测试SHP2如何调节NMDAR函数，我将衡量SHP2活性的损失或功能的损失或功能如何影响海马切片中NMDAR介导的电流。我还将通过评估缺乏整合素α3ß1或ARG的小鼠中升高的NMDAR电流来测试SHP2作为整合素ARG信号传导与NMDAR之间的机械联系的假设。我的第三个目标是了解SHP2如何调节树突状脊柱和突触结构。我们的实验室表明，整联蛋白ARG信号传导减弱了NMDAR活性以稳定刺，我假设SHP2介导了ARG信号对NMDAR功能的影响。我将使用共聚焦和电子显微镜来定量测量小鼠SHP2活性的损失和功能收益如何影响树突状脊柱，突触结构和稳定性。我还将测试激活的SHP2等位基因是否可以抑制因整联蛋白α3ß1的损失还是小鼠中的ARG导致的脊柱不稳定。