Validation of FGF14 as a New Molecular Target of GSK3

验证 FGF14 作为 GSK3 的新分子靶点

• 批准号: 8373279
• 负责人: Fernanda Laezza
• 金额: $ 38.25万
• 依托单位: UNIVERSITY OF TEXAS MED BR GALVESTON
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-08-01 至 2017-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8373279
• 关键词: Action Potentials; Addictive Behavior; Binding; Biochemical; Biochemistry; Biological Assay; Biology; Bioluminescence; Brain; Cell Culture System; Cells; Chronic; Co-Immunoprecipitations; Complex; Confocal Microscopy; Data; Development; Disease; Drug Delivery Systems; Electrophysiology (science); Enzymes; Functional disorder; Genetic; Glycogen Synthase Kinase 3; Hippocampus (Brain); Image; Intervention; Life; Link; Maintenance; Mass Spectrum Analysis; Mental Depression; Mental disorders; Modeling; Molecular; Molecular Biology; Molecular Target; Mood Disorders; Neuronal Dysfunction; Neuronal Plasticity; Neurons; Outcome Study; Pathway interactions; Peptides; Pharmaceutical Preparations; Pharmacotherapy; Phenotype; Phosphorylation; Proto-Oncogene Proteins c-akt; Repressor Proteins; Rodent Model; Role; Schizophrenia; Screening procedure; Site; Site-Directed Mutagenesis; Slice; Symptoms; Synapses; Synaptic Transmission; Technology; Testing; Therapeutic Intervention; Time; Validation; base; cellular imaging; cortico-limbic circuits; fibroblast growth factor-14; in vivo; innovation; insight; multidisciplinary; mutant; neuroadaptation; neuronal circuitry; neuronal excitability; novel; patch clamp; prevent; success; tool

DESCRIPTION (provided by applicant): Psychiatric diseases are chronic, devastating disorders thought to arise from maladaptive brain plasticity, and potent and safe pharmacotherapies are in great need. Identifying the mechanistic links that might sustain these aberrant neuroadaptations will advance our understanding of the biology of mental disorders, potentially providing new platforms for medication development. Using an innovative bioluminescence-based molecular screening approach combined with biochemical, electrophysiological, and imaging assays, we provide breakthrough results showing a link between glycogen synthase kinase 3 (GSK3), a critical enzyme found dysfunctional in mood disorders, depression and schizophrenia, and neuronal excitability, which we propose as a potential mechanism underlying dysfunction of neuronal circuitries associated with psychiatric disorders and certain addictive behaviors. Building on previous discoveries demonstrating that fibroblast growth factor 14 (FGF14) is a functionally relevant component of the Nav channelosome that controls neuronal excitability, we present exciting new data showing that the FGF14:Nav channel complex formation is bi-directionally controlled by GSK3 and by the GSK3 constitutive repressor, protein kinase B (Akt), and that GSK3 directly phosphorylates FGF14. Pharmacological inhibition of Akt and GSK increases and prevents, respectively, the FGF14:Nav channel complex formation, whereas inhibition of GSK3 occludes the effect of Akt inhibition. In hippocampal neurons, GSK3 inhibition disperses the FGF14:Nav channel complex from the axonal initial segment (AIS), the site of action potential initiation, impairs intrinsic fring and reduces excitatory synaptic transmission, whereas inhibition of Akt leads to opposite phenotypes. Furthermore, we show that Fpep1, a small interfering peptide modeled upon the FGF14:Nav channel interface, prevents the FGF14:Nav channel complex assembly, providing a tool for minimizing the effect of GSK3 on neuronal excitability in vivo. In this proposal we will employ a combination of bioluminescence-based technology, mass spectrometry, phosphorylation assays, confocal imaging and electrophysiology to determine the molecular mechanism by which GSK3 controls the FGF14:Nav channel complex formation (Aim 1) and promotes targeting of the FGF14:Nav channel complex in neurons (Aim 2) and to evaluate whether GSK3 exerts an effect on excitability and neuroplasticity in cortico-limbic circuits through the FGF14:Nav channel complex that could be reversed by pharmacological or genetic approaches targeting FGF14 (Aim 3). Positive outcomes of this study will provide new insights into the molecular mechanisms of GSK3 in the brain and offer an unprecedented opportunity for new medication development against GSK3-linked psychiatric disorders. PUBLIC HEALTH RELEVANCE: Innovative and integrated approaches are needed to enhance the success of therapeutic interventions against psychiatric disorders. Through a multidisciplinary project including molecular biology, biochemistry mass spectrometry, single cell imaging and electrophysiology in rodent models, we will validate FGF14 as a novel downstream target of GSK3, creating a novel platform for intervention against psychiatric disorders associated with GSK3 dysfunction.

描述（由申请人提供）：精神疾病是慢性的、破坏性的疾病，被认为是由适应不良的大脑可塑性引起的，并且非常需要有效且安全的药物疗法。识别可能维持这些异常神经适应的机制联系将增进我们对精神障碍生物学的理解，并有可能为药物开发提供新平台。使用基于生物发光的创新分子筛选方法，结合生化、电生理学和成像检测，我们提供了突破性的结果，显示了糖原合酶激酶 3 (GSK3) 与神经元之间的联系，糖原合酶激酶 3 是一种在情绪障碍、抑郁症和精神分裂症中功能失调的关键酶。兴奋性，我们认为这是与精神疾病和某些成瘾行为相关的神经元回路功能障碍的潜在机制。基于先前的发现，证明成纤维细胞生长因子 14 (FGF14) 是控制神经元兴奋性的 Nav 通道体的功能相关成分，我们提出了令人兴奋的新数据，表明 FGF14:Nav 通道复合物的形成受到 GSK3 和GSK3 组成型阻遏蛋白蛋白激酶 B (Akt)，并且 GSK3 直接磷酸化 FGF14。 Akt 和 GSK 的药理学抑制分别增加和阻止 FGF14:Nav 通道复合物的形成，而 GSK3 的抑制则阻断 Akt 抑制的作用。在海马神经元中，GSK3 抑制将 FGF14:Nav 通道复合物从轴突初始段 (AIS)（动作电位起始位点）分散，损害内在环并减少兴奋性突触传递，而 Akt 抑制会导致相反的表型。此外，我们还发现 Fpep1（一种以 FGF14:Nav 通道界面为模型的小干扰肽）可阻止 FGF14:Nav 通道复合物的组装，从而为最大限度地减少 GSK3 对体内神经元兴奋性的影响提供了工具。在本提案中，我们将结合使用基于生物发光的技术、质谱、磷酸化测定、共焦成像和电生理学来确定 GSK3 控制 FGF14:Nav 通道复合物形成（目标 1）并促进 FGF14 靶向的分子机制：神经元中的导航通道复合体（目标 2）并评估 GSK3 是否通过FGF14：导航通道复合体，可以通过针对 FGF14 的药理学或遗传方法来逆转（目标 3）。这项研究的积极成果将为 GSK3 在大脑中的分子机制提供新的见解，并为针对 GSK3 相关精神疾病的新药物开发提供前所未有的机会。 公共卫生相关性：需要创新和综合的方法来提高精神疾病治疗干预的成功率。通过啮齿动物模型中的分子生物学、生物化学质谱、单细胞成像和电生理学等多学科项目，我们将验证 FGF14 作为 GSK3 的新下游靶点，创建一个新的平台来干预与 GSK3 功能障碍相关的精神疾病。