Validation of FGF14 as a New Molecular Target of GSK3

描述（由申请人提供）：精神疾病是一种慢性的、毁灭性的疾病，被认为是由不适应的大脑可塑性引起的，迫切需要有效和安全的药物治疗。确定可能维持这些异常神经适应的机制联系将促进我们对精神障碍生物学的理解，可能为药物开发提供新的平台。利用一种创新的基于生物发光的分子筛选方法，结合生化、电生理和成像分析，我们提供了突破性的结果，显示糖原合成酶激酶3 （GSK3）与神经元兴奋性之间的联系，GSK3是一种在情绪障碍、抑郁症和精神分裂症中发现功能失调的关键酶。我们认为这是与精神疾病和某些成瘾行为相关的神经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通道复合物的形成由GSK3和GSK3组成抑制因子蛋白激酶B （Akt）双向控制，并且GSK3直接磷酸化FGF14。Akt和GSK的药物抑制分别增加和阻止FGF14:Nav通道复合物的形成，而GSK3的抑制则阻断了Akt抑制的作用。在海马神经元中，抑制GSK3使FGF14:Nav通道复合物从轴突初始段（AIS）（动作电位起始部位）分散，损害固有神经元并减少兴奋性突触传递，而抑制Akt则导致相反的表型。此外，我们发现Fpep1，一种模拟FGF14:Nav通道界面的小干扰肽，可以阻止FGF14:Nav通道复合物的组装，这为最小化GSK3对神经元兴奋性的影响提供了一种工具。在这个提案中，我们将采用基于生物发光的技术，质谱分析，磷酸化分析，通过共聚焦成像和电生理学来确定GSK3控制FGF14:Nav通道复合物形成的分子机制（Aim 1）和促进神经元中FGF14:Nav通道复合物的靶向性（Aim 2），并评估GSK3是否通过FGF14:Nav通道复合物对皮质边缘回路的兴奋性和神经可塑性产生影响，这种影响可以通过靶向FGF14的药理学或遗传学方法逆转（Aim 3）。本研究的积极结果将为GSK3在大脑中的分子机制提供新的见解，并为开发针对GSK3相关精神疾病的新药物提供前所未有的机会。

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.