VGSC Modulation by FHFs: Neural Functions and Mechanisms

描述(申请人提供)：电压门控钠通道(VGSC)在可兴奋细胞中产生和传播电活动。VGSC在闭合、开放和失活状态之间转换的精确动力学对于大脑、神经、心脏和肌肉功能是必不可少的，正如临床上广泛的钠通道病疾病所证明的那样。虽然分离的VGSCα亚基包括通道的孔洞、电压传感器和失活机制，但与成纤维细胞生长因子同源因子(FHFs)的相互作用对VGSC失活的动力学有巨大而复杂的影响。事实上，FHF突变是人类脊髓小脑性共济失调的原因之一，并导致神经元内在兴奋性的缺陷。虽然所有的FHF亚型都与VGSC结合，但它们调节VGSC快速失活和诱导新表征的长期失活通道状态的能力不同。本申请提出了扩展FHF神经元功能和FHF诱导的VGSC调制的物理机制的实验。目的I：我们的总体生物学假设是，FHF亚型在不同神经元及其亚细胞室的不同表达决定了细胞的兴奋和传导特性。我们将测试：(I-A)轴突起始段不同长形FHFs(A型FHFs，FHF4B)的相对丰度是否规定了神经元的兴奋特性？(I-B)A型FHs的躯体树突膜定位是否能限制神经元重复放电过程中钠动作电位的反向传播？(I-C)动作电位轴突传导不需要或可能有害的长型FHFs？目的II：更清楚地了解FHFs调节VGSC的物理机制将进一步深入了解VGSC的构象动力学，并为设计治疗癫痫和心律失常等高兴奋性疾病的药物提供合理的方法。我们将测试：(II-A)A型FHFs远端N端的长期失活颗粒是否停靠在VGSC的细胞质腔内？(II-B)对接的A型FHF灭活颗粒中的阳离子残基是否起到屏蔽钠离子传导的作用？(II-C)FHFs的三叶状核心附近的阳离子区域如何调节VGSC的快速失活？

DESCRIPTION (provided by applicant): Voltage-gated sodium channels (VGSCs) generate and propagate electrical activity in excitable cells. The precise dynamics of VGSC transitions among closed, open, and inactivated states are essential for brain, nerve, heart, and muscle function, as documented by a wide range of clinical sodium channelopathy disorders. While the VGSC alpha subunit in isolation comprises the channel's pore, its voltage sensors, and its inactivation machinery, interaction with fibroblast growth factor homologous factors (FHFs) has large and complex effects on the dynamics of VGSC inactivation. Indeed, FHF mutations are a cause of human spinocerebellar ataxia and cause defects in neuronal intrinsic excitability. While all FHF isoforms bind VGSCs, they differ in their abilities to modulate VGSC fast inactivation and to induce a newly characterized long-term inactivated channel state. This application proposes experiments to expand analysis of FHF neuronal functions and physical mechanisms of FHF- induced VGSC modulation. AIM I: Our overall biological hypothesis is that differential expression of FHF isoforms in different neurons and their subcellular compartments acts to determine excitation and conduction properties of cells. We will test: (I-A) Does the relative abundance of different long-form FHFs (A-type FHFs, FHF4B) at the axon initial segment specify the excitation properties of a neuron? (I-B) Does somatodendritic membrane localization of A-type FHFs act to limit sodium action potential backpropagation during repetitive firing of neurons? (I-C) Are long-form FHFs not required for, and potentially deleterious to, action potential axonal conduction? AIM II: A clearer understanding of physical mechanisms for VGSC modulation by FHFs will provide further insight into VGSC conformation dynamics and suggest rational approaches for designing therapeutics for managing disorders of hyperexcitability, such as epilepsies and arrhythmias. We will test: (II-A) Does the long- term inactivation particle at the distal N-terminus of A-type FHFs dock within the cytoplasmic cavern of a VGSC? (II-B) Do the cationic residues in the docked A-type FHF inactivation particle act as a shield against sodium ion conduction? (II-C) How does a cationic region near the ?-trefoil core of FHFs modulate VGSC fast inactivation?