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Structural studies of NaV1.5 and functional implications.

NaV1.5 的结构研究和功能意义。

基本信息

批准号：
9443872
负责人：
Geoffrey S Pitt
金额：
$ 38.7万
依托单位：
WEILL MEDICAL COLL OF CORNELL UNIV
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-07-01 至 2018-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9443872
关键词：
Structural studies NaV1.5 functional implications.

项目摘要

DESCRIPTION (provided by applicant): Voltage-gated Na channels (NaV) initiate electrical impulses in neurons and the heart. Mutations in SCN1A and SCN2A, which encode the neuronal channels NaV1.1 and NaV1.2, respectively, cause inherited epilepsy syndromes. Mutations in SCN5A, which encodes the major subtype of NaV channel in heart (NaV1.5), are linked to many cardiac arrhythmogenic disorders, such as Long QT syndrome (LQTS) and Brugada syndrome (BrS). The main structural components of NaV channels are four homologous repeats, which assemble in a tetrameric configuration within the membrane to control voltage-dependent Na+ conduction, and a cytosolic C-terminal domain (CTD) that serves as a binding site for many auxiliary regulatory proteins and is a hotspot for disease-causing mutations. Among these auxiliary CTD-interacting proteins, the ubiquitous Ca2+ sensor calmodulin (CaM) and fibroblast growth factor homologous factors (FHFs; FGF11-FGF14) are of particular interest: many disease- causing mutations are localized to their putative interaction domains within the CTDs of specific NaV channels; FGF14 is itself a locus for the neurodegenerative disorder spinocerebellar ataxia 27; and Ca2+ regulation of NaV channels by CaM is poorly understood. A lack of structural information for these interactions with the CTD, however, has prevented an understanding of the roles of the CTDs, these auxiliary proteins in channel regulation, and the molecular basis of mutational effects on NaV that lead to disease. Building on recent success in determining the crystal structure of a NaV CTD in complex with CaM and an FHF; and in defining new roles for FHFs in regulation of NaV channel gating and trafficking, we propose to define the mechanisms of NaV channel regulation by Ca2+/CaM, and FHFs and to uncover the molecular basis of the mutational effects on NaV CTDs and their associated proteins that lead to channelopathies. To achieve these goals, we propose the following specific aims: 1. Structure-function studies to determine how Ca2+/CaM regulate NaV channel function, using electrophysiology and biochemistry, to probe insights gained from structural determination; 2. Structure-function studies to determine how FHFs regulate NaV channel function and how FHFs co-operate with CaM for channel regulation; 3. Physiological investigations of the consequences of FHF regulation of NaV1.5. Because NaV channelopathies and mutations in FHFs are associated with neurodegenerative diseases, epilepsy syndromes, and cardiac arrhythmias, the proposed project has great potential to illuminate the molecular basis of multiple channelopathies in multiple organ systems due to NaV CTD mutations and, more generally, to uncover fundamental aspects of NaV channel structure-function.

描述（由申请人提供）：电压门控Na通道（NaV）在神经元和心脏中启动电脉冲。分别编码神经元通道NaV1.1和NaV1.2的SCN 1A和SCN 2A的突变导致遗传性癫痫综合征。SCN 5A编码心脏中NaV通道的主要亚型（NaV1.5），其突变与许多心脏致心律失常性疾病有关，如长QT综合征（LQTS）和Brugada综合征（BrS）。NaV通道的主要结构组分是四个同源重复序列，其在膜内组装成四聚体构型以控制电压依赖性Na+传导，以及胞质C末端结构域（CTD），其充当许多辅助调节蛋白的结合位点，并且是致病突变的热点。在这些辅助CTD相互作用蛋白中，普遍存在的Ca 2+传感器钙调素（CaM）和成纤维细胞生长因子同源因子FHF; FGF 11-FGF 14）是特别感兴趣的：许多致病突变定位于特定NaV通道的CTD内的它们推定的相互作用结构域; FGF 14本身是神经变性病症脊髓小脑共济失调27的基因座; Ca 2+通过CaM对NaV通道的调节还知之甚少。然而，缺乏与CTD的这些相互作用的结构信息，阻碍了对CTD、这些辅助蛋白在通道调节中的作用以及导致疾病的NaV突变效应的分子基础的理解。基于最近成功确定NaV CTD与CaM和FHF复合物的晶体结构，以及定义FHF在NaV通道门控和运输调节中的新作用，我们建议定义Ca 2 +/CaM和FHF调节NaV通道的机制，并揭示导致通道病的NaV CTD及其相关蛋白突变效应的分子基础。为实现这些目标，我们提出以下具体目标：1.结构-功能研究，以确定Ca 2 +/CaM如何调节NaV通道功能，使用电生理学和生物化学，以探索从结构测定中获得的见解; 2.结构-功能研究以确定FHF如何调节NaV通道功能以及FHF如何与CaM合作进行通道调节; 3. FHF调节NaV1.5的后果的生理学研究。由于NaV通道病和FHF突变与神经退行性疾病、癫痫综合征和心律失常相关，因此该项目具有很大的潜力，可以阐明NaV CTD突变导致的多器官系统中多通道病的分子基础，更普遍地说，可以揭示NaV通道结构-功能的基本方面。