INTRACELLULAR FGFS:NOVEL REGULATIONS OF CARDIAC NAV CHANNELS

细胞内 FGFS：心脏 NAV 通道的新颖调节

基本信息

批准号：
8031777
负责人：
JEANNE M. NERBONNE
金额：
$ 22.8万
依托单位：
WASHINGTON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-01-01 至 2012-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8031777
关键词：
Action Potentials Adult Affect Arrhythmia Atrial Fibrillation Attenuated Binding Biochemical C-terminal Cardiac Cell surface Cells Cytoplasmic Granules Disease Family Fibroblast Growth Factor Future Gene Targeting Generations Goals Heart Heart Diseases In Vitro Inherited Investigation Ion Channel Life Link Longitudinal Studies Mediating Membrane Molecular Molecular Genetics Multiprotein Complexes Mus Muscle Cells Mutation Myocardial Myocardium Neonatal Neurons Pathway interactions Physiological Play Property Proteins RNA Interference Receptor Protein-Tyrosine Kinases Regulation Reporting Research Research Proposals Risk Role Sick Sinus Syndrome Small Interfering RNA Sodium Specificity Syndrome System Testing Ventricular density fibroblast growth factor 13 genetic regulatory protein heart rhythm hippocampal pyramidal neuron in vivo insight neuronal excitability novel novel strategies programs protein complex research study small hairpin RNA therapeutic target trafficking voltage

项目摘要

DESCRIPTION (provided by applicant): Voltage-gated Na+ (Nav) channels are responsible for the rapid upstroke of the action potential in cardiac cells and play critical roles in controlling action potential durations and propagation. The primary Nav pore- forming (1) subunit in the myocardium is Nav1.5, encoded by SCN5A, and mutations in SCN5A have been linked to a number of cardiac rhythm disorders, including Long QT3 syndrome, Brugada syndrome, cardiac conduction disease, sick sinus syndrome, and atrial fibrillation. Accumulating evidence suggests that myocardial Nav channels function in multimeric protein complexes, comprising one Nav1 subunit, accessory (2) subunits and a number other accessory/regulatory proteins, although the roles of accessory and regulatory proteins in controlling channel expression, properties and subcellular distributions are not well understood. This R21 proposal will test the hypothesis that intracellular fibroblast growth factors (iFGFs) function as novel regulators of myocardial Nav1.5-encoded channels. This hypothesis is motivated by recent preliminary studies demonstrating that iFGF13 is expressed in adult and neonatal (mouse) ventricles and that iFGF13-targeted RNA interference markedly attenuates Nav current densities in (neonatal mouse ventricular) myocytes. There are two related aims in this proposal, and these will be pursued in parallel. Specifically, the studies outlined here will test the hypothesis that iFGF13 selectively regulates ventricular Nav currents and plays a physiological role in the generation of ventricular action potentials (aim #1). Parallel studies will explore the hypothesis that iFGF13 functions to regulate the stability, the trafficking and/or the subcellular localization of Nav1.5-encoded ventricular Nav channels (aim #2). To achieve these aims, the expression of iFGF13 will be manipulated in (mouse) ventricular myocytes in vitro using targeted gene "knockdown" strategies with small interfering RNAs (siRNAs), and the functional consequences of these manipulations on the properties and the cell surface expression of Nav (and other) channels will be determined. Parallel experiments will be completed on myocytes isolated from mice (Fgf13-/-) harboring a targeted disruption of the Fgf13 locus. It is anticipated that the studies proposed here will provide new and fundamentally important insights into the role(s) of the iFGFs in the dynamic regulation of myocardial Nav channels. In addition, the results of these studies will guide future investigations focused on delineating the molecular, cellular and systemic mechanisms involved in the dynamic regulation of myocardial membrane excitability and in the derangements in cardiac excitability linked to mutations in SCN5A. In the long term, it is anticipated that these studies will provide important new insights into the potential of the iFGFs as therapeutic targets to modulate Nav channel functioning in inherited and acquired cardiac rhythm disorders. PUBLIC HEALTH RELEVANCE: In the heart, voltage-gated sodium Na+ (Nav) channels are responsible for the rapid upstroke of the action potential and play roles in controlling action potential durations and propagation. These channels, therefore, are critical for the generation of normal cardiac rhythms. Changes in Nav channel expression and/or properties are observed in a number of inherited and acquired cardiac diseases, and these changes can have profound physiological consequences, including increasing the risk of potentially life-threatening cardiac arrhythmias. Accumulating evidence suggests that myocardial Nav channels function as components of macromolecular protein complexes, comprising pore-forming (1) subunits and a variety of accessory (2) subunits, although very little is presently known about the roles of these accessory subunits in the regulation of myocardial Nav channel stability, trafficking and/or properties. Combining in vivo and in vitro molecular genetic strategies with electrophysiological and biochemical approaches, this new research program is focused on defining the physiological role(s) of the novel family of Nav channel regulatory proteins, the intracellular fibroblast growth factors (iFGFs), in the regulation of myocardial Nav channel expression and functioning. These studies will provide new and fundamentally important insights into the physiological roles of the iFGFs in the dynamic regulation of myocardial Nav channels and myocardial membrane excitability. In the long term, these studies are also expected to provide important new insights into the potential of the iFGFs as therapeutic targets to modulate Nav channel functioning in inherited and acquired cardiac rhythm disorders.

描述（由申请人提供）：电压门控 Na+ (Nav) 通道负责心肌细胞中动作电位的快速上升，并在控制动作电位持续时间和传播中发挥关键作用。心肌中主要的 Nav 成孔 (1) 亚基是 Nav1.5，由 SCN5A 编码，SCN5A 的突变与许多心律失常有关，包括长 QT3 综合征、Brugada 综合征、心脏传导疾病、病态窦综合症和心房颤动。越来越多的证据表明，心肌 Nav 通道在多聚蛋白复合物中发挥作用，该复合物包含一个 Nav1 亚基、辅助 (2) 亚基和许多其他辅助/调节蛋白，尽管辅助和调节蛋白在控制通道表达、特性和亚细胞分布方面的作用尚不明确。不太理解。该 R21 提案将检验细胞内成纤维细胞生长因子 (iFGF) 作为心肌 Nav1.5 编码通道的新型调节剂的假设。这一假设源于最近的初步研究，该研究表明 iFGF13 在成人和新生儿（小鼠）心室中表达，并且 iFGF13 靶向 RNA 干扰显着减弱（新生儿小鼠心室）肌细胞中的 Nav 电流密度。该提案有两个相关的目标，这些目标将同时实现。具体来说，此处概述的研究将检验以下假设：iFGF13 选择性调节心室 Nav 电流并在心室动作电位的生成中发挥生理作用（目标#1）。平行研究将探讨 iFGF13 功能调节 Nav1.5 编码的心室 Nav 通道的稳定性、运输和/或亚细胞定位的假设（目标#2）。为了实现这些目标，将使用小干扰 RNA (siRNA) 的靶向基因“敲低”策略在体外操纵（小鼠）心室肌细胞中的 iFGF13 表达，以及这些操纵对性质和细胞表面表达的功能影响导航（和其他）频道的数量将被确定。平行实验将在从具有 Fgf13 基因座靶向破坏的小鼠 (Fgf13-/-) 中分离的肌细胞上完成。预计这里提出的研究将为 iFGF 在心肌 Nav 通道动态调节中的作用提供新的、根本性的重要见解。此外，这些研究的结果将指导未来的研究，重点是描绘心肌膜兴奋性动态调节以及与 SCN5A 突变相关的心脏兴奋性紊乱所涉及的分子、细胞和系统机制。从长远来看，预计这些研究将为 iFGF 作为调节遗传性和获得性心律失常中 Nav 通道功能的治疗靶点的潜力提供重要的新见解。公共健康相关性：在心脏中，电压门控钠 Na+ (Nav) 通道负责动作电位的快速上升，并在控制动作电位持续时间和传播方面发挥作用。因此，这些通道对于正常心律的产生至关重要。在许多遗传性和获得性心脏病中观察到 Nav 通道表达和/或特性的变化，这些变化可能产生深远的生理后果，包括增加潜在危及生命的心律失常的风险。越来越多的证据表明，心肌 Nav 通道作为大分子蛋白复合物的组成部分发挥作用，包括成孔 (1) 亚基和各种辅助 (2) 亚基，尽管目前对这些辅助亚基在调节心肌 Nav 通道稳定性、运输和/或特性。这项新研究计划将体内和体外分子遗传学策略与电生理学和生化方法相结合，重点是确定新型 Nav 通道调节蛋白家族、细胞内成纤维细胞生长因子 (iFGF) 在细胞中的生理作用。心肌 Nav 通道表达和功能的调节。这些研究将为 iFGF 在心肌 Nav 通道和心肌膜兴奋性动态调节中的生理作用提供新的、根本性的重要见解。从长远来看，这些研究也有望为 iFGF 作为调节遗传性和获得性心律失常中 Nav 通道功能的治疗靶点的潜力提供重要的新见解。