Molecular Mechanisms of Ankyrin-B-based Arrhythmia

描述（由申请人提供）：心脏兴奋性缺陷是人类心律失常和心源性猝死的基础，是发达国家死亡的主要原因。最近的研究结果表明，基于影响心脏离子通道和转运体的表达/亚细胞定位的基因突变，人类心律失常的新范式。人类4型长QT综合征（LQT4）是由膜适配器锚蛋白b （AnkB）的功能缺失突变引起的。LQT4受试者和AnkB表达降低的小鼠表现出相似的复杂心脏表型，包括心房、心室、传导缺陷和心源性猝死风险。然而，AnkB多肽的分子特性，AnkB在特殊心脏细胞中的表达范围，以及AnkB多肽在心脏兴奋性中的细胞作用仍然是关键的，但尚未解决的问题。此外，AnkB在正常心脏中的调控机制和人类心律失常的功能障碍仍未得到解决。这项研究计划的长期目标是了解AnkB在心脏中功能的分子基础。我们假设，由于离子通道/转运体运输和膜稳定性的缺陷，整个心脏中AnkB多肽的协调功能障碍创造了人类4型长QT综合征的复杂表型。具体目的是：1)表征AnkB亚型在不同心脏可兴奋细胞类型中的表达和亚细胞分布。2)使用最新开发的慢病毒技术确定AnkB在成人心肌细胞中离子通道/转运体运输和定位中的细胞作用。3)描述心脏中AnkB调控的机制，并确定与致命性心律失常相关的人类AnkB功能缺失突变如何影响这种调控。心肌细胞中离子通道和转运体靶向、定位和稳定性的细胞通路基本上是未知的，但为未来的心脏治疗提供了一个令人兴奋的新靶点。我们建议使用最近开发的表达技术来阐明ankb依赖的细胞途径在原代心肌细胞的生理背景下对离子通道和转运体的靶向、定位和稳定性的分子机制。预计这一信息将促进对基于ankb的人类致命性心律失常以及与异常Ca2+稳态相关的获得性心律失常的机制的理解，并开始确定细胞兴奋性调节的潜在未来分子靶点。

DESCRIPTION (provided by applicant): Defects in cardiac excitability are the basis for human arrhythmia and sudden cardiac death, a leading cause of mortality in developed countries. Recent findings demonstrate a new paradigm for human arrhythmia based on gene mutations that affect the expression/subcellular localization of cardiac ion channels and transporters. Human type 4 long QT syndrome (LQT4) results from loss-of-function mutations in the membrane adapter ankyrin-B (AnkB). Subjects with LQT4, and mice with reduced AnkB expression display similar complex cardiac phenotypes including atrial, ventricular, conduction defects, and risk of sudden cardiac death. However, the molecular identities of AnkB polypeptides, scope of AnkB expression in specialized cardiac cells, and cellular role(s) for AnkB polypeptides for cardiac excitability remain critical, yet unanswered questions. Moreover, the mechanisms underlying AnkB regulation in normal heart, and dysfunction in human arrhythmia remain unsolved. The long-term objective of this research proposal is to understand the molecular basis for AnkB function in the heart. We hypothesize that coordinate dysfunction of AnkB polypeptides throughout the heart create the complex phenotype of human type 4 long QT syndrome due to defects in ion channel/transporter trafficking and membrane stability. The specific aims are to: 1) Characterize the expression and subcellular distribution of AnkB isoforms in diverse excitable cell types of heart. 2) Define the cellular role(s) of AnkB for ion channel/transporter trafficking and localization in adult cardiomyocytes using recently developed lentiviral techniques. 3) Characterize the mechanisms underlying AnkB regulation in heart, and determine how human AnkB loss-of-function mutations associated with fatal arrhythmia affect this regulation. The cellular pathways underlying ion channel and transporter targeting, localization, and stability in cardiomyocytes are essentially unknown but present an exciting new target for future cardiac therapies. We propose to use recently developed expression techniques to elucidate the molecular mechanisms underlying AnkB-dependent cellular pathways for ion channel and transporter targeting, localization, and stability in the physiological context of the primary cardiomyocyte. It is anticipated that this information will advance understanding of mechanisms underlying AnkB-based human fatal human arrhythmia as well as acquired cardiac arrhythmias associated with abnormal Ca2+ homeostasis, and begin to define potential future molecular targets for the regulation of cellular excitability.