Mechanisms of cardiac Ca dysregulation and arrhythmias in ankyrin B deficiency

锚蛋白 B 缺乏导致心脏 Ca 失调和心律失常的机制

• 批准号: 8787535
• 负责人: Sanda Despa
• 金额: $ 33.96万
• 依托单位: UNIVERSITY OF KENTUCKY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-01 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8787535
• 关键词: Mechanisms; cardiac; Ca; dysregulation; arrhythmias

DESCRIPTION (provided by applicant): Ankyrin B (AnkB) is an "adaptor" protein that anchors several membrane proteins to the cytoskeleton. AnkB deficiency emerged as an important pro-arrhythmic factor a few years ago, when it was found that AnkB loss- of-function mutations generate human long QT syndrome type 4 (LQT4), the only LQT produced by alterations in a protein other than an ion channel. Besides LQT4, humans with AnkB loss-of-function mutations display a complex cardiac phenotype that also includes bradycardia, stress-induced ventricular arrhythmias and sudden cardiac death. The mechanisms responsible for such a phenotype are largely unknown. AnkB protein expression and distribution are drastically altered in the infarct border zone after myocardial infarction (MI). This may result in a mechanism for post-MI remodeling and arrhythmogenesis similar to that found in patients with AnkB loss-of-function mutations. Direct interaction with AnkB is required for the membrane targeting and stability of the Na/Ca exchanger (NCX) and Na/K-ATPase (NKA), which are essential in the regulation of cardiac [Na]i and [Ca]i and thus contractility and potential for triggered arrhythmias. The severity of the human cardiac phenotype generated by various AnkB loss-of-function mutants directly relates to the inability of those mutants to target NCX and NKA correctly to the cardiac myocyte sarcolemma. Thus, altered NCX and NKA expression and membrane distribution are key to the cardiac phenotype generated by AnkB loss-of-function. This phenotype is largely reproduced in mice heterozygous for a null mutation in AnkB (AnkB mice). Myocytes from AnkB mice show modestly reduced NCX and NKA protein expression, predominantly at the T-tubules, larger cellular and sarcoplasmic reticulum (SR) Ca load and increased frequency of early afterdepolarizations (EAD) and delayed afterdepolarizations (DAD). Despite its physiological and pathophysiological significance, the role of AnkB in regulating cardiac [Ca]i and arrhythmogenesis is poorly understood. The overall goal of this proposal is to decipher the mechanisms responsible for altered cardiac Ca regulation and triggered arrhythmias induced by AnkB loss-of-function (inherited and acquired). We will combine measurements of [Na]i and [Ca]i (in the bulk and junctional cleft), patch-clamp and molecular biology techniques in isolated cardiac myocytes in three Specific Aims. First, I will test several specific hypotheses aimed at understanding how AnkB affects intracellular Ca in the heart. Aim 2 will focus on the mechanisms responsible for the occurrence of early and delayed afterdepolarizations in myocytes with AnkB deficiency. In the final aim I will test the hypothesis that AnkB proteolysis by calpain and the ensuing remodeling in NCX and NKA is a more general mechanism for triggered ventricular arrhythmias. These studies will both advance our understanding of how AnkB affects cardiac [Ca]i regulation and will provide key mechanistic information that could lead to the development of new treatments for patients with ventricular arrhythmias associated with inherited or acquired AnkB loss-of-function.

描述（由申请人提供）：Ankyrin B（ANKB）是一种“衔接子”蛋白，可将几种膜蛋白固定在细胞骨架上。几年前，ANKB缺乏症是一种重要的促性心律失常因素，当时发现ANKB损失 - 功能突变会产生人类长QT综合征4型（LQT4），这是唯一由蛋白质以外的蛋白质改变产生的LQT离子通道。除LQT4外，具有ANKB功能丧失突变的人类还显示出复杂的心脏表型，其中还包括心动过缓，压力诱导的心室心律不齐和心脏猝死。负责这种表型的机制在很大程度上未知。心肌梗塞（MI）后，在梗塞边界区域中，ANKB蛋白表达和分布发生了巨大变化。这可能会导致一种与ANKB功能丧失突变患者相似的MI后重塑和心律失常发生的机制。与Na/Ca交换器（NCX）和Na/K-ATPase（NKA）的膜靶向和稳定性需要直接相互作用，这对于调节心脏[Na] I和[Ca] I的调节至关重要收缩性和触发心律不齐的潜力。各种ANKB功能丧失突变体产生的人类心脏表型的严重程度直接与这些突变体正确靶向NCX和NKA的能力与心肌肌细胞肌瘤正确靶向NKA。因此，改变的NCX和NKA表达和膜分布是ANKB功能丧失产生的心脏表型的关键。该表型在ANKB（ANKB小鼠）中的杂合突变中基本上是复制的。来自ANKB小鼠的肌细胞显示出适度降低的NCX和NKA蛋白表达，主要是在T管上，较大的细胞和肌浆网（SR）Ca载荷（SR）Ca负载以及早期过度后发生后（EAD）早期频率（EAD）和延迟过度过度分离的频率（爸爸）。尽管具有生理和病理生理学意义，但ANKB在调节心脏[CA] I和心律失常发生方面的作用知之甚少。该提案的总体目标是破译负责改变心脏CA调节的机制，并触发ANKB功能丧失（遗传和获取）引起的心律不齐。我们将结合[Na] I和[Ca] I（在散装和连接裂缝中），斑块钳和分子生物学技术中分离的心肌细胞中的斑块钳和分子生物学技术的测量值。首先，我将测试一些旨在了解ANKB如何影响心脏内CA的特定假设。 AIM 2将重点放在导致ANKB缺乏肌细胞早期和延迟造影后发生后发生后发生后发生后发生的机制。在最终目的中，我将检验以下假设：Calpain的ANKB蛋白水解以及NCX和NKA中随之而来的重塑是触发心律不齐的更通用机制。这些研究都将提高我们对ANKB如何影响心脏[CA]调节的理解，并将提供关键的机械信息，这可能会导致为与遗传性或获得ANKB功能丧失相关的心室心律不齐患者开发新治疗。