Molecular Physiology of KCNE Potassium Channel Subunits

KCNE 钾通道亚基的分子生理学

• 批准号: 6923262
• 负责人: Alfred L. George
• 金额: $ 36.51万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-05-15 至 2009-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6923262
• 关键词: RNA interference; SDS polyacrylamide gel electrophoresis; cardiac myocytes; cardiovascular disorder risk; clinical research; dogs; electrophysiology; gene expression; gene induction /repression; gene interaction; heart disorder; heart function; human tissue; immunocytochemistry; immunoprecipitation; laboratory mouse; membrane proteins; molecular pathology; pore forming protein; potassium channel; regulatory gene; tissue /cell culture

DESCRIPTION (provided by applicant): Voltage-gated potassium (Kv) channels are modulated by at least three distinct classes of accessory proteins including the KCNE family of single transmembrane proteins. In the human genome, KCNE proteins are encoded by five genes designated KCNE1 through KCNE5. The physiological importance of three KCNE genes has been demonstrated by associations with inherited disorders of cardiac and skeletal muscle excitability. Mutation of KCNE 1 impairs generation of the slowly activating cardiac delayed rectifier current (IKS), a critical potassium current contributing to myocardial repolarization that can be reconstituted in vitro by heterologous co-expression of recombinant KCNQ1 potassium channels with KCNE1. Other KCNE proteins also exert diverse functional effects on heterologously expressed KCNQ1. Co-expression of KCNQ1 with KCNE3, KCNE4 or KCNE5 is particularly interesting with functional effects ranging from constitutive activation (KCNE3) to suppression (KCNE4, KCNE5). We have recently demonstrated that all KCNE genes are expressed in human cardiac myocytes to varying degrees. We have further demonstrated that KCNE4 and KCNE5 exert potent inhibitory influences on heterologously expressed IKappaS and KCNE4 appears dominant over KCNE1 in this context. In addition to potential interactions with KCNQ1, KCNE proteins affect the functional properties of other cloned potassium channels. The physiological relevance of these in vitro observations remains uncertain. The observations that multiple KCNE proteins can modulate KCNQ1 channels to produce diverse biophysical phenotypes raises several intriguing questions with important implications for understanding the regulation of IKS as well as other potassium currents in normal and diseased heart. Can different KCNE subtypes compete functionally with KCNE1 for modulation of KCNQ1? Can KCNQ1 associate with more than one KCNE subtype in the same cell and at the same time? Do KCNE4 and KCNE5 participate in the pathophysiology of heart failure and other arrhythmia-prone conditions? In this proposal, we outline several experimental approaches for defining the physiological importance of KCNE subunits with a particular focus on KCNE4 and KCNE5. In Specific Aim 1, we will use a heterologous expression system to assess the range of functional and biochemical interactions possible between KCNQ1 and different KCNE subunits with a focus on KCNE4 and KCNE5. In Specific Aim 2, we will test the hypothesis that KCNE4 and KCNE5 are physiologically significant regulators of potassium currents in native cardiac myocytes by using two independent gene silencing approaches. Finally in Specific Aim 3, we will examine expression patterns of KCNQ1 and KCNE genes in diseased human heart and in an experimental canine model of ventricular arrhythmia susceptibility. Results from these studies will have implications for advancing our knowledge of the complex interplay between pore-forming and accessory subunits that comprise human potassium channels expressed in the heart and in other tissues.

描述（由申请人提供）：电压门控钾（KV）通道由至少三种不同的辅助蛋白（包括单跨膜蛋白的KCNE家族）调节。在人类基因组中，KCNE蛋白质由指定KCNE1至KCNE5的五个基因编码。与心脏和骨骼肌肉兴奋性的遗传性疾病的关联证明了三个KCNE基因的生理重要性。 KCNE 1的突变会损害缓慢激活心脏延迟整流剂电流（IKS）的产生，这是一种临界钾电流，有助于心肌复极化，可以通过与KCNE1的重组KCNQ1钾通道的异源共表达在体外重构。其他KCNE蛋白也对异源表达的KCNQ1产生了不同的功能作用。 KCNQ1与KCNE3，KCNE4或KCNE5的共表达特别有趣，功能效应从本构激活（KCNE3）到抑制（KCNE4，KCNE5）。我们最近证明，所有KCNE基因在人类心脏肌细胞中以不同程度表示。我们进一步证明，在这种情况下，KCNE4和KCNE5对异源表达的Ikappas产生了有效的抑制作用，而KCNE4在KCNE1上显得主导地位。除了与KCNQ1的潜在相互作用外，KCNE蛋白还会影响其他克隆钾通道的功能特性。这些体外观察的生理相关性仍然不确定。 观察到多种KCNE蛋白可以调节KCNQ1通道产生不同的生物物理表型，这引发了几个有趣的问题，对了解IKS的调节以及正常心脏和患病心脏的其他钾电流具有重要意义。不同的KCNE子类型是否可以与KCNE1在功能上竞争KCNQ1的调制？ KCNQ1可以在同一单元格和同时与多个KCNE亚型相关联吗？ KCNE4和KCNE5是否参与心力衰竭和其他心律失常疾病的病理生理？ 在此提案中，我们概述了几种实验方法，以定义KCNE亚基的生理重要性，特别关注KCNE4和KCNE5。在特定目标1中，我们将使用异源表达系统来评估KCNQ1和不同KCNE亚基之间可能的功能和生化相互作用的范围，重点是KCNE4和KCNE5。在特定的目标2中，我们将通过使用两种独立的基因沉默方法来检验假设KCNE4和KCNE5是天然心肌细胞中钾电流的生理意义调节剂。最后，在特定的目标3中，我们将检查患病的人心脏中KCNQ1和KCNE基因的表达模式，以及在心室心律失常易感性的实验犬模型中。这些研究的结果将对我们对孔形成孔形成和辅助亚基之间的复杂相互作用的了解有影响，这些孔形成和包括人类钾通道在心脏和其他组织中表达。