KCNQ CHANNELS: GATING AND SUBUNITS MODULATION

 描述（由申请人提供）：本项目旨在揭示KCNQ K+通道（KCNQ 1 -5）功能的主要分子基础，这些通道在心脏、大脑、内耳和上皮中非常重要。这些通道的异常功能与心律失常、癫痫、耳聋和胃癌有关。这些渠道的重要性基于两个突出的属性。首先，所有这些通道，当没有辅助的α亚基表达时，被负范围的电压激活（在-60 mV左右开始激活）。在静息膜电位之上被激活，通过神经元中KCNQ 2和3的M电流降低膜兴奋性，并作为膜放电的“制动器”。第二，KCNE家族K+通道β亚基的关联，从根本上改变了KCNQ 1的门控、渗透和药理学特性，决定了KCNQ 1的生理作用。KCNQ 1与KCNE 1结合形成心脏中调节动作电位时程的IKs通道，并与KCNE 2或KCNE 3结合形成上皮中对离子转运重要的组成性开放背景K+通道。这些特性背后的机制是什么？这是一个长期存在的问题，其答案将为理解和治疗KCNQ相关疾病提供基础。我们认为这个问题的答案在于KCNQ 1激活的新机制。在KCNQ 1的电压依赖性激活过程中，电压感受器结构域（VSD）分两步移动，从静息状态到中间状态，然后到激活状态;通道孔在VSD的中间状态和激活状态下都打开，但VSD-孔相互作用在VSD的不同状态下不同，从而改变通道门控、离子渗透和药理学。KCNE 1通过抑制通道的中间开口来调节通道功能。在这个项目中，我们希望在三个具体目标中测试这个假设的各个方面。1)我们希望确定的结构图案是重要的VSD孔相互作用在中间和激活状态，分别。KCNQ 1和KCNE 1的突变与长QT综合征（LQT）相关，该综合征使患者易于发生致命性心律失常，本研究将揭示某些LQT突变是否会改变VSD-孔相互作用。2)我们将研究是否抑制中间体和增强激活的开口是KCNE 1相关的主要功能变化的主要机制，包括对β-肾上腺素能刺激和失活门控的反应。携带KCNQ 1突变的LQT患者在β-肾上腺素能通路受到刺激时，经常出现心律失常症状，如晕厥和运动时猝死。因此，这项研究对于了解LQT的分子基础特别有意义。3)我们希望揭示KCNE 2和3是否通过改变VSD激活、孔开放或VSD-孔相互作用使通道组成性开放。我们将确定这些通道是否处于中间或激活的开放状态，以及它们对药物和细胞信号分子的反应。

 DESCRIPTION (provided by applicant): This project is to reveal the major molecular bases for the function of KCNQ K+ channels (KCNQ1-5) that are important in the heart, brain, inner ear and epithelia. The aberrant functions of these channels are associated with cardiac arrhythmia, epilepsy, deafness, and gastric cancer. The importance of these channels is based on two prominent properties. First, all these channels, when expressed without auxiliary ß subunits, are activated by voltages at negative ranges (start activating around -60 mV). Being activated just above the resting membrane potential, the M-current through KCNQ2 and 3 in neurons reduces membrane excitability and acts as a "brake" to membrane discharge. Second, the association of the KCNE family K+ channel ß subunits, which radically alter gating, permeation and pharmacological properties of KCNQ1, determines the physiological role of KCNQ1. KCNQ1 associates with KCNE1 to form the IKs channel in the heart that regulates action potential duration, and with KCNE2 or KCNE3 to form the constitutively open background K+ channels in epithelia important for ion transport. What are the mechanisms underlying these properties? This is a long-standing question whose answer will provide the basis for the understanding and treatment of KCNQ associated diseases. We propose that the answer to this question lies in a novel mechanism for KCNQ1 activation. During voltage dependent activation in KCNQ1, the voltage sensor domain (VSD) moves in two steps, from the resting to intermediate and then to activated state; the channel pore opens at both intermediate and activated states of VSD, but the VSD-pore interaction differs at different states of VSD to alter channel gating, ion permeation and pharmacology. KCNE1 modulates channel function by suppressing the intermediate openings of the channel. In this project we wish to test various aspects of this hypothesis in three specific aims. 1) We wish to identify the structural motifs that are important for the VSD-pore interaction at intermediate and activated states, respectively. Mutations in KCNQ1 and KCNE1 are associated with long QT syndrome (LQT) that predispose patients to fatal cardiac arrhythmia, this study will reveal if some of the LQT mutations alter VSD-pore interaction. 2) We will examine if suppression of the intermediate and potentiation of the activated openings is the main mechanism for major functional changes upon KCNE1 association including the response to ß- adrenergic stimulation and inactivation gating. LQT patients with KCNQ1 mutations often experience symptoms of cardiac arrhythmia such as syncope and sudden death during exercise when ß-adrenergic pathway is stimulated. This study is therefore particularly significant for the understanding of molecular bases of LQT. 3) We wish to reveal if KCNE2 and 3 make the channel constitutively open by altering VSD activation, pore opening or VSD-pore interaction. We will identify if these channels are at intermediate or activated open states and their responses to drugs and cellular signaling molecules.