Molecular Components of A-type K+ Channels

描述（由申请人提供）：快速瞬态a型K+电流（IA），如阈下激活的神经元体树突a型K+电流（ISA）和心室肌细胞的快速瞬态外向K+电流（Ito）对大脑和心脏的正常功能至关重要。在病理生理条件下（如心脏缺血和脑卒中发生期间），这些电流的异常会导致与这些疾病条件相关的异常。该项目解决了负责产生这些电流的离子通道的分子性质。它试图建立这些通道的分子组成，并阐明鉴定的成分的生理意义。在阐明ISA和Ito通道的分子组成方面取得了进展，并确定了两个关键成分，Kv4孔隙形成亚基和KCHIP相关蛋白。然而，在许多神经元中，ISA通道的动力学比由Kv4和KCHIP蛋白组成的通道更快。最近有证据表明，在脑mRNA中存在编码一种因子（称为KAF）的转录本，这可能是一种新的相关亚基，可以加速Kv4通道的动力学。此外，利用生化方法鉴定了一种新的Kv4通道相关蛋白（DPPX），并获得了该蛋白负责KAF活性的证据。本项目的目的是验证DPPX是许多神经元中Kv4通道的重要组成部分，并有助于天然a型K+通道的性质和多样性的假设。目的1将研究DPPX对异源表达系统中Kv4通道功能的影响。目的2将利用原位杂交和免疫组织化学研究DPPX蛋白在大脑中的表达位置和时间，以及它们与Kv4通道其他已知组分的关系。Aim 3将利用基因靶向和反义技术更直接地研究DPPX蛋白在神经元中的生理意义。

DESCRIPTION (provided by applicant): Fast transient A-type K+ currents (IA), such as the subthreshold-activating somato-dendritic A-type K+ currents in neurons (ISA) and the fast transient outward K+ current in cardiac ventricular myocytes (Ito) are essential for the proper functioning of the brain and the heart. During pathophysiological conditions (e.g. ischemia in the heart and during the occurrence of stroke in brain) abnormalities in these currents contribute to the abnormalities associated with these disease conditions. This project addresses the molecular nature of the ion channels responsible for the generation of these currents. It seeks to establish the molecular composition of these channels and to elucidate the physiological significance of the identified components. Progress has been made in elucidating the molecular composition of the channels mediating the ISA and the Ito, and two key components, Kv4 pore-forming subunits and KCHIP associated proteins, have been identified. However, the kinetics of ISA channels in many neurons is faster than that of channels composed of Kv4 and KCHIP proteins. Evidence has been recently obtained for the presence in brain mRNA of transcripts encoding a factor (termed KAF), probably a novel associated subunit, which accelerates the kinetics of Kv4 channels. Moreover, a novel Kv4 channel associated protein (DPPX) has been identified utilizing biochemical methods, and evidence that this protein is responsible for KAF activity has been obtained. The goal of this project is to test the hypothesis that DPPX is an important component of Kv4 channels in many neurons and contributes to the properties and diversity of native A-type K+ channels. Aim 1 will investigate the effects of DPPX on Kv4 channel function in heterologous expression systems. Aim 2 will investigate where and when DPPX proteins are expressed in brain and their relationship to the other known components of Kv4 channels utilizing in-situ hybridization and immunohistochemistry. Aim 3 will investigate more directly the physiological significance of DPPX proteins in neurons utilizing gene targeting and antisense technology.