Targeting of Voltage-gated K+ Channels to Lipid Rafts

描述：神经元和肌膜内的离子通道调节是一种 神经和精神疾病患者电兴奋性的重要决定因素 心血管系统、骨骼肌、胃肠道和子宫。电压门控K+ 通道(Kv通道)在设置静息电位中起着重要作用 并确定这些功能不同的系统中的复极化。近期 有证据表明，专门的微域通常被称为脂质 木筏存在于大多数质膜的平面内。这些域是 富含胆固醇和鞘脂，并浓缩多项信号 转导分子。在初步数据部分，我们演示了 电压门控K+通道Kv2.1、Kv1.1、Kv1.5和Kv1.4，而不是Kv4.2， 靶向异源表达系统和大鼠脑中的脂筏。在……里面 此外，Kv2.1和Kv1.5可能位于不同的浮筏隔间。 细胞胆固醇的耗尽改变Kv2.1相关的浮力 将Kv2.1的失活中点漂移30-40 mV 影响峰值电流密度或通道激活。Kv2.1的孵化 用神经酰胺合成酶抑制剂伏马菌素B表达细胞会引起 失活曲线上也有类似的变化。神经酰胺既是一种浮筏成分，也是一种 一种细胞内信号分子。因此，浮筏协会在功能上是 意义重大，因为这种失活的转变将导致 Kv2.1通道在以下范围内功能静音的百分比 生理膜电位。此外，初步数据显示， 伏马菌素B诱导Kv2.1从细胞体到远端的错误定位 培养神经元中的树突，表明RAFT相关的信号机制是 参与Kv2.1目标定位。具体目标将：1)解决机制问题 参与将Kv2.1定位于脂筏，重点放在 亚基组成和通道跨膜结构域；2)检查 Kv2.1与脂筏结构域结合的功能意义 重点研究神经酰胺信号转导途径，3)研究神经酰胺信号转导途径与 RAFT联合、神经酰胺信号转导和神经元细胞表面定位； 和4)开始初步工作，目的是纯化含Kv 2.1的脂类 大脑中的木筏。 这项拟议的研究考察了KV通道研究的一个新领域，该领域将 在多种组织系统中都有重要的意义。

DESCRIPTION: Ion channel regulation within neuronal and muscle membranes is an important determinant of electrical excitability in the nervous and cardiovascular systems, skeletal muscle, GI tract, and uterus. Voltage-gated K+ channels (Kv channels) play an important role in setting the resting potential and determining repolarization in these functionally diverse systems. Recent evidence suggests that specialized microdomains commonly referred to as lipid rafts exist within the plane of most plasma membranes. These domains are enriched in cholesterol and sphingolipids and concentrate a number of signal transduction molecules. In the Preliminary Data section, we demonstrate that the voltage-gated K+ channels, Kv2.1, Kv1.1, Kv1.5, and Kv1.4, but not Kv4.2, target to lipid rafts in both heterologous expression systems and rat brain. In addition, Kv2.1 and Kv1.5 probably reside in different raft compartments. Depletion of cellular cholesterol alters the buoyancy of the Kv2.1-associated rafts and shifts the midpoint of Kv2.1 inactivation by 30-40 mV without affecting peak current density or channel activation. Incubation of Kv2.1 expressing cells with fumonisin B, an inhibitor of ceramide synthase, causes a similar shift in the inactivation curve. Ceramide is both a raft component and an intracellular signaling molecule. Thus, raft association is functionally significant, for such a shift in the inactivation will result in a large percentage of the Kv2.1 channels being functionally silenced in the range of physiological membrane potentials. In addition, the preliminary data suggest fumonisin B induces mistargeting of Kv2.1 from the cell body to the distal dendrites in cultured neurons, suggesting raft-related signaling mechanisms are involved in Kv2.1 targeting. The Specific Aims will 1) address the mechanisms involved in the targeting of Kv2.1 to lipid rafts, with emphasis placed on subunit composition and channel transmembrane domains; 2) examine the functional significance of Kv2.1 association with lipid raft domains with emphasis on ceramide signaling pathways, 3) examine the relationship between raft association, ceramide signaling, and cell surface localization in neurons; and 4) begin initial work aimed at purification of Kv 2.1-containing lipid rafts from brain. This proposed research examines a new area in Kv channel research that will have important implications in multiple tissue systems.