Redox Regulation of Auxillary B subunits of BK Channels

描述(申请人提供)：这项工作的长期目标是了解BK型钙(钙)激活钾(K+)通道调节的分子机制。BK通道在不同类型的细胞中广泛表达，并表现出与其生理作用相适应的显著功能多样性。BK电流将膜下钙离子浓度的变化与膜电位和兴奋性的变化联系在一起。BK通道的激活与缺血发作时神经元的保护有关，氧化条件的改变对BK通道的调节可能对细胞的电兴奋性产生深远的影响。BK通道至少由四个成孔亚基组成。此外，还有四个辅助β亚基，它们不仅在组织分布方面不同，而且在所产生的BK通道的功能特性方面也不同。在脑组织中发现了三个贝塔亚基，即贝塔2、贝塔3和贝塔4，其中贝塔4是最丰富的脑亚单位。 在不同组织中，BK通道功能的差异很大程度上是由β亚基构成的，并且由于1β亚基细胞外环中含有丰富的半胱氨酸残基，因此可能是氧化条件改变BK通道功能的一个重要的潜在靶点。 本项目将利用电生理学和分子生物学相结合的方法，研究BK-1β亚氨酸半胱氨酸残基的氧化还原反应是否在调节脑内BK通道功能中发挥重要作用。首先，将考察氧化还原反应影响β亚基关键功能作用的能力，包括失活、门控明显的钙依赖、内向电流整流和药理学。第二，特殊的作用 半胱氨酸在这些影响中的残留量将被定义。第三，将测试可能影响关键半胱氨酸氧化还原条件的潜在生理机制。 这些研究将提供关于BKβ亚基在定义BK通道功能中的作用以及氧化还原反应在通道功能调节中的作用的分子机制的明确信息。BK通道在多种可兴奋细胞的正常功能中具有广泛的重要性。在不同的组织中，BK通道参与神经元兴奋性、平滑肌松弛、突触传递和激素释放的调节。此外，在缺血条件下，BK通道可能作为神经保护剂发挥关键作用。这项工作将为BK通道的这种作用可能发生的机制提供重要的见解。

DESCRIPTION (provided by applicant): The long-term aim of this work is to understand the molecular mechanisms of regulation of BK-type calcium (Ca2+)-activated potassium (K+) channels. BK channels are widely expressed among different cell types and exhibit significant functional diversity suited to their physiological roles. BK currents couple changes in sub membrane Ca2+ concentrations to changes in membrane potential and excitability. Activation of BK channels have been implicated in protection of neurons during ischemic attacks and regulation of BK channels by changes in oxidative conditions may have a profound impact on cellular electrical excitability. BK channels consist minimally of four pore-forming subunits. In addition, there are four auxiliary beta subunits, which differ not only in terms of tissue distribution, but also in terms of the functional properties of the resulting BK channels. Three beta subunits, the beta2, beta3, and beta4,are found in brain tissue, with beta4 being the most abundant brain subunit. The beta subunits account for much of the diversity of BK channel function among different tissues and, because of an abundance of cysteine residues in the 1betasubunit extracellular loop, may be an important potential target by which changes in oxidative conditions may alter BK channel function. Using methods of electrophysiology combined with molecular biology, this project will examine whether redox reactions involving cysteine residues of BK 1betasubunits particularly from brain may play an important role in modulating BK channel function in brain. First, the ability of redox reactions to influence the key functional roles of beta subunits, including inactivation, apparent Ca2+-dependence of gating, inward current rectification, and pharmacology will be examined. Second, the role of particular cysteine residues in these effects will be defined. Third, potential physiological mechanisms by which the redox conditions of key cysteines may be influenced will be tested. These studies will provide definitive information about molecular mechanisms that underlie the role of BK beta subunits in defining BK channel function and the role of redox reactions in regulation of channel function. BK channels are of broad importance in the normal functioning of a variety of excitable cells. Among different tissues, BK channels contribute to regulation of neuronal excitability, smooth muscle relaxation, synaptic transmission and hormone release. Furthermore, BK channels may play a key role as neuroprotectants during ischemic conditions. This work will provide important insight into the mechanisms by which this role of BK channels may occur.