Regulation of calcium-activated potassium channels by lipid messengers

描述（由申请人提供）：大电导Ca2+和电压门控K+ (Slo1 BK)通道起着许多生理和病理生理作用，它们的变构门控机制受到多种细胞信号通路的调节。越来越多的证据表明，某些脂质可能起到信号分子的作用。我们提出揭示两种脂质信使磷脂酰肌醇4,5-二磷酸（PIP2）和二十二碳六烯酸（DHA）（富含油性鱼类的omega-3长链多不饱和脂肪酸）调节Slo1 BK通道的生物物理和物理化学机制。最近报道了PIP2对Slo1 BK通道的影响，但其机制尚不清楚。我们将通过进行彻底的机械电生理测量来填补这一关键的知识空白。此外，我们将确定对PIP2的作用很重要的辅助β亚基的结构决定因素。我们的电生理测量将与分离物和纯化的Slo1门控环蛋白的色氨酸荧光测量相补充。作为一种新兴的脂质信使，DHA在Slo1通道上的作用的生物物理和物理化学机制也将使用电生理和荧光方法进行类似的研究。该研究结果有望提供PIP和DHA作用于Slo1通道变构门控机制的明确机制，并建立Slo1通道是一种omega-3脂肪酸受体的新范式。DHA对Slo1 BK通道的调节可能是ω -3长链脂肪酸促进健康作用的基础。

DESCRIPTION (provided by applicant): Large-conductance Ca2+- and voltage-gated K+ (Slo1 BK) channels play numerous physiological and pathophysiological roles and their allosteric gating mechanism is subject to modulation by a variety of cellular signaling pathways. Increasing evidence suggests that certain lipids may serve as signaling molecules. We propose to reveal the biophysical and physicochemical mechanisms of modulation of Slo1 BK channels by two lipid messengers, phosphatidylinositol 4,5-bisphosphate (PIP2) and docosahexaenoic acid (DHA), an omega-3 long-chain polyunsaturated fatty acid enriched in oily fish. The effect of PIP2 on the Slo1 BK channel was reported recently but the mechanism is only poorly known. We will fill this critical knowledge gap by performing thorough mechanistic electrophysiological measurements. Furthermore, we will identify the structural determinants of the auxiliary beta subunit important for the action of PIP2. Our electrophysiological measurements will be complemented with measurements of tryptophan fluorescence of the isolate and purified Slo1 gating ring protein. The biophysical and physicochemical mechanisms of the action of DHA, an emerging lipid messenger, on the Slo1 channel will be also investigated similarly using the electrophysiological and fluorescence methods. The research outcome is expected to provide definitive mechanisms of the PIP and DHA actions on the allosteric gating mechanism of the Slo1 channel and establish the novel paradigm that the Slo1 channel is an omega-3 fatty acid receptor. The modulation of the Slo1 BK channel by DHA may underlie the health-promoting effects of omega-3 long-chain fatty acids.