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Abstract：
Inwardly rectifying potassium (Kir) channels establish and regulate the resting membrane potential of excitable cells in the heart, brain, and other peripheral tissues. Phosphatidylinositol 4,5-bisphosphate (PIP2) is a key direct activator of ion channels, including Kir channels. The gasotransmitter carbon monoxide has been shown to regulate Kir channel activity by altering channel-PIP2 interactions. Here, we tested in two cellular models the effects and mechanism of action of another gasotransmitter, hydrogen sulfide (H2S), thought to play a key role in cellular responses under ischemic conditions. Direct administration of sodium hydrogen sulfide as an exogenous H2S source and expression of cystathionine γ-lyase, a key enzyme that produces endogenous H2S in specific brain tissues, resulted in comparable current inhibition of several Kir2 and Kir3 channels. This effect resulted from changes in channel-gating kinetics rather than in conductance or cell-surface localization. The extent of H2S regulation depended on the strength of the channel-PIP2 interactions. H2S regulation was attenuated when channel-PIP2 interactions were strengthened and was increased when channel-PIP2 interactions were weakened by depleting PIP2 levels. These H2S effects required specific cytoplasmic cysteine residues in Kir3.2 channels. Mutation of these residues abolished H2S inhibition, and reintroduction of specific cysteine residues back into the background of the cytoplasmic cysteine-lacking mutant rescued H2S inhibition. Molecular dynamics simulation experiments provided mechanistic insights into how potential sulfhydration of specific cysteine residues could lead to changes in channel-PIP2 interactions and channel gating.
摘要：
内向整流钾（Kir）通道建立和调节心脏中兴奋细胞的静息膜电位，大脑,和其他外周组织。磷脂酰肌醇4,5-二磷酸(PIP2)是离子通道的关键直接激活剂,包括基尔频道。已显示气体发射器一氧化碳通过改变通道-PIP2相互作用来调节Kir通道活性。这里,我们在两个细胞模型中测试了另一种气体发射器的作用和机制，硫化氢(H2S),被认为在缺血条件下的细胞反应中起关键作用。直接施用硫化氢钠作为外源性H2S来源并表达胱硫醚γ-裂解酶，一种在特定脑组织中产生内源性H2S的关键酶，导致几个Kir2和Kir3通道的相当的电流抑制。这种效应是由通道门控动力学的变化而不是电导或细胞表面定位引起的。H2S调节的程度取决于通道-PIP2相互作用的强度。当通道-PIP2相互作用增强时，H2S调节减弱，而当通过消耗PIP2水平减弱通道-PIP2相互作用时，H2S调节增强。这些H2S效应需要Kir3.2通道中特定的细胞质半胱氨酸残基。这些残基的突变消除了H2S抑制，并将特定的半胱氨酸残基重新引入细胞质中缺乏半胱氨酸的突变体拯救了H2S抑制。分子动力学模拟实验提供了有关特定半胱氨酸残基的潜在硫酸化如何导致通道-PIP2相互作用和通道门控变化的机制见解。
