Inhibition of a background potassium channel by Gq protein α-subunits
Inhibition of a background potassium channel by Gq protein α-subunits
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DOI：
10.1073/pnas.0507710103
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发表时间：
2006-02-28
影响因子：
11.1
通讯作者：
Bayliss, DA
中科院分区：
文献类型：
--
作者：
Chen, XD;Talley, EM;Bayliss, DA
双孔域K+通道提供神经元背景电流，建立静息膜电位和输入电阻；它们的调节提供了一种普遍的调节细胞兴奋性的机制。所谓的TASK通道亚基（TASK-1和TASK-3）被广泛表达，它们被通过G α q家族蛋白发出信号的受体强烈抑制。在这里，我们在完整和无细胞系统中操纵G蛋白表达和膜磷脂酰肌醇4,5-二磷酸（PIP2）水平，以提供电生理和生化证据，证明在缺乏磷脂酶C （PLC）活性的情况下，G α q连接受体对TASK通道的抑制不减弱，而是涉及活化的G α q亚基与通道的关联。与已知依赖于PLC的pip2敏感的Kir3.4（S143T）同源通道相比，受体介导的TASK通道抑制对PLC β 1-ct小基因结构的抑制更快，更不敏感。TASK通道被组成型活性G α q强烈抑制，即使是缺乏PLC激活的突变版本。在G α q/11敲除小鼠的成纤维细胞中，受体介导的TASK通道抑制需要外源性G α q表达，但在通过调节脂质磷酸酶的过表达降低PIP2水平的细胞系中，受体介导的TASK通道抑制没有减弱。直接应用活化的G α q，而不是其他G蛋白亚基，抑制了切除斑块中的TASK通道，并且组成活性的G α q亚基选择性地与TASK通道共免疫沉淀。这些数据表明，受体介导的TASK通道抑制与PIP2耗竭无关，它们提示了一种机制，即G α q通过与离子通道或密切相关的介质直接相互作用发生通道调节。
Two-pore-domain K+ channels provide neuronal background currents that establish resting membrane potential and input resistance; their modulation provides a prevalent mechanism for regulating cellular excitability. The so-called TASK channel subunits (TASK-1 and TASK-3) are widely expressed, and they are robustly inhibited by receptors that signal through G alpha q family proteins. Here, we manipulated G protein expression and membrane phosphatidylinositol 4,5-bisphosphate (PIP2) levels in intact and cell-free systems to provide electrophysiological and biochemical evidence that inhibition of TASK channels by G alpha q-linked receptors proceeds unabated in the absence of phospholipase C (PLC) activity, and instead involves association of activated G alpha q subunits with the channels. Receptor-mediated inhibition of TASK channels was faster and less sensitive to a PLC beta 1-ct minigene construct than inhibition of PIP2-sensitive Kir3.4(S143T) homomeric channels that is known to be dependent on PLC. TASK channels were strongly inhibited by constitutively active G alpha q, even by a mutated version that is deficient in PLC activation. Receptor-mediated TASK channel inhibition required exogenous G alpha q expression in fibroblasts derived from G alpha q/11 knockout mice, but proceeded unabated in a cell line in which PIP2 levels were reduced by regulated overexpression of a lipid phosphatase. Direct application of activated G alpha q, but not other G protein subunits, inhibited TASK channels in excised patches, and constitutively active G alpha q subunits were selectively coimmunoprecipitated with TASK channels. These data indicate that receptor-mediated TASK channel inhibition is independent Of PIP2 depletion, and they suggest a mechanism whereby channel modulation by G alpha q occurs through direct interaction with the ion channel or a closely associated intermediary.