Organization and regulation of small conductance Ca2+-activated K+ channel multiprotein complexes
Organization and regulation of small conductance Ca2+-activated K+ channel multiprotein complexes
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DOI：
10.1523/jneurosci.3565-06.2007
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发表时间：
2007-02-28
影响因子：
5.3
通讯作者：
Adelman, John P.
中科院分区：
文献类型：
--
作者：
Allen, Duane;Fakler, Bernd;Adelman, John P.
小电导Ca 2+激活的K+通道（SK通道）是由四个α孔形成亚基组成的复合物，每个亚基与介导Ca 2+门控的钙调蛋白（CaM）结合。蛋白质组学分析表明，SK 2通道也结合蛋白激酶CK 2（CK 2）和蛋白磷酸酶2A（PP 2A）。SK2与CaM磷酸化替代物CaM（T80 D）的共表达表明，SK2结合的CaM的CK 2磷酸化降低了SK2通道的表观Ca 2+敏感性。通过使用CK 2特异性抑制剂4，5，6，7-四溴-2-氮杂苯并咪唑，我们证实了SK2通道与CK 2共组装。PP 2A还结合SK2通道并抵消CK 2的作用，如显性负突变PP 2A以及不再能够结合PP 2A的突变SK2通道的共表达所示。体外结合研究揭示了通道亚基的N和C末端之间的相互作用以及CK 2 α和β亚基、PP 2A和通道不同结构域之间的相互作用。在通道复合物中，通道N端结构域内的赖氨酸残基121激活SK 2结合的CK 2，并且CaM的磷酸化是状态依赖性的，仅在通道关闭时发生。CK 2和PP 2A的作用表明，天然SK2通道是多蛋白复合物，含有组成性相关的钙调素，CK 2的两个亚基，和至少两个不同的亚基PP 2A。结果还表明，SK2通道的Ca 2+敏感性是以动态的方式调节的，直接通过CK 2和PP 2A，并通过CK 2磷酸化CaM的状态依赖性间接由Ca 2+本身。
Small conductance Ca2+-activated K+ channels (SK channels) are complexes of four alpha pore-forming subunits each bound by calmodulin (CaM) that mediate Ca2+ gating. Proteomic analysis indicated that SK2 channels also bind protein kinase CK2 ( CK2) and protein phosphatase 2A (PP2A). Coexpression of SK2 with the CaM phosphorylation surrogate CaM(T80D) suggested that the apparent Ca2+ sensitivity of SK2 channels is reduced by CK2 phosphorylation of SK2-bound CaM. By using 4,5,6,7-tetrabromo-2- azabenzimidazole, a CK2-specific inhibitor, we confirmed that SK2 channels coassemble with CK2. PP2A also binds to SK2 channels and counterbalances the effects of CK2, as shown by coexpression of a dominant-negative mutant PP2A as well as a mutant SK2 channel no longer able to bind PP2A. In vitro binding studies have revealed interactions between the N and C termini of the channel subunits as well as interactions among CK2 alpha and beta subunits, PP2A, and distinct domains of the channel. In the channel complex, lysine residue 121 within the N-terminal domain of the channel activates SK2-bound CK2, and phosphorylation of CaM is state dependent, occurring only when the channels are closed. The effects of CK2 and PP2A indicate that native SK2 channels are multiprotein complexes that contain constitutively associated CaM, both subunits of CK2, and at least two different subunits of PP2A. The results also show that the Ca2+ sensitivity of SK2 channels is regulated in a dynamic manner, directly through CK2 and PP2A, and indirectly by Ca2+ itself via the state dependence of CaM phosphorylation by CK2.