EF hands at the N-lobe of calmodulin are required for both SK channel gating and stable SK-calmodulin interaction.
EF hands at the N-lobe of calmodulin are required for both SK channel gating and stable SK-calmodulin interaction.
复制标题
DOI：
10.1085/jgp.200910295
复制
发表时间：
2009-10
期刊：
影响因子：
--
通讯作者：
Aldrich RW
中科院分区：
文献类型：
--
作者：
Li W;Halling DB;Hall AW;Aldrich RW
小电导钙活化钾（SK）通道通过组成相关钙调蛋白（CaM）响应细胞内Ca2+。先前的研究提出了CaM和SK通道之间相互作用的模块化设计。c -叶和CaM的连接体被认为调节本构结合，而n -叶结合Ca2+并打开SK通道。然而，我们发现突变CaM （E/Q）的共表达，其中n -叶只有一个功能EF手，导致SK通道活性的快速下降，这可以通过外源应用野生型（WT）而不是突变型CaM恢复。我们的研究结果表明，n叶EF手的突变破坏了CaM和SK通道亚基之间的稳定相互作用，因此当膜内部暴露于无CaM的溶液中时，突变的CaM与通道复合物分离。稳定相互作用的破坏不是直接由于Ca2+结合能力的丧失，因为SK通道和WT CaM可以在没有Ca2+的情况下稳定地相互作用。这些发现质疑了先前的结论，即n叶只有一个功能EF手的CaM可以稳定地支持SK通道的门控。它们不能用CaM和SK通道之间的模块化相互作用的当前模型来解释，它们暗示了n瓣EF残基与通道亚基结合的作用。此外，我们发现一种有效的SK通道增强剂，3-肟-6,7-二氯- 1h -吲哚-2,3-二酮（NS309），能够恢复与CaM （E/Q）的通道活性，这表明NS309稳定了CaM和SK通道之间的相互作用。在NS309存在的情况下，CaM （E/Q）可以调节SK通道的Ca2+依赖性门控，但其Ca2+亲和力明显低于WT CaM。
Small conductance calcium-activated potassium (SK) channels respond to intracellular Ca2+ via constitutively associated calmodulin (CaM). Previous studies have proposed a modular design for the interaction between CaM and SK channels. The C-lobe and the linker of CaM are thought to regulate the constitutive binding, whereas the N-lobe binds Ca2+ and gates SK channels. However, we found that coexpression of mutant CaM (E/Q) where the N-lobe has only one functional EF hand leads to rapid rundown of SK channel activity, which can be recovered with exogenously applied wild-type (WT), but not mutant, CaM. Our results suggest that the mutation at the N-lobe EF hand disrupts the stable interaction between CaM and SK channel subunits, such that mutant CaM dissociates from the channel complex when the inside of the membrane is exposed to CaM-free solution. The disruption of the stable interaction does not directly result from the loss of Ca2+-binding capacity because SK channels and WT CaM can stably interact in the absence of Ca2+. These findings question a previous conclusion that CaM where the N-lobe has only one functional EF hand can stably support the gating of SK channels. They cannot be explained by the current model of modular interaction between CaM and SK channels, and they imply a role for N-lobe EF hand residues in binding to the channel subunits. Additionally, we found that a potent enhancer for SK channels, 3-oxime-6,7-dichloro-1H-indole-2,3-dione (NS309), enables the recovery of channel activity with CaM (E/Q), suggesting that NS309 stabilizes the interaction between CaM and SK channels. CaM (E/Q) can regulate Ca2+-dependent gating of SK channels in the presence of NS309, but with a lower apparent Ca2+ affinity than WT CaM.
登录
查看更多内容
DOI：
10.1073/pnas.0812008106
发表时间：
2009-01-27
影响因子：
11.1
作者：
Li, Weiyan;Aldrich, Richard W.
通讯作者：
Aldrich, Richard W.
影响因子：
64.8
作者：
Schumacher, MA;Rivard, AF;Adelman, JP
通讯作者：
Adelman, JP
DOI：
10.1152/ajpheart.1999.276.3.h1107
发表时间：
1999-03-01
影响因子：
4.8
作者：
Doughty, JM;Plane, F;Langton, PD
通讯作者：
Langton, PD
DOI：
10.1016/0006-291x(82)90712-4
发表时间：
1982-01-01
影响因子：
3.1
作者：
GOPALAKRISHNA, R;ANDERSON, WB
通讯作者：
ANDERSON, WB
影响因子：
4.8
作者：
Pedarzani, P;Mosbacher, J;Fakler, B
通讯作者：
Fakler, B