Conserved Single Residue in the BK Potassium Channel Required for Activation by Alcohol and Intoxication in C. elegans
Conserved Single Residue in the BK Potassium Channel Required for Activation by Alcohol and Intoxication in C. elegans
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DOI：
10.1523/jneurosci.0838-14.2014
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发表时间：
2014-07-16
影响因子：
5.3
通讯作者：
Pierce-Shimomura, Jonathan T.
中科院分区：
文献类型：
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作者：
Davis, Scott J.;Scott, Luisa L.;Pierce-Shimomura, Jonathan T.
酒精直接调节BK钾通道，改变从无脊椎动物到人类的物种行为。在线虫秀丽隐杆线虫中，消除BK通道SLO-1的突变对乙醇中毒具有显著的抵抗力。我们假设某些保守的氨基酸对乙醇调节是关键的，但对基础通道功能不是。为了鉴定这些残基，我们筛选了C。在SLO-1通道中具有不同错义突变的线虫菌株。在RCK 1结构域中具有SLO-1错义突变T381 I的菌株对中毒具有高度抗性。这种突变不干扰其他BK通道依赖性行为，表明突变通道保留了正常的体内功能。野生型蠕虫或人类BK通道的敲入挽救了在β-1-无效突变背景下的中毒和其他BK通道依赖性行为。相反，敲入蠕虫T381 I或等效的人T352 I突变体BK通道选择性地拯救BK通道依赖性行为，同时传递对中毒的抗性。单通道膜片钳记录证实，人类BK通道工程与T352 I错义突变是不敏感的激活乙醇，但在其他方面有正常的电导，钾的选择性，只有微妙的差异，电压依赖性。总之，我们的行为和电生理学结果表明，T352 I突变选择性地破坏BK通道的乙醇调制。T352 I突变可能会改变乙醇的结合位点和/或干扰乙醇诱导的构象变化，这对乙醇的行为反应至关重要。
Alcohol directly modulates the BK potassium channel to alter behaviors in species ranging from invertebrates to humans. In the nematode Caenorhabditis elegans, mutations that eliminate the BK channel, SLO-1, convey dramatic resistance to intoxication by ethanol. We hypothesized that certain conserved amino acids are critical for ethanol modulation, but not for basal channel function. To identify such residues, we screened C. elegans strains with different missense mutations in the SLO-1 channel. A strain with the SLO-1 missense mutation T381I in the RCK1 domain was highly resistant to intoxication. This mutation did not interfere with other BK channel-dependent behaviors, suggesting that the mutant channel retained normal in vivo function. Knock-in of wild-type versions of the worm or human BK channel rescued intoxication and other BK channel-dependent behaviors in a slo-1-null mutant background. In contrast, knock-in of the worm T381I or equivalent human T352I mutant BK channel selectively rescued BK channel-dependent behaviors while conveying resistance to intoxication. Single-channel patch-clamp recordings confirmed that the human BK channel engineered with the T352I missense mutation was insensitive to activation by ethanol, but otherwise had normal conductance, potassium selectivity, and only subtle differences in voltage dependence. Together, our behavioral and electrophysiological results demonstrate that the T352I mutation selectively disrupts ethanol modulation of the BK channel. The T352I mutation may alter a binding site for ethanol and/or interfere with ethanol-induced conformational changes that are critical for behavioral responses to ethanol.