The glycine hinge of transmembrane segment 2 modulates the subcellular localization and gating properties in TREK channels
The glycine hinge of transmembrane segment 2 modulates the subcellular localization and gating properties in TREK channels
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跨膜片段 2 的甘氨酸铰链调节 TREK 通道中的亚细胞定位和门控特性
DOI：
10.1016/j.bbrc.2017.06.200
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发表时间：
2017
影响因子：
3.1
通讯作者：
Ma Xiao-Yun
中科院分区：
文献类型：
--
作者：
Zhuo Ren-Gong;Peng Peng;Zheng Jian-Quan;Zhang Yun-Long;Wen Lei;Wei Xiao-Li;Ma Xiao-Yun
TWIK-Related K+channels（TREK），包括TREK-1和TREK-2，属于双孔结构域K+（K2 P）家族的TREK/TRAAK亚类。跨膜片段4（M4）-甘氨酸铰链在TREK通道门控中的重要功能已被表征，但M2-铰链（M4-铰链的等效残基）的作用尚不清楚。在这里，通过表征其M2铰链突变体（G166 A的TREK-1和G196 A的TREK-2）的宏观电流、亚细胞定位和门控特性，我们研究了M2铰链的功能。G166 A显示降低的全细胞电流，而G196 A不产生电流。亚细胞分析表明，这两个突变体聚集在核周区附近，其中大部分滞留在内质网（ER）。接下来，为了探索M2铰链在门控机制中的作用，我们测试了相关M2铰链突变体对2-氨基乙氧基二苯基硼酸酯（2-APB）和胞外pH改变（ΔpHo）的响应。与TREK-1 mut 7相比，TREK-1 mut 7-G166 A对2-APB激活的敏感性降低，但对ΔpHo的敏感性相似。WT-ΔpCt是一种TREK-2串联二聚体，用于评估M2铰链在TREK-2的膜型门控中的功能。G196 A-ΔpCt对2-APB和Δ pHo的敏感性较WT-ΔpCt降低。总之，我们的研究结果表明，TREK通道的M2铰链控制其宏观电流，亚细胞定位和门控过程。
TWIK-Related K+channels (TREK), including TREK-1 and TREK-2, belong to the TREK/TRAAK subclass of two-pore domain K+(K2P) family. The important functions of transmembrane segment 4 (M4)-glycine hinge in TREK channel gating have been characterized, but the roles of M2-hinge (the equivalent residue of M4-hinge) remain unclear. Here, by characterizing the macroscopic currents, subcellular localization and gating properties of their M2-hinge mutants (G166A for TREK-1 and G196A for TREK-2), we investigated the functions of M2-hinge. G166A displayed decreased whole-cell currents, whereas no current was produced by G196A. Subcellular analysis indicated that both mutants were aggregated near the perinuclear region, and most of them were retented within the endoplasmic reticulum (ER). Next, to explore the roles of M2-hinge in the gating mechanism, we tested the responses of the related M2-hinge mutants to 2-Aminoethoxydiphenyl borate (2-APB) and extracellular pH alteration (ΔpHo). TREK-1mut7-G166A displayed reduced sensitivity to 2-APB activation, but similar sensitivity to ΔpHo, when compared with TREK-1mut7. WT-ΔpCt, a TREK-2 tandom dimer, was used to assess the function of M2-hinge in thecis-type gating of TREK-2. The sensitivities of G196A-ΔpCt to both 2-APB and ΔpHodecreased compared with WT-ΔpCt. Taken together, our results reveal that the M2-hinge of TREK channels control their macroscopic current, subcellular localization and gating process.