Heterodimerization within the TREK channel subfamily produces a diverse family of highly regulated potassium channels
Heterodimerization within the TREK channel subfamily produces a diverse family of highly regulated potassium channels
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DOI：
10.1073/pnas.1522459113
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发表时间：
2016-04-12
影响因子：
11.1
通讯作者：
Sandoz, Guillaume
中科院分区：
文献类型：
--
作者：
Levitz, Joshua;Royal, Perrine;Sandoz, Guillaume
Twik相关的K+通道1（TREK 1）、TREK 2和Twik相关的花生四烯酸刺激的K+通道（TRAAK）组成了双孔结构域K+（K-2 P）通道的TREK亚家族。尽管在神经系统中共享高达78%的序列同源性和重叠的表达谱，但这些通道在生理刺激的调节中显示出重大差异。例如，TREK 1被外部酸化抑制，而TREK 2被激活。在这里，我们研究了TREK亚家族成员组装形成具有新特性的功能性异聚体通道的能力。使用HEK细胞裂解物的单分子下拉（SiMPull）和活细胞质膜中的亚基计数，我们表明TREK 1，TREK 2和TRAAK容易共组装。TREK 1和TREK 2可以各自与TRAAK异源二聚化，但这样做的效率低于彼此。我们的功能特点的异二聚体，并发现所有的组合形式向外整流钾选择性通道，但可变的电压敏感性和pH调节。TREK 1-TREK 2异二聚体在生理外部pH下显示出低水平的活性，但与其相应的同二聚体不同，其可被酸性和碱性条件激活。基于最近的晶体结构的建模，沿着突变分析，表明TREK 1-TREK 2通道内的每个亚基通过可滴定的His独立调节。最后，TREK 1/TRAAK异源二聚体在功能上与TRAAK同源二聚体有两个关键的不同：它们被细胞内酸化和碱化激活，并受磷脂酶D2的调节。因此，异源二聚化提供了通过K-2 P通道内通道类型的扩展来使功能多样化的手段。
Twik-related K+ channel 1 (TREK1), TREK2, and Twik-related arachidonic-acid stimulated K+ channel (TRAAK) form the TREK subfamily of two-pore-domain K+ (K-2P) channels. Despite sharing up to 78% sequence homology and overlapping expression profiles in the nervous system, these channels show major differences in their regulation by physiological stimuli. For instance, TREK1 is inhibited by external acidification, whereas TREK2 is activated. Here, we investigated the ability of the members of the TREK subfamily to assemble to form functional heteromeric channels with novel properties. Using single-molecule pull-down (SiMPull) from HEK cell lysate and subunit counting in the plasma membrane of living cells, we show that TREK1, TREK2, and TRAAK readily coassemble. TREK1 and TREK2 can each heterodimerize with TRAAK, but do so less efficiently than with each other. We functionally characterized the heterodimers and found that all combinations form outwardly rectifying potassium-selective channels but with variable voltage sensitivity and pH regulation. TREK1-TREK2 heterodimers show low levels of activity at physiological external pH but, unlike their corresponding homodimers, are activated by both acidic and alkaline conditions. Modeling based on recent crystal structures, along with mutational analysis, suggests that each subunit within a TREK1-TREK2 channel is regulated independently via titratable His. Finally, TREK1/TRAAK heterodimers differ in function from TRAAK homodimers in two critical ways: they are activated by both intracellular acidification and alkalinization and are regulated by the enzyme phospholipase D2. Thus, heterodimerization provides a means for diversifying functionality through an expansion of the channel types within the K-2P channels.