Acid-sensing ion channel (ASIC) 1a/2a heteromers have a flexible 2:1/1:2 stoichiometry
Acid-sensing ion channel (ASIC) 1a/2a heteromers have a flexible 2:1/1:2 stoichiometry
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DOI：
10.1073/pnas.1324060111
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发表时间：
2014-06-03
影响因子：
11.1
通讯作者：
Ulbrich, Maximilian H.
中科院分区：
文献类型：
--
作者：
Bartoi, Tudor;Augustinowski, Katrin;Ulbrich, Maximilian H.
酸敏感离子通道 (ASIC) 是广泛表达的质子化 Na+ 通道，在组织酸中毒和疼痛中发挥作用。结晶表明了 ASIC 的三聚体组成。在非洲爪蟾卵母细胞中共表达 ASIC1a 和 ASIC2a 时，我们通过电生理学观察到异聚体的形成及其与同聚体的共存，但无法确定异聚复合物是否具有固定的亚基化学计量或某些化学计量是否优于其他化学计量。因此，我们对在单分子水平上标记有绿色和红色荧光蛋白的 ASIC 进行成像，计算 GFP 的漂白步骤，并将它们与许多单独复合物的红色串联四聚体 mCherry 共定位。组合分析提出了 ASIC1a 和 ASIC2a 亚基随机混合的模型，以产生 2:1 和 1:2 ASIC1a:ASIC2a 异聚体以及 ASIC1a 和 ASIC2a 同聚体。
Acid-sensing ion channels (ASICs) are widely expressed protongated Na+ channels playing a role in tissue acidosis and pain. A trimeric composition of ASICs has been suggested by crystallization. Upon coexpression of ASIC1a and ASIC2a in Xenopus oocytes, we observed the formation of heteromers and their coexistence with homomers by electrophysiology, but could not determine whether heteromeric complexes have a fixed subunit stoichiometry or whether certain stoichiometries are preferred over others. We therefore imaged ASICs labeled with green and red fluorescent proteins on a single-molecule level, counted bleaching steps from GFP and colocalized them with red tandem tetrameric mCherry for many individual complexes. Combinatorial analysis suggests a model of random mixing of ASIC1a and ASIC2a subunits to yield both 2:1 and 1:2 ASIC1a: ASIC2a heteromers together with ASIC1a and ASIC2a homomers.