Assembly of ROMK1 (Kir 1.1a) inward rectifier K+ channel subunits involves multiple interaction sites.
Assembly of ROMK1 (Kir 1.1a) inward rectifier K+ channel subunits involves multiple interaction sites.
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ROMK1 (Kir 1.1a) 内向整流 K 通道亚基的组装涉及多个相互作用位点。
DOI：
10.1016/s0006-3495(98)77892-0
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发表时间：
1998
期刊：
影响因子：
--
通讯作者：
Ho,K
中科院分区：
文献类型：
--
作者：
Koster,JC;Bentle,KA;Nichols,CG;Ho,K
ROMK 1（Kir 1.1a）通道由亚基的四聚体复合物形成，每个亚基的特征在于细胞质N-和C-末端以及位于孔形成片段侧翼的两个跨膜螺旋的核心区域。为了描述介导ROMK 1亚基组装的一般区域，我们构建了表位标记的N-末端、C-末端和跨膜片段缺失突变体。当与野生型ROMK 1亚基在异种卵母细胞中共表达时，N-末端、核心区和C-末端缺失的非功能性亚基具有显性负效应。与此相反，这些非功能性亚基与Kv 2.1（DRK 1）的共表达并没有抑制对照卵母细胞中的Kv 2.1电流。通过免疫沉淀[35 S]标记的卵母细胞膜蛋白，平行研究了表位标记的突变体和野生型ROMK 1亚基之间的相互作用。复合物含有野生型和突变体亚基，保留H5，M2，和C-末端区域的共免疫沉淀在更大程度上比复合物组成的野生型和突变体亚基与核心区域和/或C-末端缺失。目前的研究结果是一致的假设，即位于ROMK 1亚基的核心区域和胞质末端的多个相互作用位点介导同源多聚体组装。
The ROMK1 (Kir 1.1a) channel is formed by a tetrameric complex of subunits, each characterized by cytoplasmic N- and C-termini and a core region of two transmembrane helices flanking a pore-forming segment. To delineate the general regions mediating the assembly of ROMK1 subunits we constructed epitope-tagged N-terminal, C-terminal, and transmembrane segment deletion mutants. Nonfunctional subunits with N-terminal, core region, and C-terminal deletions had dominant negative effects when coexpressed with wild-type ROMK1 subunits inXenopusoocytes. In contrast, coexpression of these nonfunctional subunits with Kv 2.1 (DRK1) did not suppress Kv 2.1 currents in control oocytes. Interactions between epitope-tagged mutant and wild-type ROMK1 subunits were studied in parallel by immunoprecipitating [35S]-labeled oocyte membrane proteins. Complexes containing both wild-type and mutant subunits that retained H5, M2, and C-terminal regions were coimmunoprecipitated to a greater extent than complexes consisting of wild-type and mutant subunits with core region and/or C-terminal deletions. The present findings are consistent with the hypothesis that multiple interaction sites located in the core region and cytoplasmic termini of ROMK1 subunits mediate homomultimeric assembly.