Maljevic Snezana, Lerche Christian, Seebohm Guiscard, Alekov Alexi K, Busch Andreas E, Lerche Holger
Department of Applied Physiology, University of Ulm, Germany.
J Physiol. 2003 Apr 15;548(Pt 2):353-60. doi: 10.1113/jphysiol.2003.040980. Epub 2003 Mar 14.
Coexpression of KCNQ2 and KCNQ3 channels results in a 10-fold increased current amplitude compared to that of KCNQ2 alone, suggesting the formation of heteromultimeric channels. There is no interaction of either channel with KCNQ1. We evaluated the C-terminus as a potential interaction domain by construction of chimeras with interchanged C-termini of KCNQ1, KCNQ2 and KCNQ3 and functional expression in Xenopus oocytes. The chimera of KCNQ1 with a KCNQ2 C-terminus (Q1ctQ2) showed an 8-fold increase in current amplitude, and Q1ctQ3 a 3-fold increase when coexpressed with KCNQ3 and KCNQ2, respectively, indicating that the C-terminus contains an interaction domain. To characterize this interacting region, we studied further chimeras of KCNQ1 containing different parts of the KCNQ3 C-terminus for interaction with KCNQ2. We also evaluated short sequences of the KCNQ2 C-terminus for a dominant-negative effect on Q1ctQ3. According to the results of these experiments, functional interaction of KCNQ2 and KCNQ3 requires a highly conserved region of about 80 amino acids, previously called the A-domain, plus either 40 residues downstream of the A-domain (B-domain) or the proximal C-terminus between S6 and the A-domain. Furthermore, the chimeras Q1ctQ3 and Q2ctQ3 showed > 10-fold increased current amplitudes compared to KCNQ1 or KCNQ2 alone and a strong depolarizing shift of voltage-dependent activation. The proximal part of the KCNQ3 C-terminus was necessary to produce these effects. Our results indicate that specific parts of the C-terminus enable the interaction between KCNQ2 and KCNQ3 channels and that different parts of the KCNQ3 C-terminus are important for regulating current amplitude.
与单独的KCNQ2相比，KCNQ2和KCNQ3通道的共表达使电流幅度增加了10倍，这表明形成了异源多聚体通道。这两种通道与KCNQ1均无相互作用。我们通过构建KCNQ1、KCNQ2和KCNQ3的C末端互换的嵌合体并在非洲爪蟾卵母细胞中进行功能表达，来评估C末端作为潜在相互作用结构域。带有KCNQ2 C末端的KCNQ1嵌合体（Q1ctQ2）在分别与KCNQ3和KCNQ2共表达时，电流幅度增加了8倍，而Q1ctQ3增加了3倍，这表明C末端包含一个相互作用结构域。为了表征这个相互作用区域，我们进一步研究了包含KCNQ3 C末端不同部分的KCNQ1嵌合体与KCNQ2的相互作用。我们还评估了KCNQ2 C末端的短序列对Q1ctQ3的显性负效应。根据这些实验结果，KCNQ2和KCNQ3的功能相互作用需要一个约80个氨基酸的高度保守区域，该区域以前称为A结构域，加上A结构域下游的40个残基（B结构域）或S6与A结构域之间的近端C末端。此外，与单独的KCNQ1或KCNQ2相比，嵌合体Q1ctQ3和Q2ctQ3的电流幅度增加了10倍以上，并且电压依赖性激活发生了强烈的去极化偏移。KCNQ3 C末端的近端部分是产生这些效应所必需的。我们的结果表明，C末端的特定部分使KCNQ2和KCNQ3通道之间能够相互作用，并且KCNQ3 C末端的不同部分对于调节电流幅度很重要。
