Mechanistic basis for the pathogenesis of long QT syndrome associated with a common splicing mutation in KCNQ1 gene
Mechanistic basis for the pathogenesis of long QT syndrome associated with a common splicing mutation in KCNQ1 gene
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DOI：
10.1016/j.yjmcc.2006.12.015
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发表时间：
2007-03-01
影响因子：
5
通讯作者：
Horie, Minoru
中科院分区：
文献类型：
--
作者：
Tsuji, Keiko;Akao, Masaharu;Horie, Minoru
编码心肌延迟整流钾通道基因KCNQ1的突变可导致长QT综合征(LQTS)。我们研究了3个LQTS家系，在这些家系中，发现了先前报道的常见剪接点突变，即外显子7(c.1032G>；A)最后一个碱基的鸟嘌呤转变为腺嘌呤。我们使用实时逆转录聚合酶链式反应对该突变引起的KCNQ1mRNAs的外显子跳跃进行了定量测量。与具有少量剪接变异体的正常人相比(Delta 7-8：0.1%，Delta 8：6.9%，占KCNQ1转录本总数的6.9%)，患者外显子跳跃mRNAs显著增加(Delta 7：23.5%，Delta 7-8：16.8%，Delta 8：4.5%)。非洲爪哇卵母细胞异源表达野生型(WT)通道或跳过外显子的KCNQ1通道(Delta 7、Delta 7-8或Delta 8)的电流记录表明，这些突变体都没有产生任何可测得的电流，而且当它们与野生型(WT)共表达时，它们对WT电流表现出突变体特有的显性-负效应。共聚焦显微镜分析表明，荧光蛋白标记的WT主要表达在质膜上，而突变体则分布在细胞内。当WT与突变体共表达时，大部分WT与突变体共定位于细胞内。最后，我们提供了证据表明WT和突变体之间直接的蛋白质-蛋白质相互作用，通过荧光共振能量转移。因此，突变体可能通过将WT捕获到细胞内，从而干扰其向质膜的转运，从而发挥其显性-负效应。总之，我们的数据为KCNQ1剪接突变引起的LQTS的发病机制提供了基础。(C)2007 Elsevier Inc.保留所有权利。
Mutations in KCNQ1, the gene encoding the delayed rectifier K+ channel in cardiac muscle, cause long QT syndrome (LQTS). We studied 3 families with LQTS, in whom a guanine to adenine change in the last base of exon 7 (c. 1032G >A), previously reported as a common splice-site mutation, was identified. We performed quantitative measurements of exon-skipping KCNQ1 mRNAs caused by this mutation using real-time reverse transcription polymerase chain reaction. Compared with normal individuals who have minor fractions of splicing variants (Delta 7-8: 0.1%, Delta 8: 6.9%, of total KCNQ1 transcripts), the affected individuals showed remarkable increases of exon-skipping mRNAs (Delta 7: 23.5%, Delta 7-8: 16.8%, Delta 8: 4.5%). Current recordings from Xenopus laevis oocytes heterologously expressing channels of wild-type (WT) or exon-skipping KCNQ1 (Delta 7, Delta 7-8, or Delta 8) revealed that none of the mutants produced any measurable currents, and moreover they displayed mutant-specific degree of dominant-negative effects on WT currents, when co-expressed with WT. Confocal microscopy analysis showed that fluorescent protein-tagged WT was predominantly expressed on the plasma membrane, whereas the mutants showed intracellular distribution. When WT was co-expressed with mutants, the majority of WT co-localized with the mutants in the intracellular space. Finally, we provide evidence showing direct protein-protein interactions between WT and the mutants, by using fluorescence resonance energy transfer. Thus, the mutants may exert their dominant-negative effects by trapping WT intracellularly and thereby interfering its translocation to the plasma membrane. In conclusion, our data provide a mechanistic basis for the pathogenesis of LQTS caused by a splicing mutation in KCNQ1. (c) 2007 Elsevier Inc. All rights reserved.