长QT综合征相关基因新突变G52R-KCNE1的功能研究

Electrophysiological characterization of a novel long QT syndrome mutation G52R-KCNE1

目的 了解长QT综合征相关基因新突变G52R-KCNE1的功能。方法 采用交叠PCR方法体外制备突变体G52R-KCNE1,并克隆到原核细胞表达载体中;体外合成RNA、显微注射入卵母细胞,在卵母细胞中共表达野生型KCNQ1(WT-KCNQ1)、野生型KCNE1(WT-KCNE1)和突变体G52R-KCNE1,采用标准双电极电压钳了解突变体的功能。结果 突变体G52R-KCNE1不能放大WT-KCNQ1通道的电流强度;当WT-KCNE1和G52R-KCNE1等量注射时,G52R-KCNE1可降低WT-KCNQ1／WT-KCNE1通道约50％的电流强度,而不影响该通道的激活动力学。结论 位于跨膜区第52位的甘氨酸对维持KCNE1的功能非常重要;突变体G52R-KCNE1明显抑制WT-KCNE1通道的功能,这可能是突变基因携带者心肌细胞复极化延缓,QT间期延长的分子电生理机制。

Objective To identify the electrophysiological properties of a novel long QT syndrome mutation G52R-KCNE1 in vitro. Methods Mutations was constructed by overlap PCR and cRNA were produced by T7 RNA polymerase. The electrophysiological properties of the mutation were studied in the Xenopus oocyte heterologous expression system. Results Compared with wild-type KCNE1/KCNQ1 channels, coexpression of G52R-KCNE1 with KCNQ1 in Xenopus oocytes did not amplify the KCNQ1 current amplitudes. Coexpression of KCNQ1 together with wild type KCNE1 and G52R-KCNE1 reduced the wild-type Iks current amplitude by 50% , whereas other biophysical properties of the Iks were not altered. Conclusions Our findings indicate that Glycine52 in the transmembrane domain is critical for KCNE1 function. The mutant G52R-KCNE1 has a dominant negative effect on Iks current, which may lead to a prolongation of the cardiac action potential and contribute to ventricular arrhythmias and sudden death in those patients.