ABSTRACT

Background : Mutations in SCN5A that decrease Na current underlie arrhythmia syndromes such as the Brugada syndrome (BrS). SCN5A in humans has two splice variants, one lacking a glutamine at position 1077 (Q1077del) and one containing Q1077. We investigated the effect of splice variant background on loss-of-function and rescue for R1512W, a mutation reported to cause BrS. Methods and results : We made the mutation in both variants and expressed them in HEK-293 cells for voltage-clamp study. After 24 hours of transfection, the current expression level of R1512W was reduced by ~50% in both Q1077del and Q1077 compared to the wild-type (WT) channel, respectively. The activation and inactivation midpoint were not different between WT and mutant channels in both splice variant backgrounds. However, slower time constants of recovery and enhanced intermediate inactivation were observed for R1512W/Q1077 compared with WT-Q1077, while the recovery and intermediate inactivation parameters of R1512W/Q1077del were similar to WT-Q1077del. Furthermore, both mexiletine and the common polymorphism H558R restored peak sodium current (I _Na) amplitude of the mutant channel by increasing the cell surface expression of SCN5A. Conclusion : These findings provide further evidence that the splice variant affects the molecular phenotype with implications for the clinical phenotype, and they provide insight into the expression defect mechanisms and potential treatment in BrS.

摘要

背景 ：SCN5A突变可降低Na电流，这是心律失常综合征（例如Brugada综合征（BrS））的基础。人类的SCN5A具有两种剪接变体，一种在1077位（Q1077del）缺少谷氨酰胺，另一种含有Q1077。我们调查了剪接变体背景对R1512W功能丧失和抢救的影响，R1512W是据报道会引起BrS的突变。方法与结果 ：我们在两个变体中都进行了突变，并在HEK-293细胞中表达它们，用于电压钳研究。转染24小时后，与野生型（WT）通道相比，R1012W在Q1077del和Q1077中的当前表达水平分别降低了约50％。在两个剪接变体背景下，WT和突变通道之间的激活和失活中点没有差异。然而，与WT-Q1077相比，R1512W / Q1077的恢复时间常数和中间灭活的增强，而R1512W / Q1077del的恢复和中间灭活参数与WT-Q1077del相似。此外，美西律汀和常见多态性H558R均恢复了钠电流峰值（I _Na通过增加SCN5A的细胞表面表达来确定突变通道的幅度。结论：这些发现提供了进一步的证据，表明剪接变体影响分子表型并影响临床表型，并且它们提供了对BrS中表达缺陷机制和潜在治疗的见解。