Molecular Mechanism of QT Prolongation due to dysfunction of HERG K^+ Channels

HERG编码Ikr电流，这是心脏复极的重要因素，HERG基因缺陷导致一种遗传性长QT综合征（LQT2）。HERG/Ikr电流是引起药物性长QT综合征的各种心脏和非心脏药物的靶点。我们研究了在LQT2家族中发现的基因缺陷中HERG电流抑制的分子机制以及已知阻断Ikr电流的因子。采用异源表达系统分析了非洲爪蟾卵母细胞中herg突变T474I、A614V和V630L的功能异常。突变体单独不表达电流，但与野生型共注射，各突变体均产生显性负抑制（DNS），其程度依次为V63OL>A614V>T474I。V60L和A614V在稳态失活曲线上进一步产生负移，并发生固定失活。接下来，我们分析了S4的R534C突变，假定是电压传感器。R534C转移电压依赖激活，确认S4作为电压传感器，并产生快速失活。但是，快速失活不能解释R534C QT延长的原因，提示存在另一个未知因素。HERG c端突变S818L单独突变时不表达电流，与野生型共注射时不表现出DNS。然而，野生型和过量的S818L共注射会产生DNS和激活曲线移位，激活和失活动力学快速。结果表明，S818L可与野生型形成异聚体，生成具有野生型的功能通道，并且HERG的c端可能参与了该通道的激活过程。因此，不同类型和位置的HERG突变以不同的机制引起电流抑制。我们还探讨了酸中毒诱导的HERG电流的抑制主要是由于对激活过程的影响。

HERG encodes Ikr current, an important factor for cardiac repolarization and gene defects in HERG cause one form of inherited long QT syndrome (LQT2). HERG/Ikr currents are the targets of various cardiac and non-cardiac drugs that causes drug-induced long QT syndrome. We studied molecular mechanism of HERG current suppression in gene defects found in LQT2 families and by factors known to block Ikr currents. Functional abnormality ofHERG mutations in T474I, A614V and V630L were analyzed using heterologous expression system in Xenopus oocytes. The mutant alone could not express current, but co-injection with wild type and each mutant produced dominant negative suppression (DNS) with its degree increasing in the order of V63OL>A614V>T474I.V60L and A614V further produced negative shift in steady state inactivation curve and fastened inactivation. We next analyzed R534C mutation in S4, presumed voltage sensor. R534C shifted voltage-dependent activation confirming S4 playing as voltage sensor, and produced fast deactivation. But, fast deactivation could not explain the reason for QT prolongation in R534C, suggesting the presence of another unknown factor. The mutation in HERG C-terminus, S818L, did not express the current by mutant alone and co-injection with wild type did not show DNS.However co-injection of wild type and excess amount of cRNA of S818L produced DNS and shifted activation curve with fast activation and deactivation kinetics. The results suggest that S818L can form heteromultimer with wild type to yield functional channels with wild type, and that the C-terminus of HERG may participate in activation process of this channels. Therefore, various types and locations of HERG mutaions cause current suppression with different mechanisms. We also explored acidosis-induced suppression of HERG current due mainly to effects on activation process.