Novel intracellular transport-refractory mutations in KCNH2 identified in patients with symptomatic long QT syndrome

作者：Fukumoto Daisuke; Ding Wei Guang; Wada Yuko; Fujii Yusuke; Ichikawa Mari; Takayama Koichiro; Fukuyama Megumi; Kato Koichi; Itoh Hideki; Makiyama Takeru; Omatsu Kanbe Mariko; Matsuura Hiroshi; Hone Minoru; Ohno Seiko^*

来源：Journal of Cardiology, 2018, 71(3-4): 401-408.

DOI：10.1016/j.jjcc.2017.10.004

• Long QT syndrome
• KCNH2
• Mutation
• Patch-clamp
• Immunaluorescence

Background: Missense mutations in KCNH2, a gene encoding the Kv11.1 channel, cause long QT syndrome (LQTS) type 2 primarily by disrupting the intracellular transport of Kv11.1 to the plasma membrane. The present study aimed to clarify the functional changes by two novel KCNH2 missense mutations.
Methods: We performed genetic screening of three unrelated symptomatic LQTS probands with family histories of cardiac symptoms. Chinese hamster ovary cells were transfected with wild-type (WT) and/or mutant KCNH2 plasmid and examined by patch-clamp technique. Immunostaining and confocal microscopy were performed to evaluate the intracellular localization of WT and homozygous mutant Kv11.1 in human embryonic kidney cells. For the study of trafficking rescue, we used low-temperature incubation (30 CC). We also examined pharmacological rescue of homozygous mutant Kv11.1 current in cells treated with E-4031 or dofetilide.
Results: We identified two novel KCNH2 missense mutations, G785D and T8261. Electrophysiological study showed that both mutant channels were nonfunctional in homozygous condition and reduced current densities by half in heterozygous condition compared with WT Kv11.1. Heterozygous Kv11.1-G785D produced a significant positive shift in activation and a significant negative shift in inactivation, whereas heterozygous Kv11.1-T8261 caused no kinetic changes. lmmunostaining revealed that both were transport-refractory mutations. Incubation at 30 degrees C rescued plasma membrane expression of Kv11.1-T8261 but not G785D. We confirmed low-temperature-induced restoration of homozygous Kv11.1-T8261 transport by functional current measurements. In contrast, incubation with E-4031 or dofetilide failed to produce measurable currents in both homozygous mutant channels.
Conclusions: Two novel KCNH2 mutations disrupted the intracellular transport of Kv11.1. Low-temperature incubation rescued plasma membrane expression of Kv11.1-T8261 but not G785D. Both mutations exerted loss-of-function effects on Kv11.1 and explained the phenotypes of the mutation carriers.

• 出版日期2018-4