title: RNA interference targeting E637K mutation rescues hERG channel currents and restores its kinetic properties.
abstract: BACKGROUND
  Long QT syndrome (LQTS) is a monogenic proarrhythmic disorder that predisposes affected individuals to sudden death from tachyarrhythmia. As an inherited disease, LQTS cannot be completely cured by conventional treatment modalities. Individualized gene therapy is a promising therapeutic approach.
  
  
  OBJECTIVE
  The purpose of this study was to investigate the role of small interference RNA (siRNA) on expression of E637K-hERG (human ether-a-go-go-related gene) mutant and whether it can be used to rescue the mutant's dominant-negative suppressive effects on hERG protein channel function.
  
  
  METHODS
  Western blot was performed to select the most sensitive siRNAs to target E637K-hERG mutant knockdown. Confocal laser scanning microscope was performed to monitor cellular localization of wild-type (WT)-hERG and E637K-hERG with or without siRNA. Patch-clamp technique was used to assess the effect of siRNA on the electrophysiologic characteristics of the rapidly activating delayed rectifier K(+) current I(Kr) of the hERG protein channel.
  
  
  RESULTS
  siRNA led to a significant decrease in the level of E637K-hERG protein but did not affect the level of WT-hERG protein. WT-hERG localization in cells coexpressing E637K-hERG mutant was restored to the membrane by siRNA. The siRNA-mediated inhibition of E637K-hERG mutant restored the maximum current and tail current amplitudes. Furthermore, siRNA treatment rescued the kinetic properties of WT/E637K-hERG protein channel to a level comparable to that of WT-hERG protein channel.
  
  
  CONCLUSIONS
  Our findings illustrated that siRNA can effectively inhibit E637K-hERG protein expression and rescue the dominant-negative effect of this mutation by restoring the kinetic properties of hERG protein channel. It has potential clinical implications with regard to the possibility of using siRNA in the treatment of LQTS.
authors.affiliation: LiHuiLi Hospital, Medical School of NingBo University, NingBo, China.
authors.forename: Xiaoli
authors.initials: X
authors.lastname: Lu
authors.forename: Xi
authors.initials: X
authors.lastname: Yang
authors.forename: Xiaoyan
authors.initials: X
authors.lastname: Huang
authors.forename: Chen
authors.initials: C
authors.lastname: Huang
authors.forename: Huan Huan
authors.initials: HH
authors.lastname: Sun
authors.forename: Lihua
authors.initials: L
authors.lastname: Jin
authors.forename: Weifeng
authors.initials: W
authors.lastname: Xu
authors.forename: Haiyan
authors.initials: H
authors.lastname: Mao
authors.forename: Junming
authors.initials: J
authors.lastname: Guo
authors.forename: Jianqing
authors.initials: J
authors.lastname: Zhou
authors.forename: Jiangfang
authors.initials: J
authors.lastname: Lian
mesh_terms: D015153:Blotting, Western; D049109:Cell Proliferation; D000072237:ERG1 Potassium Channel; D051638:Ether-A-Go-Go Potassium Channels / Q000235:genetics*; D015316:Genetic Therapy / Q000379:methods; D006801:Humans; D008133:Long QT Syndrome / Q000235:genetics*; D018613:Microscopy, Confocal; D009154:Mutation; D018408:Patch-Clamp Techniques; D016133:Polymerase Chain Reaction; D034741:RNA, Small Interfering / Q000235:genetics* / Q000627:therapeutic use*
chemical_list: D000072237:ERG1 Potassium Channel; D051638:Ether-A-Go-Go Potassium Channels; C000606913:KCNH2 protein, human; D034741:RNA, Small Interfering