title: HERG-F463L potassium channels linked to long QT syndrome reduce I(Kr) current by a trafficking-deficient mechanism.
abstract: 1. Congenital long QT syndrome (LQTS) is a genetically heterogeneous disease. The aim of the present study was to identify the gene mutation in a Chinese family with LQTS and investigate the functional changes associated with the mutation. 2. Polymerase chain reaction and DNA sequencing were used to screen for the KCNH2 mutation in the proband. A mutant F463L HERG channel was expressed in HEK293 cells using a lipofectamine method. The IKr current was recorded using the whole-cell voltage clamp technique. Expression of HERG protein was detected by western blotting and the subcellular location of HERG channels in cell was analysed by confocal microscopy. 3. The novel heterozygous missense mutation F463L in KCNH2 was detected. We found that the F463L mutation did not lead to any expression of detectable I(Kr) current, which was consistent with western blotting analysis indicating that the F463L mutation only expressed a band at 135 kDa. When coexpressed with wild-type HERG, F463L HERG exhibited strong dominant-negative current suppression, resulting in a decrease in I(Kr) current density, and induced a positive shift in the voltage dependence of activation, as well as interference with trafficking of wild-type channel protein. The processing of the F463L channels was partly corrected in cells incubated in E4031. In addition, confocal microscopy demonstrated that F463L subunits could be inserted into the cell membrane when forming heteromultimeric channels with wild-type channel subunits. 4. The results of the present study suggest that the F463L mutation leads to loss of function in HERG through a dominant-negative effect caused by impaired trafficking of the channel.
authors.affiliation: Department of Cardiology, First Affiliated Hospital, Ion Channel Disease Laboratory, Key Laboratory of Environment and Genes Related to Diseases (Xi'an Jiaotong University), Ministry of Education, Medical College of Xi'an Jiaotong University, Xi'an, Shaanxi, PR China.
authors.forename: Hai-Tao
authors.initials: HT
authors.lastname: Yang
authors.forename: Chao-Feng
authors.initials: CF
authors.lastname: Sun
authors.forename: Chang-Cong
authors.initials: CC
authors.lastname: Cui
authors.forename: Xiao-Lin
authors.initials: XL
authors.lastname: Xue
authors.forename: Ai-Feng
authors.initials: AF
authors.lastname: Zhang
authors.forename: Hong-Bing
authors.initials: HB
authors.lastname: Li
authors.forename: Dong-Qi
authors.initials: DQ
authors.lastname: Wang
authors.forename: Juan
authors.initials: J
authors.lastname: Shu
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001483:Base Sequence; D015153:Blotting, Western; D002460:Cell Line; D003596:Cytosine / Q000378:metabolism; D000072237:ERG1 Potassium Channel; D004562:Electrocardiography; D004721:Endoplasmic Reticulum / Q000235:genetics / Q000378:metabolism; D051638:Ether-A-Go-Go Potassium Channels / Q000235:genetics* / Q000378:metabolism; D006579:Heterozygote; D006801:Humans; D007930:Leucine / Q000235:genetics*; D008133:Long QT Syndrome / Q000151:congenital / Q000235:genetics* / Q000378:metabolism; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D020125:Mutation, Missense; D018408:Patch-Clamp Techniques; D010375:Pedigree; D010649:Phenylalanine / Q000235:genetics*; D024642:Potassium Channels, Voltage-Gated / Q000378:metabolism*; D021122:Protein Subunits; D021381:Protein Transport / Q000235:genetics; D013941:Thymine / Q000378:metabolism; D014162:Transfection
chemical_list: D000072237:ERG1 Potassium Channel; D051638:Ether-A-Go-Go Potassium Channels; C000606913:KCNH2 protein, human; D024642:Potassium Channels, Voltage-Gated; D021122:Protein Subunits; C059983:potassium channel protein I(sk); D010649:Phenylalanine; D003596:Cytosine; D007930:Leucine; D013941:Thymine