title: [Functional analysis of a novel SCN5A mutation G1712C identified in Brugada syndrome].
abstract: OBJECTIVE
  To elucidate the molecular and electrophysiological mechanisms of Brugada syndrome through functional analysis of a novel SCN5A gene mutation G1712C.
  
  
  METHODS
  A recombinant plasmid pRc<CMV-hH1 containing the mutant human cardiac sodium channel α subunit (hH1) cDNA was constructed using in vitro PCR-based site-directed mutagenesis technique. LipofectamineTM 3000 was used to transfect the plasmid DNA into HEK293 cell line to induce stable expression of Na+channel β1-subunit, and the positive colonies were selected by screening with G418.The standard liposome method was used to transiently transfect HEK293 cells with either the wild-type or mutant Na+channel subunits (hH1 and mhH1, respectively), and the macroscopic Na+currents were recorded using whole-cell patch-clamp technique. Data acquisition and analysis, generation of voltage commands and curve fitting were accomplished with EPC-10, PatchMaster and IGOR Pro 6.0.
  
  
  RESULTS
  An HEK293 cell line that stably expressed Na+channel β1-subunit was successfully established. After transient transfection with the WT subunit, large Na+currents were recorded from the stable β1-cell line. Transient transfection with the G1712C subunit, however, did not elicit a Na+current in the cells.
  
  
  CONCLUSIONS
  Compared with normal Na+channel, the wild-type channel exhibits a similar sodium current. The characteristic kinetics of sodium channel of WT-hH1 was identical to that in normal cardiac muscle cell, and the missense mutation (G1712C) in the P-loop region of the domain IV may have caused the failure of sodium channel expression.
authors.affiliation: Department of Cardiology, Southern Medical University, Nanfang Hospital, Guangzhou 510515, China. E-mail: m15622156083@163.com.
authors.forename: Yan-Yu
authors.initials: YY
authors.lastname: Chen
authors.forename: Shen-Rong
authors.initials: SR
authors.lastname: Liu
authors.forename: Liang-Zhen
authors.initials: LZ
authors.lastname: Xie
authors.forename: Ting-Yan
authors.initials: TY
authors.lastname: Zhu
authors.forename: Yi-Zhen
authors.initials: YZ
authors.lastname: Chen
authors.forename: Xiao-Jiang
authors.initials: XJ
authors.lastname: Deng
authors.forename: Su-Rong
authors.initials: SR
authors.lastname: Meng
authors.forename: Jian
authors.initials: J
authors.lastname: Peng
mesh_terms: D053840:Brugada Syndrome / Q000235:genetics*; D005838:Genotype; D057809:HEK293 Cells; D006801:Humans; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D062554:NAV1.5 Voltage-Gated Sodium Channel / Q000235:genetics*; D018408:Patch-Clamp Techniques; D016133:Polymerase Chain Reaction; D014162:Transfection
chemical_list: D062554:NAV1.5 Voltage-Gated Sodium Channel; C568320:SCN5A protein, human