title: Cryptic 5' splice site activation in SCN5A associated with Brugada syndrome.
abstract: The Brugada syndrome (BS) is characterized by ST segment elevation in the right precordial leads and sudden cardiac death. The disease is linked to mutations in SCN5A in approximately 20% of cases. We collected a large family with BS and have identified a novel intronic mutation. We performed the clinical, genetic, molecular and biophysical characterization of this disease-causing mutation. With direct sequencing we identified an intronic insertion of TGGG 5 bp from the end of the Exon 27 of SCN5A. For transcript analysis, we investigated Epstein-Barr-transformed lymphoblastoid cell lines from patients and controls. Total RNA was extracted and RT-PCR experiments were performed to analyze the splicing patterns in exon 27 and 28. We identified two bands, one of the expected size and the other which showed a 96 bp deletion in exon 27, leading to a 32 amino acid in-frame deletion involving segments 2 and 3 of Domain IV of the SCN5A protein. This finding indicates that the intronic mutation creates a cryptic splice site inside Exon 27. Biophysical analysis using whole-cell patch-clamp techniques showed a complete loss of function of the mutated channels when heterologously expressed. In summary, this is the first report of a dysfunctional sodium channel created by an intronic mutation giving rise to cryptic splice site activation in SCN5A in a family with the BS. The deletion of fragments of segments 2 and 3 of Domain IV leads to complete loss of function, consistent with the biophysical data found in several mutations causing BS.
authors.affiliation: Molecular Genetics, Masonic Medical Research Laboratory, 2150 Bleecker Street, Utica, NY 13501, USA.
authors.forename: Kui
authors.initials: K
authors.lastname: Hong
authors.forename: Alejandra
authors.initials: A
authors.lastname: Guerchicoff
authors.forename: Guido D
authors.initials: GD
authors.lastname: Pollevick
authors.forename: Antonio
authors.initials: A
authors.lastname: Oliva
authors.forename: Robert
authors.initials: R
authors.lastname: Dumaine
authors.forename: Mark
authors.initials: M
authors.lastname: de Zutter
authors.forename: Elena
authors.initials: E
authors.lastname: Burashnikov
authors.forename: Yue Sheng
authors.initials: YS
authors.lastname: Wu
authors.forename: Josep
authors.initials: J
authors.lastname: Brugada
authors.forename: Pedro
authors.initials: P
authors.lastname: Brugada
authors.forename: Ramon
authors.initials: R
authors.lastname: Brugada
mesh_terms: D001145:Arrhythmias, Cardiac / Q000235:genetics*; D001483:Base Sequence; D016757:Death, Sudden, Cardiac / Q000209:etiology*; D005091:Exons / Q000235:genetics; D005260:Female; D020022:Genetic Predisposition to Disease; D006801:Humans; D007438:Introns / Q000235:genetics; D008297:Male; D008969:Molecular Sequence Data; D009154:Mutation; D062554:NAV1.5 Voltage-Gated Sodium Channel; D018408:Patch-Clamp Techniques; D022821:RNA Splice Sites / Q000235:genetics*; D015222:Sodium Channels / Q000235:genetics*; D013577:Syndrome
chemical_list: D062554:NAV1.5 Voltage-Gated Sodium Channel; D022821:RNA Splice Sites; C568320:SCN5A protein, human; D015222:Sodium Channels