title: Characterization of SEMA3A-encoded semaphorin as a naturally occurring Kv4.3 protein inhibitor and its contribution to Brugada syndrome.
abstract: BACKGROUND
  Semaphorin 3A (SEMA3A)-encoded semaphorin is a chemorepellent that disrupts neural patterning in the nervous and cardiac systems. In addition, SEMA3A has an amino acid motif that is analogous to hanatoxin, an inhibitor of voltage-gated K(+) channels. SEMA3A-knockout mice exhibit an abnormal ECG pattern and are prone to ventricular arrhythmias and sudden cardiac death.
  
  
  OBJECTIVE
  Our aim was to determine whether SEMA3A is a naturally occurring protein inhibitor of Kv4.3 (Ito) channels and its potential contribution to Brugada syndrome.
  
  
  RESULTS
  Kv4.3, Nav1.5, Cav1.2, or Kv4.2 were coexpressed or perfused with SEMA3A in HEK293 cells, and electrophysiological properties were examined via whole-cell patch clamp technique. SEMA3A selectively altered Kv4.3 by significantly reducing peak current density without perturbing Kv4.3 cell surface protein expression. SEMA3A also reduced Ito current density in cardiomyocytes derived from human-induced pluripotent stem cells. Disruption of a putative toxin binding domain on Kv4.3 was used to assess physical interactions between SEMA3A and Kv4.3. These findings in combination with coimmunoprecipitations of SEMA3A and Kv4.3 revealed a potential direct binding interaction between these proteins. Comprehensive mutational analysis of SEMA3A was performed on 198 unrelated SCN5A genotype-negative patients with Brugada syndrome, and 2 rare SEMA3A missense mutations were identified. The SEMA3A mutations disrupted SEMA3A's ability to inhibit Kv4.3 channels, resulting in a significant gain of Kv4.3 current compared with wild-type SEMA3A.
  
  
  CONCLUSIONS
  This study is the first to demonstrate SEMA3A as a naturally occurring protein that selectively inhibits Kv4.3 and SEMA3A as a possible Brugada syndrome susceptibility gene through a Kv4.3 gain-of-function mechanism.
authors: From the Center for Clinical and Translational Science (N.J.B.) and Mayo Graduate School (N.J.B.), Department Molecular Pharmacology and Experimental Therapeutics, Windland Smith Rice Sudden Death Genomics Laboratory (D.Y., D.J.T., J.R.G., M.J.A.), Department of Medicine (J.R.G.), Division of Cardiovascular Diseases (M.J.A.), and Division of Pediatric Cardiology (M.J.A.), Mayo Clinic, Rochester, MN; Department of Molecular Biology and Pharmacology, Washington University School of Medicine, St Louis, MO (E.K.J., W.W., J.M.N.); Department of Molecular Medicine, University of Pavia, Pavia, Italy (L.C.); Center for Cardiac Arrhythmias of Genetic Origin, IRCCS Istituto Auxologico Italiano, Milan, Italy (L.C., F.D., M.T., P.J.S.); Institute of Human Genetics, Helmholtz Zentrum Munich, Neuherberg, Germany (L.C.); and Department of Clinical and Experimental Cardiology, Academic Medical Center, Amsterdam, The Netherlands (Y.M., M.A., A.A.M.W.).
authors: Nicole J
authors: NJ
authors: Boczek
authors: From the Center for Clinical and Translational Science (N.J.B.) and Mayo Graduate School (N.J.B.), Department Molecular Pharmacology and Experimental Therapeutics, Windland Smith Rice Sudden Death Genomics Laboratory (D.Y., D.J.T., J.R.G., M.J.A.), Department of Medicine (J.R.G.), Division of Cardiovascular Diseases (M.J.A.), and Division of Pediatric Cardiology (M.J.A.), Mayo Clinic, Rochester, MN; Department of Molecular Biology and Pharmacology, Washington University School of Medicine, St Louis, MO (E.K.J., W.W., J.M.N.); Department of Molecular Medicine, University of Pavia, Pavia, Italy (L.C.); Center for Cardiac Arrhythmias of Genetic Origin, IRCCS Istituto Auxologico Italiano, Milan, Italy (L.C., F.D., M.T., P.J.S.); Institute of Human Genetics, Helmholtz Zentrum Munich, Neuherberg, Germany (L.C.); and Department of Clinical and Experimental Cardiology, Academic Medical Center, Amsterdam, The Netherlands (Y.M., M.A., A.A.M.W.).
authors: Dan
authors: D
authors: Ye
authors: From the Center for Clinical and Translational Science (N.J.B.) and Mayo Graduate School (N.J.B.), Department Molecular Pharmacology and Experimental Therapeutics, Windland Smith Rice Sudden Death Genomics Laboratory (D.Y., D.J.T., J.R.G., M.J.A.), Department of Medicine (J.R.G.), Division of Cardiovascular Diseases (M.J.A.), and Division of Pediatric Cardiology (M.J.A.), Mayo Clinic, Rochester, MN; Department of Molecular Biology and Pharmacology, Washington University School of Medicine, St Louis, MO (E.K.J., W.W., J.M.N.); Department of Molecular Medicine, University of Pavia, Pavia, Italy (L.C.); Center for Cardiac Arrhythmias of Genetic Origin, IRCCS Istituto Auxologico Italiano, Milan, Italy (L.C., F.D., M.T., P.J.S.); Institute of Human Genetics, Helmholtz Zentrum Munich, Neuherberg, Germany (L.C.); and Department of Clinical and Experimental Cardiology, Academic Medical Center, Amsterdam, The Netherlands (Y.M., M.A., A.A.M.W.).
authors: Eric K
authors: EK
authors: Johnson
authors: From the Center for Clinical and Translational Science (N.J.B.) and Mayo Graduate School (N.J.B.), Department Molecular Pharmacology and Experimental Therapeutics, Windland Smith Rice Sudden Death Genomics Laboratory (D.Y., D.J.T., J.R.G., M.J.A.), Department of Medicine (J.R.G.), Division of Cardiovascular Diseases (M.J.A.), and Division of Pediatric Cardiology (M.J.A.), Mayo Clinic, Rochester, MN; Department of Molecular Biology and Pharmacology, Washington University School of Medicine, St Louis, MO (E.K.J., W.W., J.M.N.); Department of Molecular Medicine, University of Pavia, Pavia, Italy (L.C.); Center for Cardiac Arrhythmias of Genetic Origin, IRCCS Istituto Auxologico Italiano, Milan, Italy (L.C., F.D., M.T., P.J.S.); Institute of Human Genetics, Helmholtz Zentrum Munich, Neuherberg, Germany (L.C.); and Department of Clinical and Experimental Cardiology, Academic Medical Center, Amsterdam, The Netherlands (Y.M., M.A., A.A.M.W.).
authors: Wei
authors: W
authors: Wang
authors: From the Center for Clinical and Translational Science (N.J.B.) and Mayo Graduate School (N.J.B.), Department Molecular Pharmacology and Experimental Therapeutics, Windland Smith Rice Sudden Death Genomics Laboratory (D.Y., D.J.T., J.R.G., M.J.A.), Department of Medicine (J.R.G.), Division of Cardiovascular Diseases (M.J.A.), and Division of Pediatric Cardiology (M.J.A.), Mayo Clinic, Rochester, MN; Department of Molecular Biology and Pharmacology, Washington University School of Medicine, St Louis, MO (E.K.J., W.W., J.M.N.); Department of Molecular Medicine, University of Pavia, Pavia, Italy (L.C.); Center for Cardiac Arrhythmias of Genetic Origin, IRCCS Istituto Auxologico Italiano, Milan, Italy (L.C., F.D., M.T., P.J.S.); Institute of Human Genetics, Helmholtz Zentrum Munich, Neuherberg, Germany (L.C.); and Department of Clinical and Experimental Cardiology, Academic Medical Center, Amsterdam, The Netherlands (Y.M., M.A., A.A.M.W.).
authors: Lia
authors: L
authors: Crotti
authors: From the Center for Clinical and Translational Science (N.J.B.) and Mayo Graduate School (N.J.B.), Department Molecular Pharmacology and Experimental Therapeutics, Windland Smith Rice Sudden Death Genomics Laboratory (D.Y., D.J.T., J.R.G., M.J.A.), Department of Medicine (J.R.G.), Division of Cardiovascular Diseases (M.J.A.), and Division of Pediatric Cardiology (M.J.A.), Mayo Clinic, Rochester, MN; Department of Molecular Biology and Pharmacology, Washington University School of Medicine, St Louis, MO (E.K.J., W.W., J.M.N.); Department of Molecular Medicine, University of Pavia, Pavia, Italy (L.C.); Center for Cardiac Arrhythmias of Genetic Origin, IRCCS Istituto Auxologico Italiano, Milan, Italy (L.C., F.D., M.T., P.J.S.); Institute of Human Genetics, Helmholtz Zentrum Munich, Neuherberg, Germany (L.C.); and Department of Clinical and Experimental Cardiology, Academic Medical Center, Amsterdam, The Netherlands (Y.M., M.A., A.A.M.W.).
authors: David J
authors: DJ
authors: Tester
authors: From the Center for Clinical and Translational Science (N.J.B.) and Mayo Graduate School (N.J.B.), Department Molecular Pharmacology and Experimental Therapeutics, Windland Smith Rice Sudden Death Genomics Laboratory (D.Y., D.J.T., J.R.G., M.J.A.), Department of Medicine (J.R.G.), Division of Cardiovascular Diseases (M.J.A.), and Division of Pediatric Cardiology (M.J.A.), Mayo Clinic, Rochester, MN; Department of Molecular Biology and Pharmacology, Washington University School of Medicine, St Louis, MO (E.K.J., W.W., J.M.N.); Department of Molecular Medicine, University of Pavia, Pavia, Italy (L.C.); Center for Cardiac Arrhythmias of Genetic Origin, IRCCS Istituto Auxologico Italiano, Milan, Italy (L.C., F.D., M.T., P.J.S.); Institute of Human Genetics, Helmholtz Zentrum Munich, Neuherberg, Germany (L.C.); and Department of Clinical and Experimental Cardiology, Academic Medical Center, Amsterdam, The Netherlands (Y.M., M.A., A.A.M.W.).
authors: Federica
authors: F
authors: Dagradi
authors: From the Center for Clinical and Translational Science (N.J.B.) and Mayo Graduate School (N.J.B.), Department Molecular Pharmacology and Experimental Therapeutics, Windland Smith Rice Sudden Death Genomics Laboratory (D.Y., D.J.T., J.R.G., M.J.A.), Department of Medicine (J.R.G.), Division of Cardiovascular Diseases (M.J.A.), and Division of Pediatric Cardiology (M.J.A.), Mayo Clinic, Rochester, MN; Department of Molecular Biology and Pharmacology, Washington University School of Medicine, St Louis, MO (E.K.J., W.W., J.M.N.); Department of Molecular Medicine, University of Pavia, Pavia, Italy (L.C.); Center for Cardiac Arrhythmias of Genetic Origin, IRCCS Istituto Auxologico Italiano, Milan, Italy (L.C., F.D., M.T., P.J.S.); Institute of Human Genetics, Helmholtz Zentrum Munich, Neuherberg, Germany (L.C.); and Department of Clinical and Experimental Cardiology, Academic Medical Center, Amsterdam, The Netherlands (Y.M., M.A., A.A.M.W.).
authors: Yuka
authors: Y
authors: Mizusawa
authors: From the Center for Clinical and Translational Science (N.J.B.) and Mayo Graduate School (N.J.B.), Department Molecular Pharmacology and Experimental Therapeutics, Windland Smith Rice Sudden Death Genomics Laboratory (D.Y., D.J.T., J.R.G., M.J.A.), Department of Medicine (J.R.G.), Division of Cardiovascular Diseases (M.J.A.), and Division of Pediatric Cardiology (M.J.A.), Mayo Clinic, Rochester, MN; Department of Molecular Biology and Pharmacology, Washington University School of Medicine, St Louis, MO (E.K.J., W.W., J.M.N.); Department of Molecular Medicine, University of Pavia, Pavia, Italy (L.C.); Center for Cardiac Arrhythmias of Genetic Origin, IRCCS Istituto Auxologico Italiano, Milan, Italy (L.C., F.D., M.T., P.J.S.); Institute of Human Genetics, Helmholtz Zentrum Munich, Neuherberg, Germany (L.C.); and Department of Clinical and Experimental Cardiology, Academic Medical Center, Amsterdam, The Netherlands (Y.M., M.A., A.A.M.W.).
authors: Margherita
authors: M
authors: Torchio
authors: From the Center for Clinical and Translational Science (N.J.B.) and Mayo Graduate School (N.J.B.), Department Molecular Pharmacology and Experimental Therapeutics, Windland Smith Rice Sudden Death Genomics Laboratory (D.Y., D.J.T., J.R.G., M.J.A.), Department of Medicine (J.R.G.), Division of Cardiovascular Diseases (M.J.A.), and Division of Pediatric Cardiology (M.J.A.), Mayo Clinic, Rochester, MN; Department of Molecular Biology and Pharmacology, Washington University School of Medicine, St Louis, MO (E.K.J., W.W., J.M.N.); Department of Molecular Medicine, University of Pavia, Pavia, Italy (L.C.); Center for Cardiac Arrhythmias of Genetic Origin, IRCCS Istituto Auxologico Italiano, Milan, Italy (L.C., F.D., M.T., P.J.S.); Institute of Human Genetics, Helmholtz Zentrum Munich, Neuherberg, Germany (L.C.); and Department of Clinical and Experimental Cardiology, Academic Medical Center, Amsterdam, The Netherlands (Y.M., M.A., A.A.M.W.).
authors: Marielle
authors: M
authors: Alders
authors: From the Center for Clinical and Translational Science (N.J.B.) and Mayo Graduate School (N.J.B.), Department Molecular Pharmacology and Experimental Therapeutics, Windland Smith Rice Sudden Death Genomics Laboratory (D.Y., D.J.T., J.R.G., M.J.A.), Department of Medicine (J.R.G.), Division of Cardiovascular Diseases (M.J.A.), and Division of Pediatric Cardiology (M.J.A.), Mayo Clinic, Rochester, MN; Department of Molecular Biology and Pharmacology, Washington University School of Medicine, St Louis, MO (E.K.J., W.W., J.M.N.); Department of Molecular Medicine, University of Pavia, Pavia, Italy (L.C.); Center for Cardiac Arrhythmias of Genetic Origin, IRCCS Istituto Auxologico Italiano, Milan, Italy (L.C., F.D., M.T., P.J.S.); Institute of Human Genetics, Helmholtz Zentrum Munich, Neuherberg, Germany (L.C.); and Department of Clinical and Experimental Cardiology, Academic Medical Center, Amsterdam, The Netherlands (Y.M., M.A., A.A.M.W.).
authors: John R
authors: JR
authors: Giudicessi
authors: From the Center for Clinical and Translational Science (N.J.B.) and Mayo Graduate School (N.J.B.), Department Molecular Pharmacology and Experimental Therapeutics, Windland Smith Rice Sudden Death Genomics Laboratory (D.Y., D.J.T., J.R.G., M.J.A.), Department of Medicine (J.R.G.), Division of Cardiovascular Diseases (M.J.A.), and Division of Pediatric Cardiology (M.J.A.), Mayo Clinic, Rochester, MN; Department of Molecular Biology and Pharmacology, Washington University School of Medicine, St Louis, MO (E.K.J., W.W., J.M.N.); Department of Molecular Medicine, University of Pavia, Pavia, Italy (L.C.); Center for Cardiac Arrhythmias of Genetic Origin, IRCCS Istituto Auxologico Italiano, Milan, Italy (L.C., F.D., M.T., P.J.S.); Institute of Human Genetics, Helmholtz Zentrum Munich, Neuherberg, Germany (L.C.); and Department of Clinical and Experimental Cardiology, Academic Medical Center, Amsterdam, The Netherlands (Y.M., M.A., A.A.M.W.).
authors: Arthur A M
authors: AA
authors: Wilde
authors: From the Center for Clinical and Translational Science (N.J.B.) and Mayo Graduate School (N.J.B.), Department Molecular Pharmacology and Experimental Therapeutics, Windland Smith Rice Sudden Death Genomics Laboratory (D.Y., D.J.T., J.R.G., M.J.A.), Department of Medicine (J.R.G.), Division of Cardiovascular Diseases (M.J.A.), and Division of Pediatric Cardiology (M.J.A.), Mayo Clinic, Rochester, MN; Department of Molecular Biology and Pharmacology, Washington University School of Medicine, St Louis, MO (E.K.J., W.W., J.M.N.); Department of Molecular Medicine, University of Pavia, Pavia, Italy (L.C.); Center for Cardiac Arrhythmias of Genetic Origin, IRCCS Istituto Auxologico Italiano, Milan, Italy (L.C., F.D., M.T., P.J.S.); Institute of Human Genetics, Helmholtz Zentrum Munich, Neuherberg, Germany (L.C.); and Department of Clinical and Experimental Cardiology, Academic Medical Center, Amsterdam, The Netherlands (Y.M., M.A., A.A.M.W.).
authors: Peter J
authors: PJ
authors: Schwartz
authors: From the Center for Clinical and Translational Science (N.J.B.) and Mayo Graduate School (N.J.B.), Department Molecular Pharmacology and Experimental Therapeutics, Windland Smith Rice Sudden Death Genomics Laboratory (D.Y., D.J.T., J.R.G., M.J.A.), Department of Medicine (J.R.G.), Division of Cardiovascular Diseases (M.J.A.), and Division of Pediatric Cardiology (M.J.A.), Mayo Clinic, Rochester, MN; Department of Molecular Biology and Pharmacology, Washington University School of Medicine, St Louis, MO (E.K.J., W.W., J.M.N.); Department of Molecular Medicine, University of Pavia, Pavia, Italy (L.C.); Center for Cardiac Arrhythmias of Genetic Origin, IRCCS Istituto Auxologico Italiano, Milan, Italy (L.C., F.D., M.T., P.J.S.); Institute of Human Genetics, Helmholtz Zentrum Munich, Neuherberg, Germany (L.C.); and Department of Clinical and Experimental Cardiology, Academic Medical Center, Amsterdam, The Netherlands (Y.M., M.A., A.A.M.W.).
authors: Jeanne M
authors: JM
authors: Nerbonne
authors: From the Center for Clinical and Translational Science (N.J.B.) and Mayo Graduate School (N.J.B.), Department Molecular Pharmacology and Experimental Therapeutics, Windland Smith Rice Sudden Death Genomics Laboratory (D.Y., D.J.T., J.R.G., M.J.A.), Department of Medicine (J.R.G.), Division of Cardiovascular Diseases (M.J.A.), and Division of Pediatric Cardiology (M.J.A.), Mayo Clinic, Rochester, MN; Department of Molecular Biology and Pharmacology, Washington University School of Medicine, St Louis, MO (E.K.J., W.W., J.M.N.); Department of Molecular Medicine, University of Pavia, Pavia, Italy (L.C.); Center for Cardiac Arrhythmias of Genetic Origin, IRCCS Istituto Auxologico Italiano, Milan, Italy (L.C., F.D., M.T., P.J.S.); Institute of Human Genetics, Helmholtz Zentrum Munich, Neuherberg, Germany (L.C.); and Department of Clinical and Experimental Cardiology, Academic Medical Center, Amsterdam, The Netherlands (Y.M., M.A., A.A.M.W.). ackerman.michael@mayo.edu.
authors: Michael J
authors: MJ
authors: Ackerman
mesh_terms: D000328:Adult; D053840:Brugada Syndrome / Q000235:genetics / Q000378:metabolism* / Q000503:physiopathology; D004252:DNA Mutational Analysis; D004305:Dose-Response Relationship, Drug; D004562:Electrocardiography; D005260:Female; D020022:Genetic Predisposition to Disease; D057809:HEK293 Cells; D006801:Humans; D047468:Immunoprecipitation; D007700:Kinetics; D008297:Male; D008564:Membrane Potentials; D008875:Middle Aged; D020125:Mutation, Missense; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000378:metabolism*; D010641:Phenotype; D026902:Potassium Channel Blockers / Q000494:pharmacology; D011485:Protein Binding; D040121:Semaphorin-3A / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D051672:Shal Potassium Channels / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D014162:Transfection
chemical_list: C497021:KCND3 protein, human; D026902:Potassium Channel Blockers; C108639:SEMA3A protein, human; D040121:Semaphorin-3A; D051672:Shal Potassium Channels
keywords: Brugada syndrome; genetics, medical; ion channels; potassium channels; semaphorin-3A