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International Journal of Legal Medicine

, Volume 128, Issue 1, pp 105–115 | Cite as

Identification and characterization of a novel genetic mutation with prolonged QT syndrome in an unexplained postoperative death

  • Yukiko Hata
  • Hisashi Mori
  • Ayumi Tanaka
  • Yosuke Fujita
  • Takeshi Shimomura
  • Toshihide Tabata
  • Koshi Kinoshita
  • Yoshiaki Yamaguchi
  • Fukiko Ichida
  • Yoshihiko Kominato
  • Noriaki Ikeda
  • Naoki NishidaEmail author
Original Article

Abstract

Introduction

The human ether-à-go-go-related gene (hERG) encodes the α-subunit of a cardiac potassium channel. Various mutations of hERG, including missense mutations, have been reported to cause long QT syndrome (LQTS) and severe arrhythmic disorders such as sudden cardiac death. We identified a novel hERG frameshift mutation (hERG(ΔAT)) in the S5-pore region from a LQTS patient who died suddenly and analyzed its genetic profile and the molecular and electrophysiological behaviors of the protein product to assess the pathogenicity of hERG(ΔAT).

Methods and results

We performed direct sequencing of hERG and evaluated its transcript level by using a whole blood sample from the patient. We performed immunoblotting, immunocytochemistry, and patch-clamp recordings of HEK-293 T cells transfected with hERG(ΔAT), wild-type hERG (hERG(WT)), or both. The patient demonstrated an AT deletion (c.1735_1736del) in hERG and a decrease in hERG mRNA transcripts. HEK-293 T cells showed lower production and cell surface expression of hERG(ΔAT) compared with hERG(WT) protein. In addition, the hERG(∆AT) protein failed to form functional channels, while the activation kinetics of functional channels, presumably consisting of hERG(WT) subunits, were unaffected.

Conclusion

The ΔAT mutation may decrease the number of functional hERG channels by impairing the posttranscriptional and posttranslational processing of the mutant product. This decrease may partly explain the cardiac symptoms of the patient who was heterozygous for hERG(ΔAT).

Keywords

M579fs + 75X frameshift mutation Human ether-à-go-go-related gene Long QT syndrome Patch-clamp Transmembrane pore domain Arrhythmia 

Notes

Acknowledgments

We thank Drs. Kenshi Hayashi (Kanazawa University Graduate School of Medical Science, Kanazawa, Japan), Sabina Kupershmidt (Vanderbilt University School of Medicine, Nashville, USA), and Jun-ichi Miyazaki (Osaka University Medical School, Osaka, Japan) for providing plasmids. We also thank Prof. K. Fukurotani for the opportunity to perform this work. This work was supported in part by KAKENHI grants from MEXT, Japan, to T.T. (19045019, 20022025, 20500284, 21026011, and 23500384) and a KAKENHI grant from JSPS, Japan, to Y.H. (24590852).

Conflict of interest

The authors declare that they have no conflict of interest.

Ethics statement

The experiments described in this manuscript conform to the Declaration of Helsinki, and the protocols of gene sampling and manipulation were approved by the University of Toyama’s committee on the usage of human genetic material (#24-1).

References

  1. 1.
    Splawski I, Shen J, Timothy KW, Lehmann MH, Priori S, Robinson JL, Moss AJ, Schwartz PJ, Towbin JA, Vincent GM, Keating MT (2000) Spectrum of mutation in long-QT syndrome genes: KVLQT1, HERG, SCN5A, KCNE1, and KCNE2. Circulation 102:1178–1185PubMedCrossRefGoogle Scholar
  2. 2.
    Splawski I, Timothy KW, Sharpe LM, Decher N, Kumar P, Bloise R, Napolitano C, Schwartz PJ, Joseph RM, Condouris K, Tager-Flusberg H, Priori SG, Sanguinetti MC, Keating MT (2004) Cav1.2 calcium channel dysfunction causes a multisystem disorder including arrhythmia and autism. Cell 119:19–31PubMedCrossRefGoogle Scholar
  3. 3.
    Vincent GM, Timothy KW, Leppert M, Keating M (1992) The spectrum of symptoms and QT intervals in carriers of the gene for the long-QT syndrome. N Engl J Med 327:846–852PubMedCrossRefGoogle Scholar
  4. 4.
    Hedley PL, Jørgensen P, Schlamowitz S, Wangari R, Moolman-Smook J, Brink PA, Kanters JK, Corfield VA, Christiansen M (2009) The genetic basis of long QT and short QT syndromes: a mutation update. Hum Mutat 30:1486–1511PubMedCrossRefGoogle Scholar
  5. 5.
    Yang Y, Yang Y, Liang B, Liu J, Li J, Grunnet M, Olesen SP, Rasmussen HB, Ellinor PT, Gao L, Lin X, Li L, Wang L, Xiao J, Liu Y, Liu Y, Zhang S, Liang D, Peng L, Jespersen T, Chen YH (2010) Identification of a Kir3.4 mutation in congenital long QT syndrome. Am J Hum Genet 86:872–880PubMedCentralPubMedCrossRefGoogle Scholar
  6. 6.
    Cerrone M, Priori SG (2011) Genetics of sudden death: focus on inherited channelopathies. Eur Heart J 32:2109–2118PubMedCrossRefGoogle Scholar
  7. 7.
    Perrin MJ, Subbiah RN, Vandenberg JI, Hill AP (2008) Human ether-a-go-go related gene (hERG) K + channels: function and dysfunction. Prog Biophys Mol Biol 98:137–148PubMedCrossRefGoogle Scholar
  8. 8.
    Shimizu W, Moss AJ, Wilde AA, Towbin JA, Ackerman MJ, January CT, Tester DJ, Zareba W, Robinson JL, Qi M, Vincent GM, Kaufman ES, Hofman N, Noda T, Kamakura S, Miyamoto Y, Shah S, Amin V, Goldenberg I, Andrews ML, McNitt S (2009) Genotype–phenotype aspects of type 2 long QT syndrome. J Am Coll Cardiol 54:2052–2062PubMedCentralPubMedCrossRefGoogle Scholar
  9. 9.
    Nishida N, Ikeda N, Kudo K, Tsuji A, Kiyoshima A (2002) Forensic significance of conduction system abnormalities as a precise cause of accidental death. Int J Legal Med 116:344–349PubMedGoogle Scholar
  10. 10.
    Harvey PA, Leinwand LA (2011) The cell biology of disease: cellular mechanisms of cardiomyopathy. J Cell Biol 194:355–365PubMedCrossRefGoogle Scholar
  11. 11.
    Hata Y, Kominato Y, Takizawa H (2007) Identification and characterization of a novel antisense RNA transcribed from the opposite strand of the human blood group ABO gene. Transfusion 47:842–851PubMedCrossRefGoogle Scholar
  12. 12.
    Snyders DJ, Chaudhary A (1996) High-affinity open channel block by dofetilide of HERG expressed in a human cell line. Mol Pharmacol 49:949–955PubMedGoogle Scholar
  13. 13.
    Niwa H, Yamamura K, Miyazaki J (1991) Efficient selection for high-expression transfectants with a novel eukaryotic vector. Gene 108:193–199PubMedCrossRefGoogle Scholar
  14. 14.
    Ho SN, Hunt HD, Horton RM, Pullen JK, Pease LR (1898) Site-directed mutagenesis by overlap extension using the polymerase chain reaction. Gene 77:51–59CrossRefGoogle Scholar
  15. 15.
    Ochman H, Gerber AS, Hartl DL (1988) Genetic applications of an inverse polymerase chain reaction. Genetics 120:621–623PubMedGoogle Scholar
  16. 16.
    Barahona-Dussault C, Benito B, Campuzano O, Iglesias A, Leung TL, Robb L, Talajic M, Brugada R (2010) Role of genetic testing in arrhythmogenic right ventricular cardiomyopathy/dysplasia. Clin Genet 77:37–48PubMedCrossRefGoogle Scholar
  17. 17.
    Gong Q, Keeney DR, Robinson JC, Zhou Z (2004) Defective assembly and trafficking of mutant HERG channels with C-terminal truncations in long QT syndrome. J Mol Cell Cardiol 37:1225–1233PubMedGoogle Scholar
  18. 18.
    Kinoshita K, Yamaguchi Y, Nishide K, Kimoto K, Nonobe Y, Fujita A, Asano K, Tabata T, Mori H, Inoue H, Hata Y, Fukurotani K, Nishida N (2012) A novel missense mutation causing a G487R substitution in the S2–S3 loop of human ether-à-go-go-related gene channel. J Cardiovasc Electrophysiol 23:1246–1253PubMedCrossRefGoogle Scholar
  19. 19.
    Tester DJ, Will ML, Haglund CM, Ackerman MJ (2005) Compendium of cardiac channel mutation in 541 consecutive unrelated patients referred for long QT syndrome genetic testing. Heart Rhythm 2:507–517PubMedCrossRefGoogle Scholar
  20. 20.
    Millat G, Chevalier P, Restier-Miron L, Da Costa A, Bouvagnet P, Kugener B, Fayol L, Gonzàlez Armengod C, Oddou B, Chanavat V, Froidefond E, Perraudin R, Rousson R, Rodriguez-Lafrasse C (2006) Spectrum of pathogenic mutations and associated polymorphisms in a cohort of 44 unrelated patients with long QT syndrome. Clin Genet 70:214–227PubMedCrossRefGoogle Scholar
  21. 21.
    Nagy E, Maquat LE (1998) A rule for termination-codon position within intron-containing genes: when nonsense affects RNA abundance. Trends Biochem Sci 23:198–199PubMedCrossRefGoogle Scholar
  22. 22.
    Stephenson LS, Maquat LE (1996) Cytoplasmic mRNA for human triosephosphate isomerase is immune to nonsense-mediated decay despite forming polysomes. Biochimie 78:1043–1047PubMedCrossRefGoogle Scholar
  23. 23.
    Yan Y, Teng S, Li N, Zhang Y, Boyden PA, Pu J (2009) Aminoglycoside antibiotics restore functional expression of truncated HERG channels produced by nonsense mutations. Hear Rhythm 6:553–560CrossRefGoogle Scholar
  24. 24.
    Teng S, Ma L, Dong Y, Lin C, Ye J, Bähring R, Vardanyan V, Yang Y, Lin Z, Pongs O, Hui R (2004) Clinical and electrophysiological characterization of a novel mutation R863X in HERG C-terminus associated with long QT syndrome. J Mol Med 82:189–196PubMedCrossRefGoogle Scholar
  25. 25.
    Zarraga IG, Zhang L, Stump MR, Gong Q, Vincent GM, Zhou Z (2011) Nonsense-mediated mRNA decay caused by a frameshift mutation in a large kindred of type 2 long QT syndrome. Hear Rhythm 8:1200–1206CrossRefGoogle Scholar
  26. 26.
    Gong Q, Zhang L, Vincent M, Horne BD, Zhou Z (2007) Nonsense mutations in hERG cause a decrease in mutant mRNA transcripts by nonsense-mediated mRNA decay in human long-QT syndrome. Circulation 116:17–24PubMedCentralPubMedCrossRefGoogle Scholar
  27. 27.
    Gong Q, Stump MR, Zhou Z (2011) Inhibition of nonsense-mediated mRNA decay by antisense morpholino oligonucleotides restores functional expression of hERG nonsense and frameshift mutations in long-QT syndrome. J Mol Cell Cardiol 50:223–229PubMedCentralPubMedCrossRefGoogle Scholar
  28. 28.
    Sanguinetti MC, Tristani-Firouzi M (2006) hERG potassium channels and cardiac arrhythmia. Nature 440:463–469PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2013

Authors and Affiliations

  • Yukiko Hata
    • 1
  • Hisashi Mori
    • 2
  • Ayumi Tanaka
    • 2
  • Yosuke Fujita
    • 3
  • Takeshi Shimomura
    • 3
  • Toshihide Tabata
    • 3
  • Koshi Kinoshita
    • 1
  • Yoshiaki Yamaguchi
    • 4
  • Fukiko Ichida
    • 5
  • Yoshihiko Kominato
    • 6
  • Noriaki Ikeda
    • 7
  • Naoki Nishida
    • 1
    Email author
  1. 1.Department of Legal Medicine, Faculty of Medicine, Graduate School of Medicine and Pharmaceutical Sciences for ResearchUniversity of ToyamaToyamaJapan
  2. 2.Department of Molecular Neuroscience, Graduate School of Medicine and Pharmaceutical Sciences for ResearchUniversity of ToyamaToyamaJapan
  3. 3.Laboratory for Neural Information Technology, Graduate School of Sciences and EngineeringUniversity of ToyamaToyamaJapan
  4. 4.Second Department of Internal Medicine, Graduate School of Medicine and Pharmaceutical Sciences for ResearchUniversity of ToyamaToyamaJapan
  5. 5.Department of Pediatrics, Graduate School of Medicine and Pharmaceutical Sciences for ResearchUniversity of ToyamaToyamaJapan
  6. 6.Department of Legal Medicine, Graduate School of MedicineGunma UniversityMaebashiJapan
  7. 7.Department of Forensic Pathology and Sciences, Graduate School of Medical SciencesKyushu UniversityFukuokaJapan

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