Advertisement

Archives of Virology

, Volume 158, Issue 5, pp 1003–1011 | Cite as

Attenuation of an influenza A virus due to alteration of its hemagglutinin-neuraminidase functional balance in mice

  • Fumihiro Gen
  • Shinya Yamada
  • Kentaro Kato
  • Hiroomi Akashi
  • Yoshihiro Kawaoka
  • Taisuke HorimotoEmail author
Original Article

Abstract

Influenza A viruses possess two surface glycoproteins, hemagglutinin (HA), which binds to sialic-acid-containing receptors, and neuraminidase (NA), which removes sialic acid from host cells. It is well established that the HA-NA functional balance regulates the efficiency of virus replication. Here, we selected a plaque variant of the WSN (H1N1) strain that grew better than the wild-type virus in NA-expressing MDCK cell culture. A reverse genetics study revealed that the single mutation HA E190K, which occurs infrequently in naturally isolated H1N1 viruses, was responsible for the phenotype of this variant. Receptor assays indicated that this mutation did not affect the receptor specificity of HA but enhanced its receptor-binding affinity, resulting in altered HA-NA functional balance relative to that of the wild-type virus. We also found that this variant replicated in nasal turbinates at an equivalent level but in lungs at a lower level compared with wild-type virus, demonstrating its attenuation in mice. Together, our data demonstrated the importance of the HA-NA functional balance for influenza virus replication in an in vivo biological setting.

Keywords

Sialic Acid H1N1 Virus MDCK Cell Parent Virus Large Plaque 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Notes

Acknowledgments

We thank Dr. Emi Takashita and Susan Watson for providing anti-NA antibody and editing the manuscript. This work was supported by grants-in-aid for Specially Promoted Research and for Scientific Research (B), from the Ministry of Education, Culture, Sports, Science, and Technology, by grants-in-aid from the Ministry of Health, Labor and Welfare, by ERATO, and by National Institute of Allergy and Infectious Diseases Public Health Service research grants.

Conflict of interest

The authors declare that they have no conflicts of interest

References

  1. 1.
    Chen H, Wen X, To KK, Wang P, Tse H, chan JF, Tsoi HW, Fung KS, Tse CW, Lee RA, Chan KH, Yuen KY (2010) Quasispecies of the D225G substitution in the hemagglutinin of pandemic influenza A(H1N1) 2009 virus from patients with severe disease in Hong Kong, China. J Infect Dis 201:1517–1521Google Scholar
  2. 2.
    Francis T, Moore AL (1940) A study of the neurotropic tendency in strains of the virus of epidemic influenza. J Exp Med 72:717–728PubMedCrossRefGoogle Scholar
  3. 3.
    Gamblin SJ, Skehel JJ (2010) Influenza hemagglutinin and neuraminidase membrane glycoproteins. J Biol Chem 285:28403–28409PubMedCrossRefGoogle Scholar
  4. 4.
    Horimoto T, Kawaoka Y (2005) Influenza lessons from past pandemics, warnings from current incidents. Nat Rev Microbiol 3:591–600PubMedCrossRefGoogle Scholar
  5. 5.
    Hughes MT, Matrosovich M, Rodgers ME, McGregor M, Kawaoka Y (2000) Influenza A viruses lacking sialidase activity can undergo multiple cycles of replication in cell culture, eggs, or mice. J Virol 74:5206–5212PubMedCrossRefGoogle Scholar
  6. 6.
    Hughes MT, McGregor M, Suzuki T, Suzuki Y, Kawaoka Y (2001) Adaptation of influenza A viruses to cells expressing low levels of sialic acid leads to loss of neuraminidase activity. J Virol 75:3766–3770PubMedCrossRefGoogle Scholar
  7. 7.
    Ito T, Suzuki Y, Mitnaul L, Vines A, Kida H, Kawaoka Y (1997) Receptor specificity of influenza A viruses correlates with the agglutinin of erythrocytes from different animal species. Virology 227:493–499PubMedCrossRefGoogle Scholar
  8. 8.
    Ito T, Suzuki Y, Takada A, Kawamoto K, Otsuki K, Masuda H, Yamada M, Suzuki T, Kida H, Kawaoka Y (1997) Differences in sialic acid-galactose linkages in the chicken egg amnion and allantois influence human influenza virus receptor specificity and variant selection. J Virol 71:3357–3362PubMedGoogle Scholar
  9. 9.
    Jin H, Lu B, Zhou H, Ma C, Zhao J, Yang CF, Kemble G, Greenberg H (2003) Multiple amino acid residues confer temperature sensitivity to human influenza virus vaccine strain (FluMist) derived from cold-adapted A/Ann Arbor/6/60. Virology 306:18–24PubMedCrossRefGoogle Scholar
  10. 10.
    Liu Y, Childs RA, Matrosovich T, Wharton S, Palma AS, Chai W, Daniels R, Gregory V, Uhlendorff J, Kiso M, Klenk H-D, Hay A, Feizi T, Matrosovich M (2010) Altered receptor specificity and cell tropism of D222G hemagglutinin mutants isolated from fatal cases of pandemic A(H1N1) 2009 influenza virus. J Virol 84:12069–12074PubMedCrossRefGoogle Scholar
  11. 11.
    Matrosovich M, Tuzikov A, Bovin N, Gambaryan A, Klimov A, Castrucci MR, Donatelli I, Kawaoka Y (2000) Early alterations of the receptor-binding properties of H1, H2, and H3 avian influenza virus hemagglutinins after their introduction into mammals. J Virol 74:8502–8512PubMedCrossRefGoogle Scholar
  12. 12.
    Medeiros R, Escriou N, Naffakh N, Manuguerra J-C, van der Werf S (2001) Hemagglutinin residues of recent human A(H3N2) influenza viruses that contribute to the inability to agglutinate chicken erythrocytes. Virology 289:74–85PubMedCrossRefGoogle Scholar
  13. 13.
    Mitnaul LJ, Matrosovich M, Castrucci MR, Tuzikov AB, Bovin NV, Kobasa D, Kawaoka Y (2000) Balanced hemagglutinin and neuraminidase activities are critical for efficient replication of influenza A virus. J Virol 74:6015–6020PubMedCrossRefGoogle Scholar
  14. 14.
    Mochalova L, Kurova V, Shtyrya Y, Korchagina E, Gambaryan A, Belyanchikov I, Bovin N (2007) Oligosaccharide specificity of influenza H1N1 virus neuraminidases. Arch Virol 152:2047–2057PubMedCrossRefGoogle Scholar
  15. 15.
    Neumann G, Watanabe T, Ito H, Watanabe S, Goto H, Gao P, Hughes MT, Perez DR, Donis R, Hoffmann E, Hobom G, Kawaoka Y (1999) Generation of influenza A viruses entirely from cloned cDNAs. Proc Natl Acad Sci USA 96:9345–9350PubMedCrossRefGoogle Scholar
  16. 16.
    Smith W, Andrews CH, Laidlaw PP (1933) A virus obtained from influenza patients. Lancet 2:66–68CrossRefGoogle Scholar
  17. 17.
    Totani K, Kubota T, Kuroda T, Murata T, Hidari KI, Suzuki T, Suzuki Y, Kobayashi K, Ashida H, Yamamoto K, Usui T (2003) Chemoenzymatic synthesis and application of glycopolymers containing multivalent sialyloligosaccharides with a poly(l-glutamic acid) backbone for inhibition of infection by influenza viruses. Glycobiology 13:315–326PubMedCrossRefGoogle Scholar
  18. 18.
    Wagner R, Matrosovich M, Klenk H-D (2002) Functional balance between haemagglutinin and neuraminidase in influenza virus infections. Rev Med Virol 1:159–166CrossRefGoogle Scholar
  19. 19.
    Wagner R, Wolff T, Herwig A, Pleschka S, Klenk H-D (2000) Interdependence of hemagglutinin glycosylation and neuraminidase as regulators of influenza virus growth: a study by reverse genetics. J Virol 74:6316–6323PubMedCrossRefGoogle Scholar
  20. 20.
    Yen HL, Liang CH, Wu CY, Forrest HL, Ferguson A, Choy KT, Jones J, Wong DD, Cheung PP, Hsu CH, Li OT, Yuen KM, Chan RW, Poon LL, Chan MC, Nicholls JM, Krauss S, Wong CH, Guan Y, Webster RG, Webby RJ, Peiris M (2011) Hemagglutinin-neuraminidase balance confers respiratory-droplet transmissibility of the pandemic H1N1 influenza virus in ferrets. Proc Natl Acad Sci USA 108:14264–14269PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag Wien 2012

Authors and Affiliations

  • Fumihiro Gen
    • 1
  • Shinya Yamada
    • 2
  • Kentaro Kato
    • 1
  • Hiroomi Akashi
    • 1
  • Yoshihiro Kawaoka
    • 2
    • 3
    • 4
    • 5
  • Taisuke Horimoto
    • 1
    Email author
  1. 1.Department of Veterinary Microbiology, Graduate School of Agricultural and Life SciencesThe University of TokyoTokyoJapan
  2. 2.Division of Virology, Department of Microbiology and Immunology, Institute of Medical ScienceThe University of TokyoTokyoJapan
  3. 3.ERATO Infection-Induced Host Responses ProjectSaitamaJapan
  4. 4.Department of Pathobiological Sciences, School of Veterinary MedicineUniversity of Wisconsin-MadisonMadisonUSA
  5. 5.International Research Center for Infectious Diseases, Institute of Medical ScienceThe University of TokyoTokyoJapan

Personalised recommendations