1887

Abstract

The flavivirus NS2A protein is a small, multifunctional protein, involved in replication, virion formation and regulation of the innate immune response. Using the Kunjin strain of West Nile virus (WNV) we previously demonstrated that a single amino acid change from alanine to proline at position 30 of the NS2A protein (A30P) reduced viral cytopathicity in cells and virulence in mice. To further investigate functions of the NS2A protein we have substituted alanine at position 30 with different amino acids (A30 mutants) in a WNV infectious clone. The virulence of mutant viruses in wild-type (WT) and IRF3/IRF7 double-knockout mice was influenced by the amino acid change and ranged from high to low in the order of WT>A30L>A30E>A30P/A30G. Moreover, infection of beta interferon (IFN-β)-deficient Vero cells with A30P virus showed less pronounced chromosomal DNA degradation and lower percentage of cells with positive TUNEL labelling than in WT virus infection, indicating a role for the WT NS2A protein in IFN-independent apoptotic cell death.

Loading

Article metrics loading...

/content/journal/jgv/10.1099/vir.0.047076-0
2013-02-01
2020-01-24
Loading full text...

Full text loading...

/deliver/fulltext/jgv/94/2/308.html?itemId=/content/journal/jgv/10.1099/vir.0.047076-0&mimeType=html&fmt=ahah

References

  1. Ashour J., Laurent-Rolle M., Shi P. Y., García-Sastre A.. ( 2009;). NS5 of dengue virus mediates STAT2 binding and degradation. . J Virol 83:, 5408–5418. [CrossRef][PubMed]
    [Google Scholar]
  2. Chang Y. S., Liao C. L., Tsao C. H., Chen M. C., Liu C. I., Chen L. K., Lin Y. L.. ( 1999;). Membrane permeabilization by small hydrophobic nonstructural proteins of Japanese encephalitis virus. . J Virol 73:, 6257–6264.[PubMed]
    [Google Scholar]
  3. Chu J. J., Ng M. L.. ( 2003;). The mechanism of cell death during West Nile virus infection is dependent on initial infectious dose. . J Gen Virol 84:, 3305–3314. [CrossRef][PubMed]
    [Google Scholar]
  4. Coia G., Parker M. D., Speight G., Byrne M. E., Westaway E. G.. ( 1988;). Nucleotide and complete amino acid sequences of Kunjin virus: definitive gene order and characteristics of the virus-specified proteins. . J Gen Virol 69:, 1–21. [CrossRef][PubMed]
    [Google Scholar]
  5. Daffis S., Suthar M. S., Szretter K. J., Gale M. Jr, Diamond M. S.. ( 2009;). Induction of IFN-beta and the innate antiviral response in myeloid cells occurs through an IPS-1-dependent signal that does not require IRF-3 and IRF-7. . PLoS Pathog 5:, e1000607. [CrossRef][PubMed]
    [Google Scholar]
  6. Diamond M. S.. ( 2009a;). Mechanisms of evasion of the type I interferon antiviral response by flaviviruses. . J Interferon Cytokine Res 29:, 521–530. [CrossRef][PubMed]
    [Google Scholar]
  7. Diamond M. S.. ( 2009b;). West Nile Encephalitis Virus Infection. New York:: Springer;. [CrossRef]
    [Google Scholar]
  8. Firth A. E., Atkins J. F.. ( 2009;). A conserved predicted pseudoknot in the NS2A-encoding sequence of West Nile and Japanese encephalitis flaviviruses suggests NS1′ may derive from ribosomal frameshifting. . Virol J 6:, 14. [CrossRef][PubMed]
    [Google Scholar]
  9. Kleinschmidt M. C., Michaelis M., Ogbomo H., Doerr H. W., Cinatl J. Jr. ( 2007;). Inhibition of apoptosis prevents West Nile virus induced cell death. . BMC Microbiol 7:, 49. [CrossRef][PubMed]
    [Google Scholar]
  10. Koh W. L., Ng M. L.. ( 2005;). Molecular mechanisms of West Nile virus pathogenesis in brain cell. . Emerg Infect Dis 11:, 629–632. [CrossRef][PubMed]
    [Google Scholar]
  11. Kümmerer B. M., Rice C. M.. ( 2002;). Mutations in the yellow fever virus nonstructural protein NS2A selectively block production of infectious particles. . J Virol 76:, 4773–4784. [CrossRef][PubMed]
    [Google Scholar]
  12. Leung J. Y., Pijlman G. P., Kondratieva N., Hyde J., Mackenzie J. M., Khromykh A. A.. ( 2008;). Role of nonstructural protein NS2A in flavivirus assembly. . J Virol 82:, 4731–4741. [CrossRef][PubMed]
    [Google Scholar]
  13. Lindenbach B. D., Rice C. M.. ( 1997;). trans-Complementation of yellow fever virus NS1 reveals a role in early RNA replication. . J Virol 71:, 9608–9617.[PubMed]
    [Google Scholar]
  14. Lindenbach B. D., Rice C. M.. ( 2001;). Flaviviridae: their viruses and their replication. . In Fields Virology, , 4th edn., pp. 991–1041. Edited by Knipe D. M., Howley P. M... Philadelphia:: Lippincott Williams & Wilkins;.
    [Google Scholar]
  15. Liu W. J., Chen H. B., Khromykh A. A.. ( 2003;). Molecular and functional analyses of Kunjin virus infectious cDNA clones demonstrate the essential roles for NS2A in virus assembly and for a nonconservative residue in NS3 in RNA replication. . J Virol 77:, 7804–7813. [CrossRef][PubMed]
    [Google Scholar]
  16. Liu W. J., Wang X. J., Mokhonov V. V., Shi P. Y., Randall R., Khromykh A. A.. ( 2005;). Inhibition of interferon signaling by the New York 99 strain and Kunjin subtype of West Nile virus involves blockage of STAT1 and STAT2 activation by nonstructural proteins. . J Virol 79:, 1934–1942. [CrossRef][PubMed]
    [Google Scholar]
  17. Liu W. J., Wang X. J., Clark D. C., Lobigs M., Hall R. A., Khromykh A. A.. ( 2006;). A single amino acid substitution in the West Nile virus nonstructural protein NS2A disables its ability to inhibit alpha/beta interferon induction and attenuates virus virulence in mice. . J Virol 80:, 2396–2404. [CrossRef][PubMed]
    [Google Scholar]
  18. Mackenzie J. M., Khromykh A. A., Jones M. K., Westaway E. G.. ( 1998;). Subcellular localization and some biochemical properties of the flavivirus Kunjin nonstructural proteins NS2A and NS4A. . Virology 245:, 203–215. [CrossRef][PubMed]
    [Google Scholar]
  19. McAllister C. S., Samuel C. E.. ( 2009;). The RNA-activated protein kinase enhances the induction of interferon-beta and apoptosis mediated by cytoplasmic RNA sensors. . J Biol Chem 284:, 1644–1651. [CrossRef][PubMed]
    [Google Scholar]
  20. McElroy K. L., Tsetsarkin K. A., Vanlandingham D. L., Higgs S.. ( 2006;). Manipulation of the yellow fever virus non-structural genes 2A and 4B and the 3’non-coding region to evaluate genetic determinants of viral dissemination from the Aedes aegypti midgut. . Am J Trop Med Hyg 75:, 1158–1164.[PubMed]
    [Google Scholar]
  21. Melian E. B., Hinzman E., Nagasaki T., Firth A. E., Wills N. M., Nouwens A. S., Blitvich B. J., Leung J., Funk A.. & other authors ( 2010;). NS1′ of flaviviruses in the Japanese encephalitis virus serogroup is a product of ribosomal frameshifting and plays a role in viral neuroinvasiveness. . J Virol 84:, 1641–1647. [CrossRef][PubMed]
    [Google Scholar]
  22. Nestorowicz A., Chambers T. J., Rice C. M.. ( 1994;). Mutagenesis of the yellow fever virus NS2A/2B cleavage site: effects on proteolytic processing, viral replication, and evidence for alternative processing of the NS2A protein. . Virology 199:, 114–123. [CrossRef][PubMed]
    [Google Scholar]
  23. Patkar C. G., Kuhn R. J.. ( 2008;). Yellow fever virus NS3 plays an essential role in virus assembly independent of its known enzymatic functions. . J Virol 82:, 3342–3352. [CrossRef][PubMed]
    [Google Scholar]
  24. Prikhod’ko G. G., Prikhod’ko E. A., Pletnev A. G., Cohen J. I.. ( 2002;). Langat flavivirus protease NS3 binds caspase-8 and induces apoptosis. . J Virol 76:, 5701–5710. [CrossRef][PubMed]
    [Google Scholar]
  25. Rossi S. L., Fayzulin R., Dewsbury N., Bourne N., Mason P. W.. ( 2007;). Mutations in West Nile virus nonstructural proteins that facilitate replicon persistence in vitro attenuate virus replication in vitro and in vivo. . Virology 364:, 184–195. [CrossRef][PubMed]
    [Google Scholar]
  26. Samuel M. A., Morrey J. D., Diamond M. S.. ( 2007;). Caspase 3-dependent cell death of neurons contributes to the pathogenesis of West Nile virus encephalitis. . J Virol 81:, 2614–2623. [CrossRef][PubMed]
    [Google Scholar]
  27. Scherbik S. V., Stockman B. M., Brinton M. A.. ( 2007;). Differential expression of interferon (IFN) regulatory factors and IFN-stimulated genes at early times after West Nile virus infection of mouse embryo fibroblasts. . J Virol 81:, 12005–12018. [CrossRef][PubMed]
    [Google Scholar]
  28. Schuessler A., Funk A., Lazear H. M., Cooper D. A., Torres S., Daffis S., Jha B. K., Kumagai Y., Takeuchi O.. & other authors ( 2012;). West Nile virus noncoding subgenomic RNA contributes to viral evasion of the type I interferon-mediated antiviral response. . J Virol 86:, 5708–5718. [CrossRef][PubMed]
    [Google Scholar]
  29. Westaway E. G., Mackenzie J. M., Khromykh A. A.. ( 2003;). Kunjin RNA replication and applications of Kunjin replicons. . Adv Virus Res 59:, 99–140. [CrossRef][PubMed]
    [Google Scholar]
  30. Wicker J. A., Whiteman M. C., Beasley D. W., Davis C. T., Zhang S., Schneider B. S., Higgs S., Kinney R. M., Barrett A. D.. ( 2006;). A single amino acid substitution in the central portion of the West Nile virus NS4B protein confers a highly attenuated phenotype in mice. . Virology 349:, 245–253. [CrossRef][PubMed]
    [Google Scholar]
  31. Yang T. C., Shiu S. L., Chuang P. H., Lin Y. J., Wan L., Lan Y. C., Lin C. W.. ( 2009;). Japanese encephalitis virus NS2B-NS3 protease induces caspase 3 activation and mitochondria-mediated apoptosis in human medulloblastoma cells. . Virus Res 143:, 77–85. [CrossRef][PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/vir.0.047076-0
Loading
/content/journal/jgv/10.1099/vir.0.047076-0
Loading

Data & Media loading...

Most cited articles

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error