This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Liu, W. J.
Right arrow Articles by Khromykh, A. A.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Liu, W. J.
Right arrow Articles by Khromykh, A. A.

 Previous Article  |  Next Article 

Journal of Virology, November 2004, p. 12225-12235, Vol. 78, No. 22
0022-538X/04/$08.00+0     DOI: 10.1128/JVI.78.22.12225-12235.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Analysis of Adaptive Mutations in Kunjin Virus Replicon RNA Reveals a Novel Role for the Flavivirus Nonstructural Protein NS2A in Inhibition of Beta Interferon Promoter-Driven Transcription{dagger}

Wen Jun Liu,1,2 Hua Bo Chen,1,2 Xiang Ju Wang,1,2 Hester Huang,1,2 and Alexander A. Khromykh1,2*

Sir Albert Sakzewski Virus Research Centre, Royal Children's Hospital,1 Clinical Medical Virology Centre, University of Queensland, Brisbane, Queensland, Australia2

Received 1 April 2004/ Accepted 20 July 2004

The establishment of persistent noncytopathic replication by replicon RNAs of a number of positive-strand RNA viruses usually leads to generation of adaptive mutations in nonstructural genes. Some of these adaptive mutations (e.g., in hepatitis C virus) increase the ability of RNA replication to resist the antiviral action of alpha/beta interferon (IFN-{alpha}/ß); others (e.g., in Sindbis virus) may also lead to more efficient IFN production. Using puromycin-selectable Kunjin virus (KUN) replicon RNA, we identified two adaptive mutations in the NS2A gene (producing Ala30-to-Pro and Asn101-to-Asp mutations in the gene product; for simplicity, these will be referred to hereafter as Ala30-to-Pro and Asn101-to-Asp mutations) that, when introduced individually or together into the original wild-type (wt) replicon RNA, resulted in ~15- to 50-fold more efficient establishment of persistent replication in hamster (BHK21) and human (HEK293 and HEp-2) cell lines. Transfection with a reporter plasmid carrying the luciferase gene under the control of the IFN-ß promoter resulted in ~6- to 7-fold-higher luciferase expression in HEp-2 cells stably expressing KUN replicon RNA with an Ala30-to-Pro mutation in the NS2A gene compared to that observed in HEp-2 cells stably expressing KUN replicon RNA with the wt NS2A gene. Moreover, cotransfection of plasmids expressing individual wt or Ala30-to-Pro-mutated NS2A genes with the IFN-ß promoter reporter plasmid, followed by infection with Semliki Forest virus to activate IFN-ß promoter-driven transcription, showed ~7-fold inhibition of luciferase expression by the wt but not by the Ala30-to-Pro-mutated NS2A protein. The results show for the first time a role for the flavivirus nonstructural protein NS2A in inhibition of IFN-ß promoter-driven transcription and identify a single-amino-acid mutation in NS2A that dramatically reduces this inhibitory activity. The findings determine a new function for NS2A in virus-host interactions, extend the range of KUN replicon vectors for noncytopathic gene expression, and identify NS2A as a new target for attenuation in the development of live flavivirus vaccines.

* Corresponding author. Mailing address: Sir Albert Sakzewski Virus Research Centre, Royal Children's Hospital, Brisbane, Queensland 4029, Australia. Phone: 617 36361568. Fax: 617 36361401. E-mail: a.khromykh{at}uq.edu.au.

{dagger} This is publication number 206 from the Clinical Medical Virology Centre and the Sir Albert Sakzewski Virus Research Centre.

Journal of Virology, November 2004, p. 12225-12235, Vol. 78, No. 22
0022-538X/04/$08.00+0     DOI: 10.1128/JVI.78.22.12225-12235.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Wilson, J. R., de Sessions, P. F., Leon, M. A., Scholle, F. (2008). West Nile Virus Nonstructural Protein 1 Inhibits TLR3 Signal Transduction. J. Virol. 82: 8262-8271 [Abstract] [Full Text]  
  • 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 [Abstract] [Full Text]  
  • Randall, R. E., Goodbourn, S. (2008). Interferons and viruses: an interplay between induction, signalling, antiviral responses and virus countermeasures. J. Gen. Virol. 89: 1-47 [Abstract] [Full Text]  
  • Noueiry, A. O., Olivo, P. D., Slomczynska, U., Zhou, Y., Buscher, B., Geiss, B., Engle, M., Roth, R. M., Chung, K. M., Samuel, M., Diamond, M. S. (2007). Identification of Novel Small-Molecule Inhibitors of West Nile Virus Infection. J. Virol. 81: 11992-12004 [Abstract] [Full Text]  
  • Garmashova, N., Gorchakov, R., Volkova, E., Paessler, S., Frolova, E., Frolov, I. (2007). The Old World and New World Alphaviruses Use Different Virus-Specific Proteins for Induction of Transcriptional Shutoff. J. Virol. 81: 2472-2484 [Abstract] [Full Text]  
  • Kinney, R. M., Huang, C. Y.-H., Whiteman, M. C., Bowen, R. A., Langevin, S. A., Miller, B. R., Brault, A. C. (2006). Avian virulence and thermostable replication of the North American strain of West Nile virus. J. Gen. Virol. 87: 3611-3622 [Abstract] [Full Text]  
  • Pijlman, G. P., Kondratieva, N., Khromykh, A. A. (2006). Translation of the Flavivirus Kunjin NS3 Gene in cis but Not Its RNA Sequence or Secondary Structure Is Essential for Efficient RNA Packaging. J. Virol. 80: 11255-11264 [Abstract] [Full Text]  
  • 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 [Abstract] [Full Text]  
  • Samuel, M. A., Diamond, M. S. (2005). Alpha/Beta Interferon Protects against Lethal West Nile Virus Infection by Restricting Cellular Tropism and Enhancing Neuronal Survival. J. Virol. 79: 13350-13361 [Abstract] [Full Text]  
  • Best, S. M., Morris, K. L., Shannon, J. G., Robertson, S. J., Mitzel, D. N., Park, G. S., Boer, E., Wolfinbarger, J. B., Bloom, M. E. (2005). Inhibition of Interferon-Stimulated JAK-STAT Signaling by a Tick-Borne Flavivirus and Identification of NS5 as an Interferon Antagonist. J. Virol. 79: 12828-12839 [Abstract] [Full Text]  
  • Kuno, G., Chang, G.-J. J. (2005). Biological Transmission of Arboviruses: Reexamination of and New Insights into Components, Mechanisms, and Unique Traits as Well as Their Evolutionary Trends. Clin. Microbiol. Rev. 18: 608-637 [Abstract] [Full Text]  
  • Munoz-Jordan, J. L., Laurent-Rolle, M., Ashour, J., Martinez-Sobrido, L., Ashok, M., Lipkin, W. I., Garcia-Sastre, A. (2005). Inhibition of Alpha/Beta Interferon Signaling by the NS4B Protein of Flaviviruses. J. Virol. 79: 8004-8013 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»