Non-structural protein 1 (NS1) binds small interfering RNA and suppresses RNA silencing in plants, but the underlying mechanism of this suppression is not well understood. Therefore, here we characterized NS1 encoded by the avian influenza virus H9N2. The NS1 protein was able to suppress RNA silencing induced by either sense RNA or double-stranded RNA (dsRNA). Using deletion and point mutants, we discovered that the first 70 residues of NS1 could suppress RNA silencing triggered by sense transgene, but this sequence was not sufficient to block dsRNA-induced silencing. Any mutations of two arginine residues (35R and 46R) of NS1, which contribute to its homodimeric structure, caused the loss of its silencing suppression activity. These results indicate that the region after residue 70 of NS1 is essential for the repression activity on dsRNA-induced RNA silencing, and that the dimeric structure of NS1 plays a critical role in its RNA silencing suppression function.


Article metrics loading...

Loading full text...

Full text loading...



  1. Bivalkar-Mehla S., Vakharia J., Mehla R., Abreha M., Kanwar J. R., Tikoo A., Chauhan A. (2011). Viral RNA silencing suppressors (RSS): novel strategy of viruses to ablate the host RNA interference (RNAi) defense system. Virus Res 155, 19. [View Article][PubMed] [Google Scholar]
  2. Bucher E., Hemmes H., de Haan P., Goldbach R., Prins M. (2004). The influenza A virus NS1 protein binds small interfering RNAs and suppresses RNA silencing in plants. J Gen Virol 85, 983991. [View Article][PubMed] [Google Scholar]
  3. Burgyán J., Havelda Z. (2011). Viral suppressors of RNA silencing. Trends Plant Sci 16, 265272. [View Article][PubMed] [Google Scholar]
  4. Chao J. A., Lee J. H., Chapados B. R., Debler E. W., Schneemann A., Williamson J. R. (2005). Dual modes of RNA-silencing suppression by Flock House virus protein B2. Nat Struct Mol Biol 12, 952957. [View Article][PubMed] [Google Scholar]
  5. Chapman E. J., Prokhnevsky A. I., Gopinath K., Dolja V. V., Carrington J. C. (2004). Viral RNA silencing suppressors inhibit the microRNA pathway at an intermediate step. Genes Dev 18, 11791186. [View Article][PubMed] [Google Scholar]
  6. Cheng A., Wong S. M., Yuan Y. A. (2009). Structural basis for dsRNA recognition by NS1 protein of influenza A virus. Cell Res 19, 187195. [View Article][PubMed] [Google Scholar]
  7. Chien C. Y., Xu Y., Xiao R., Aramini J. M., Sahasrabudhe P. V., Krug R. M., Montelione G. T. (2004). Biophysical characterization of the complex between double-stranded RNA and the N-terminal domain of the NS1 protein from influenza A virus: evidence for a novel RNA-binding mode. Biochemistry 43, 19501962. [View Article][PubMed] [Google Scholar]
  8. de Vries W., Haasnoot J., Fouchier R., de Haan P., Berkhout B. (2009). Differential RNA silencing suppression activity of NS1 proteins from different influenza A virus strains. J Gen Virol 90, 19161922. [View Article][PubMed] [Google Scholar]
  9. Deleris A., Gallego-Bartolome J., Bao J., Kasschau K. D., Carrington J. C., Voinnet O. (2006). Hierarchical action and inhibition of plant Dicer-like proteins in antiviral defense. Science 313, 6871. [View Article][PubMed] [Google Scholar]
  10. Delgadillo M. O., Sáenz P., Salvador B., García J. A., Simón-Mateo C. (2004). Human influenza virus NS1 protein enhances viral pathogenicity and acts as an RNA silencing suppressor in plants. J Gen Virol 85, 993999. [View Article][PubMed] [Google Scholar]
  11. Hatada E., Fukuda R. (1992). Binding of influenza A virus NS1 protein to dsRNA in vitro. J Gen Virol 73, 33253329. [View Article][PubMed] [Google Scholar]
  12. Hsieh Y. C., Omarov R. T., Scholthof H. B. (2009). Diverse and newly recognized effects associated with short interfering RNA binding site modifications on the Tomato bushy stunt virus p19 silencing suppressor. J Virol 83, 21882200. [View Article][PubMed] [Google Scholar]
  13. Jing X. L., Fan M. N., Jia G., Liu L. W., Ma L., Zheng C. C., Zhu X. P., Liu H. M., Wang X. Y. (2011). A multifunctional protein encoded by turkey herpesvirus suppresses RNA silencing in Nicotiana benthamiana . J Virol 85, 1279212803. [View Article][PubMed] [Google Scholar]
  14. Lakatos L., Csorba T., Pantaleo V., Chapman E. J., Carrington J. C., Liu Y. P., Dolja V. V., Calvino L. F., Lopez-Moya J. J., Burgyan J. (2006). Small RNA binding is a common strategy to suppress RNA silencing by several viral suppressors. EMBO J 25, 27682780.[CrossRef] [Google Scholar]
  15. Liu J., Lynch P. A., Chien C. Y., Montelione G. T., Krug R. M., Berman H. M., Dolja V. V., Calvino L. F., Liu J. (1997). Crystal structure of the unique RNA-binding domain of the influenza virus NS1 protein. Nat Struct Biol 4, 896899. [View Article][PubMed] [Google Scholar]
  16. Mérai Z., Kerényi Z., Kertész S., Magna M., Lakatos L., Silhavy D. (2006). Double-stranded RNA binding may be a general plant RNA viral strategy to suppress RNA silencing. J Virol 80, 57475756. [View Article][PubMed] [Google Scholar]
  17. Murayama R., Harada Y., Shibata T., Kuroda K., Hayakawa S., Shimizu K., Tanaka T. (2007). Influenza A virus non-structural protein 1 (NS1) interacts with cellular multifunctional protein nucleolin during infection. Biochem Biophys Res Commun 362, 880885. [View Article][PubMed] [Google Scholar]
  18. Omarov R., Sparks K., Smith L., Zindovic J., Scholthof H. B. (2006). Biological relevance of a stable biochemical interaction between the tombusvirus-encoded P19 and short interfering RNAs. J Virol 80, 30003008. [View Article][PubMed] [Google Scholar]
  19. Qian X. Y., Chien C. Y., Lu Y., Montelione G. T., Krug R. M. (1995). An amino-terminal polypeptide fragment of the influenza virus NS1 protein possesses specific RNA-binding activity and largely helical backbone structure. RNA 1, 948956.[PubMed] [Google Scholar]
  20. Silhavy D., Molnár A., Lucioli A., Szittya G., Hornyik C., Tavazza M., Burgyán J. (2002). A viral protein suppresses RNA silencing and binds silencing-generated, 21- to 25-nucleotide double-stranded RNAs. EMBO J 21, 30703080. [View Article][PubMed] [Google Scholar]
  21. Várallyay E., Oláh E., Havelda Z. (2014). Independent parallel functions of p19 plant viral suppressor of RNA silencing required for effective suppressor activity. Nucleic Acids Res 42, 599608. [View Article][PubMed] [Google Scholar]
  22. Wang W., Riedel K., Lynch P., Chien C. Y., Montelione G. T., Krug R. M. (1999). RNA binding by the novel helical domain of the influenza virus NS1 protein requires its dimer structure and a small number of specific basic amino acids. RNA 5, 195205. [View Article][PubMed] [Google Scholar]
  23. Wang X., Basler C. F., Williams B. R., Silverman R. H., Palese P., García-Sastre A. (2002). Functional replacement of the carboxy-terminal two-thirds of the influenza A virus NS1 protein with short heterologous dimerization domains. J Virol 76, 1295112962. [View Article][PubMed] [Google Scholar]
  24. Yin C., Khan J. A., Swapna G. V., Ertekin A., Krug R. M., Tong L., Montelione G. T. (2007). Conserved surface features form the double-stranded RNA binding site of non-structural protein 1 (NS1) from influenza A and B viruses. J Biol Chem 282, 2058420592. [View Article][PubMed] [Google Scholar]
  25. Zhang X., Yuan Y. R., Pei Y., Lin S. S., Tuschl T., Patel D. J., Chua N. H. (2006). Cucumber mosaic virus-encoded 2b suppressor inhibits Arabidopsis Argonaute1 cleavage activity to counter plant defense. Genes Dev 20, 32553268. [View Article][PubMed] [Google Scholar]

Data & Media loading...

Most cited this month Most Cited RSS feed

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