2′-O methylation of the viral mRNA cap evades host restriction by IFIT family members

Article metrics

Abstract

Cellular messenger RNA (mRNA) of higher eukaryotes and many viral RNAs are methylated at the N-7 and 2′-O positions of the 5′ guanosine cap by specific nuclear and cytoplasmic methyltransferases (MTases), respectively. Whereas N-7 methylation is essential for RNA translation and stability1, the function of 2′-O methylation has remained uncertain since its discovery 35 years ago2,3,4. Here we show that a West Nile virus (WNV) mutant (E218A) that lacks 2′-O MTase activity was attenuated in wild-type primary cells and mice but was pathogenic in the absence of type I interferon (IFN) signalling. 2′-O methylation of viral RNA did not affect IFN induction in WNV-infected fibroblasts but instead modulated the antiviral effects of IFN-induced proteins with tetratricopeptide repeats (IFIT), which are interferon-stimulated genes (ISGs) implicated in regulation of protein translation. Poxvirus and coronavirus mutants that lacked 2′-O MTase activity similarly showed enhanced sensitivity to the antiviral actions of IFN and, specifically, IFIT proteins. Our results demonstrate that the 2′-O methylation of the 5′ cap of viral RNA functions to subvert innate host antiviral responses through escape of IFIT-mediated suppression, and suggest an evolutionary explanation for 2′-O methylation of cellular mRNA: to distinguish self from non-self RNA. Differential methylation of cytoplasmic RNA probably serves as an example for pattern recognition and restriction of propagation of foreign viral RNA in host cells.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Figure 1: WNV-E218A is attenuated in wild-type mice and cells but is virulent in Ifnar1 −/− mice and cells.
Figure 2: 2′- O methylation of viral RNA alters the sensitivity of WNV to the antiviral effects of IFN.
Figure 3: WNV-E218A is more sensitive to the antiviral actions of Ifit genes.
Figure 4: Poxvirus and coronavirus mutants lacking 2′- O methylation are more sensitive to the antiviral effects of murine IFIT-2.

References

  1. 1

    Furuichi, Y. & Shatkin, A. J. Viral and cellular mRNA capping: past and prospects. Adv. Virus Res. 55, 135–184 (2000)

  2. 2

    Wei, C. M. & Moss, B. Methylated nucleotides block 5′-terminus of vaccinia virus messenger RNA. Proc. Natl Acad. Sci. USA 72, 318–322 (1975)

  3. 3

    Wei, C. M., Gershowitz, A. & Moss, B. Methylated nucleotides block 5′ terminus of HeLa cell messenger RNA. Cell 4, 379–386 (1975)

  4. 4

    Muthukrishnan, S., Moss, B., Cooper, J. A. & Maxwell, E. S. Influence of 5′-terminal cap structure on the initiation of translation of vaccinia virus mRNA. J. Biol. Chem. 253, 1710–1715 (1978)

  5. 5

    Langberg, S. R. & Moss, B. Post-transcriptional modifications of mRNA. Purification and characterization of cap I and cap II RNA (nucleoside-2′-)-methyltransferases from HeLa cells. J. Biol. Chem. 256, 10054–10060 (1981)

  6. 6

    Fechter, P. & Brownlee, G. G. Recognition of mRNA cap structures by viral and cellular proteins. J. Gen. Virol. 86, 1239–1249 (2005)

  7. 7

    Dong, H., Zhang, B. & Shi, P. Y. Flavivirus methyltransferase: a novel antiviral target. Antiviral Res. 80, 1–10 (2008)

  8. 8

    Zhou, Y. et al. Structure and function of flavivirus NS5 methyltransferase. J. Virol. 81, 3891–3903 (2007)

  9. 9

    Samuel, M. A. & Diamond, M. S. Alpha/beta IFN protects against lethal West Nile virus infection by restricting cellular tropism and enhancing neuronal survival. J. Virol. 79, 13350–13361 (2005)

  10. 10

    Daffis, S. et al. Interferon regulatory factor IRF-7 induces the antiviral alpha interferon response and protects against lethal West Nile virus infection. J. Virol. 82, 8465–8475 (2008)

  11. 11

    Daffis, S., Suthar, M. S., Szretter, K. J., Gale, M., Jr & Diamond, M. S. Induction of IFN-β 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 (2009)

  12. 12

    Daffis, S., Samuel, M. A., Suthar, M. S., Gale, M., Jr & Diamond, M. S. Toll-like receptor 3 has a protective role against West Nile virus infection. J. Virol. 82, 10349–10358 (2008)

  13. 13

    Der, S. D., Zhou, A., Williams, B. R. & Silverman, R. H. Identification of genes differentially regulated by interferon α, β, or γ using oligonucleotide arrays. Proc. Natl Acad. Sci. USA 95, 15623–15628 (1998)

  14. 14

    Wacher, C. et al. Coordinated regulation and widespread cellular expression of interferon-stimulated genes (ISG) ISG-49, ISG-54, and ISG-56 in the central nervous system after infection with distinct viruses. J. Virol. 81, 860–871 (2007)

  15. 15

    Daffis, S., Samuel, M. A., Keller, B. C., Gale, M., Jr & Diamond, M. S. Cell-specific IRF-3 responses protect against West Nile virus infection by interferon-dependent and -independent mechanisms. PLoS Pathog. 3, e106 (2007)

  16. 16

    Fensterl, V., White, C. L., Yamashita, M. & Sen, G. C. Novel characteristics of the function and induction of murine p56 family proteins. J. Virol. 82, 11045–11053 (2008)

  17. 17

    Sumpter, R., Jr et al. Regulating intracellular antiviral defense and permissiveness to hepatitis C virus RNA replication through a cellular RNA helicase, RIG-I. J. Virol. 79, 2689–2699 (2005)

  18. 18

    Zhang, Y., Burke, C. W., Ryman, K. D. & Klimstra, W. B. Identification and characterization of interferon-induced proteins that inhibit alphavirus replication. J. Virol. 81, 11246–11255 (2007)

  19. 19

    Terenzi, F., Hui, D. J., Merrick, W. C. & Sen, G. C. Distinct induction patterns and functions of two closely related interferon-inducible human genes, ISG54 and ISG56. J. Biol. Chem. 281, 34064–34071 (2006)

  20. 20

    Hui, D. J., Terenzi, F., Merrick, W. C. & Sen, G. C. Mouse p56 blocks a distinct function of eukaryotic initiation factor 3 in translation initiation. J. Biol. Chem. 280, 3433–3440 (2005)

  21. 21

    Sarkar, S. N. & Sen, G. C. Novel functions of proteins encoded by viral stress-inducible genes. Pharmacol. Ther. 103, 245–259 (2004)

  22. 22

    Latner, D. R., Thompson, J. M., Gershon, P. D., Storrs, C. & Condit, R. C. The positive transcription elongation factor activity of the vaccinia virus J3 protein is independent from its (nucleoside-2′-O-) methyltransferase and poly(A) polymerase stimulatory functions. Virology 301, 64–80 (2002)

  23. 23

    Decroly, E. et al. Coronavirus nonstructural protein 16 is a cap-0 binding enzyme possessing (nucleoside-2′O)-methyltransferase activity. J. Virol. 82, 8071–8084 (2008)

  24. 24

    Hui, D. J., Bhasker, C. R., Merrick, W. C. & Sen, G. C. Viral stress-inducible protein p56 inhibits translation by blocking the interaction of eIF3 with the ternary complex eIF2.GTP.Met-tRNAi. J. Biol. Chem. 278, 39477–39482 (2003)

  25. 25

    Coley, S. E. et al. Recombinant mouse hepatitis virus strain A59 from cloned, full-length cDNA replicates to high titers in vitro and is fully pathogenic in vivo. J. Virol. 79, 3097–3106 (2005)

Download references

Acknowledgements

This work was supported by National Institutes of Health grants U54 AI081680, U19 AI083019 and R01 AI074973 (to M.G. and M.S.D.), R01 AI56540 (to S.S.), U54 AI057160 (to R.M.B.) and U54 AI057158 (to P.Y.S.), R01 CA068782 (to G.C.S.) the Swiss National Science Foundation, 3100A0-118425/1, and the Novartis Foundation (to V.T. and R.Z.). We thank H. Virgin and B. Moss for reading the manuscript.

Author information

S.D., K.J.S., R.M.B., T.C.P., M.G., P.-Y.S. and M.S.D. designed the experiments. S.D., K.J.S., J.S., J.L., T.-Y.L. and H.D. performed the experiments. S.D., K.J.S., J.S., R.M.B., M.G., P.-Y.S. and M.S.D. analysed the data. S.Y., V.F., G.C.S., W.B.K., R.Z. and V.T. provided key reagents and expertise. S.D., K.J.S., R.M.B, T.C.P., M.G., P.-Y.S. and M.S.D. wrote and edited the manuscript.

Correspondence to Michael S. Diamond.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Information

This file contains Supplementary Figures 1-8 with legends, Supplementary Methods and additional references. (PDF 2375 kb)

PowerPoint slides

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Daffis, S., Szretter, K., Schriewer, J. et al. 2′-O methylation of the viral mRNA cap evades host restriction by IFIT family members. Nature 468, 452–456 (2010) doi:10.1038/nature09489

Download citation

Further reading

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.