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Induction and suppression of antiviral RNA interference by influenza A virus in mammalian cells

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Abstract

Influenza A virus (IAV) causes annual epidemics and occasional pandemics, and is one of the best-characterized human RNA viral pathogens1. However, a physiologically relevant role for the RNA interference (RNAi) suppressor activity of the IAV non-structural protein 1 (NS1), reported over a decade ago2, remains unknown3. Plant and insect viruses have evolved diverse virulence proteins to suppress RNAi as their hosts produce virus-derived small interfering RNAs (siRNAs) that direct specific antiviral defence47 by an RNAi mechanism dependent on the slicing activity of Argonaute proteins (AGOs)8,9. Recent studies have documented induction and suppression of antiviral RNAi in mouse embryonic stem cells and suckling mice10,11. However, it is still under debate whether infection by IAV or any other RNA virus that infects humans induces and/or suppresses antiviral RNAi in mature mammalian somatic cells1221. Here, we demonstrate that mature human somatic cells produce abundant virus-derived siRNAs co-immunoprecipitated with AGOs in response to IAV infection. We show that the biogenesis of viral siRNAs from IAV double-stranded RNA (dsRNA) precursors in infected cells is mediated by wild-type human Dicer and potently suppressed by both NS1 of IAV as well as virion protein 35 (VP35) of Ebola and Marburg filoviruses. We further demonstrate that the slicing catalytic activity of AGO2 inhibits IAV and other RNA viruses in mature mammalian cells, in an interferon-independent fashion. Altogether, our work shows that IAV infection induces and suppresses antiviral RNAi in differentiated mammalian somatic cells.

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Figure 1: Production of viral siRNAs in mature human somatic cells.
Figure 2: Wild-type (WT) hDicer is necessary and sufficient for the biogenesis of human vsiRNAs in differentiated somatic cells.
Figure 3: Induction and suppression of influenza vsiRNA biogenesis in distinct human and monkey somatic cells.
Figure 4: AGO2 slicing activity restricts IAV, EMCV and VSV in mammalian somatic cells.

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  • 14 July 2017

    In the PDF version of this article previously published, the year of publication provided in the footer of each page and in the 'How to cite' section was erroneously given as 2017, it should have been 2016. This error has now been corrected. The HTML version of the article was not affected.

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Acknowledgements

The authors thank L.A. Ball, C. Basler, B.R. Cullen, A. Garcia-Sastre, C. Rice, M. McDonald, K.L. Johnson, Q. Liu and P. Palese for providing materials, A. Tarakhovsky for scientific discussions and support, and F. Uhl and A.E. Handte-Reinecker for technical assistance. G. Hannon provided Ago2D587A MEF lines. This study was supported by NIH grants R01AI107087 (to K.L.J.), MGH Executive Committee on Research (ECOR) funds (to K.L.J.), R01AI52447 (to S.W.D.) and R56AI110579 (to S.W.D.), CNAS of UC Riverside (to S.W.D.), AI113333 and DK068181 (to H.C.R.), a Department of Defense PRCRP fellowship CA120212 (to S.C.) and NHMRC grants 1027020 and 1083596 (to P.H.).

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Contributions

Y.L. and S.D. performed all virus infection experiments in 293T, A549 and Vero cells. M.B. performed and analysed all virus infection experiments in Ago2D597A cells. J.L. performed all bioinformatic analyses of small RNA libraries. Y.H., W.-X.L. and F.V.K. assisted with cloning of small RNAs. D.A.C. and J.T.P. assisted with viral infections. H.C.R and P.H. provided reagents and interpreted results. S.C. provided Ago2D597A MEFs. S.W.D. and K.L.J conceived of the study, designed experiments, interpreted results and wrote the final manuscript.

Corresponding authors

Correspondence to Shou-Wei Ding or Kate L. Jeffrey.

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The authors declare no competing financial interests.

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Supplementary Tables 1 and 2, Supplementary Figures 1–11. (PDF 14391 kb)

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Li, Y., Basavappa, M., Lu, J. et al. Induction and suppression of antiviral RNA interference by influenza A virus in mammalian cells. Nat Microbiol 2, 16250 (2017). https://doi.org/10.1038/nmicrobiol.2016.250

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