1887

Abstract

Limitations of the current vaccines and antivirals against influenza A virus (IAV) pandemic underscore the urgent need for developing novel anti-influenza strategies. RNA interference (RNAi) induced by small interfering RNA (siRNA) has become a powerful new means to inhibit viral infection in a gene-specific manner. However, the efficacy of the siRNA delivery platform and the relatively high cost of administration have hindered widespread application of siRNA. In this study, we developed a microRNA (miRNA)-30-based lentivirus delivery system by embedding a synthetic short hairpin RNA (shRNA) stem into the context of endogenous precursor of miRNA-30 (shRNAmir) to express a silencer of the influenza gene. We showed that the miRNA-based lentivirus vector was able to express and process a single nucleoprotein (NP)-targeting shRNAmir, which could potently inhibit IAV replication. We further showed that miRNA-based lentivirus vector carrying tandemly linked NP and polymerase PB1 shRNAmirs could express and process double shRNAmirs. Despite the relatively low levels of NP and PB1 miRNAs produced in the stably transduced cells, the combination of two miRNAs exerted a great degree of inhibition on influenza infection. Given the advantage of combinatorial RNAi in preventing emergence of mutant virus, miRNA-based lentiviral vectors are valuable tools for anitiviral activities. To the best of our knowledge, this is the first study demonstrating that a miRNA-based RNAi strategy can be applied for better control of influenza virus infection.

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2015-10-01
2019-10-22
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References

  1. Boudreau R.L., Martins I., Davidson B.L.. ( 2009;). Artificial microRNAs as siRNA shuttles: improved safety as compared to shRNAs in vitro and in vivo. Mol Ther 17: 169–175 [CrossRef] [PubMed].
    [Google Scholar]
  2. Bouvier N.M., Palese P.. ( 2008;). The biology of influenza viruses. Vaccine 26: (Suppl. 4), D49–D53 [CrossRef] [PubMed].
    [Google Scholar]
  3. Bridge A.J., Pebernard S., Ducraux A., Nicoulaz A.-L., Iggo R.. ( 2003;). Induction of an interferon response by RNAi vectors in mammalian cells. Nat Genet 34: 263–264 [CrossRef] [PubMed].
    [Google Scholar]
  4. Castanotto D., Sakurai K., Lingeman R., Li H., Shively L., Aagaard L., Soifer H., Gatignol A., Riggs A., Rossi J.J.. ( 2007;). Combinatorial delivery of small interfering RNAs reduces RNAi efficacy by selective incorporation into RISC. Nucleic Acids Res 35: 5154–5164 [CrossRef] [PubMed].
    [Google Scholar]
  5. Chang K., Elledge S.J., Hannon G.J.. ( 2006;). Lessons from nature: microRNA-based shRNA libraries. Nat Methods 3: 707–714 [CrossRef] [PubMed].
    [Google Scholar]
  6. Chang K., Marran K., Valentine A., Hannon G.J.. ( 2013;). Creating an miR30-based shRNA vector. Cold Spring Harb Protoc 2013:. [CrossRef]
    [Google Scholar]
  7. Chi S.W., Hannon G.J., Darnell R.B.. ( 2012;). An alternative mode of microRNA target recognition. Nat Struct Mol Biol 19: 321–327 [CrossRef] [PubMed].
    [Google Scholar]
  8. Chung K.-H., Hart C.C., Al-Bassam S., Avery A., Taylor J., Patel P.D., Vojtek A.B., Turner D.L.. ( 2006;). Polycistronic RNA polymerase II expression vectors for RNA interference based on BIC/miR-155. Nucleic Acids Res 34: e53 [CrossRef] [PubMed].
    [Google Scholar]
  9. Cullen B.R.. ( 2004;). Transcription and processing of human microRNA precursors. Mol Cell 16: 861–865 [CrossRef] [PubMed].
    [Google Scholar]
  10. DeKelver R.C., Choi V.M., Moehle E.A., Paschon D.E., Hockemeyer D., Meijsing S.H., Sancak Y., Cui X., Steine E.J., other authors. ( 2010;). Functional genomics, proteomics, and regulatory DNA analysis in isogenic settings using zinc finger nuclease-driven transgenesis into a safe harbor locus in the human genome. Genome Res 20: 1133–1142 [CrossRef] [PubMed].
    [Google Scholar]
  11. Dykxhoorn D.M., Lieberman J.. ( 2006;). Silencing viral infection. PLoS Med 3: e242 [CrossRef] [PubMed].
    [Google Scholar]
  12. Fellmann C., Hoffmann T., Sridhar V., Hopfgartner B., Muhar M., Roth M., Lai D.Y., Barbosa I.A.M., Kwon J.S., other authors. ( 2013;). An optimized microRNA backbone for effective single-copy RNAi. Cell Reports 5: 1704–1713 [CrossRef] [PubMed].
    [Google Scholar]
  13. Ge Q., McManus M.T., Nguyen T., Shen C.-H., Sharp P.A., Eisen H.N., Chen J.. ( 2003;). RNA interference of influenza virus production by directly targeting mRNA for degradation and indirectly inhibiting all viral RNA transcription. Proc Natl Acad Sci U S A 100: 2718–2723 [CrossRef] [PubMed].
    [Google Scholar]
  14. Ge Q., Filip L., Bai A., Nguyen T., Eisen H.N., Chen J.. ( 2004;). Inhibition of influenza virus production in virus-infected mice by RNA interference. Proc Natl Acad Sci U S A 101: 8676–8681 [CrossRef] [PubMed].
    [Google Scholar]
  15. Gitlin L., Stone J.K., Andino R.. ( 2005;). Poliovirus escape from RNA interference: short interfering RNA: target recognition and implications for therapeutic approaches. J Virol 79: 1027–1035 [CrossRef] [PubMed].
    [Google Scholar]
  16. Grimm D., Kay M.A.. ( 2007;). Combinatorial RNAi: a winning strategy for the race against evolving targets?. Mol Ther 15: 878–888 [PubMed].
    [Google Scholar]
  17. Grimm D., Streetz K.L., Jopling C.L., Storm T.A., Pandey K., Davis C.R., Marion P., Salazar F., Kay M.A.. ( 2006;). Fatality in mice due to oversaturation of cellular microRNA/short hairpin RNA pathways. Nature 441: 537–541 [CrossRef] [PubMed].
    [Google Scholar]
  18. Grimson A., Farh K.K.-H., Johnston W.K., Garrett-Engele P., Lim L.P., Bartel D.P.. ( 2007;). MicroRNA targeting specificity in mammals: determinants beyond seed pairing. Mol Cell 27: 91–105 [CrossRef] [PubMed].
    [Google Scholar]
  19. Haasnoot J., Berkhout B.. ( 2006;). RNA interference: its use as antiviral therapy. . In RNA Towards Medicine (Handbook of Experimental Pharmacology)vol. 173, pp. 117–150 Berlin:: Springer;. [CrossRef]
    [Google Scholar]
  20. Haasnoot P.C., Cupac D., Berkhout B.. ( 2003;). Inhibition of virus replication by RNA interference. J Biomed Sci 10: 607–616 [CrossRef] [PubMed].
    [Google Scholar]
  21. Hannon G.J.. ( 2002;). RNA interference. Nature 418: 244–251 [CrossRef] [PubMed].
    [Google Scholar]
  22. Jackson A.L., Bartz S.R., Schelter J., Kobayashi S.V., Burchard J., Mao M., Li B., Cavet G., Linsley P.S.. ( 2003;). Expression profiling reveals off-target gene regulation by RNAi. Nat Biotechnol 21: 635–637 [CrossRef] [PubMed].
    [Google Scholar]
  23. Khan I.F., Hirata R.K., Russell D.W.. ( 2011;). AAV-mediated gene targeting methods for human cells. Nat Protoc 6: 482–501 [CrossRef] [PubMed].
    [Google Scholar]
  24. Kim V.N.. ( 2005;). MicroRNA biogenesis: coordinated cropping and dicing. Nat Rev Mol Cell Biol 6: 376–385 [CrossRef] [PubMed].
    [Google Scholar]
  25. Lee Y., Jeon K., Lee J.T., Kim S., Kim V.N.. ( 2002;). MicroRNA maturation: stepwise processing and subcellular localization. EMBO J 21: 4663–4670 [CrossRef] [PubMed].
    [Google Scholar]
  26. Li Y., Anderson D.H., Liu Q., Zhou Y.. ( 2008;). Mechanism of influenza A virus NS1 protein interaction with the p85β, but not the p85α, subunit of phosphatidylinositol 3-kinase (PI3K) and up-regulation of PI3K activity. J Biol Chem 283: 23397–23409 [CrossRef] [PubMed].
    [Google Scholar]
  27. Lin L., Liu Q., Berube N., Detmer S., Zhou Y.. ( 2012a;). 5′-Triphosphate-short interfering RNA: potent inhibition of influenza A virus infection by gene silencing and RIG-I activation. J Virol 86: 10359–10369 [CrossRef] [PubMed].
    [Google Scholar]
  28. Lin L., Li Y., Pyo H.-M., Lu X., Raman S.N.T., Liu Q., Brown E.G., Zhou Y.. ( 2012b;). Identification of RNA helicase A as a cellular factor that interacts with influenza A virus NS1 protein and its role in the virus life cycle. J Virol 86: 1942–1954 [CrossRef] [PubMed].
    [Google Scholar]
  29. Liu Y.P., Haasnoot J., ter Brake O., Berkhout B., Konstantinova P.. ( 2008;). Inhibition of HIV-1 by multiple siRNAs expressed from a single microRNA polycistron. Nucleic Acids Res 36: 2811–2824 [CrossRef] [PubMed].
    [Google Scholar]
  30. Liu Y.P., Berkhout B.. ( 2013;). Design of lentivirally expressed siRNAs. Methods Mol Biol 942: 233–257 [CrossRef] [PubMed].
    [Google Scholar]
  31. Martin H.C., Wani S., Steptoe A.L., Krishnan K., Nones K., Nourbakhsh E., Vlassov A., Grimmond S.M., Cloonan N.. ( 2014;). Imperfect centered miRNA binding sites are common and can mediate repression of target mRNAs. Genome Biol 15: R51 [CrossRef] [PubMed].
    [Google Scholar]
  32. McBride J.L., Boudreau R.L., Harper S.Q., Staber P.D., Monteys A.M., Martins I., Gilmore B.L., Burstein H., Peluso R.W., other authors. ( 2008;). Artificial miRNAs mitigate shRNA-mediated toxicity in the brain: implications for the therapeutic development of RNAi. Proc Natl Acad Sci U S A 105: 5868–5873 [CrossRef] [PubMed].
    [Google Scholar]
  33. Saladino R., Barontini M., Crucianelli M., Nencioni L., Sgarbanti R., Palamara A.T.. ( 2010;). Current advances in anti-influenza therapy. Curr Med Chem 17: 2101–2140 [CrossRef] [PubMed].
    [Google Scholar]
  34. Shin Y.-K., Liu Q., Tikoo S.K., Babiuk L.A., Zhou Y.. ( 2007a;). Influenza A virus NS1 protein activates the phosphatidylinositol 3-kinase (PI3K)/Akt pathway by direct interaction with the p85 subunit of PI3K. J Gen Virol 88: 13–18 [CrossRef] [PubMed].
    [Google Scholar]
  35. Shin Y.-K., Liu Q., Tikoo S.K., Babiuk L.A., Zhou Y.. ( 2007b;). Effect of the phosphatidylinositol 3-kinase/Akt pathway on influenza A virus propagation. J Gen Virol 88: 942–950 [CrossRef] [PubMed].
    [Google Scholar]
  36. Tiscornia G., Singer O., Ikawa M., Verma I.M.. ( 2003;). A general method for gene knockdown in mice by using lentiviral vectors expressing small interfering RNA. Proc Natl Acad Sci U S A 100: 1844–1848 [CrossRef] [PubMed].
    [Google Scholar]
  37. Tompkins S.M., Lo C.-Y., Tumpey T.M., Epstein S.L.. ( 2004;). Protection against lethal influenza virus challenge by RNA interference in vivo. Proc Natl Acad Sci U S A 101: 8682–8686 [CrossRef] [PubMed].
    [Google Scholar]
  38. Tumpey T.M., Belser J.A.. ( 2009;). Resurrected pandemic influenza viruses. Annu Rev Microbiol 63: 79–98 [CrossRef] [PubMed].
    [Google Scholar]
  39. Wilson J.A., Richardson C.D.. ( 2005;). Hepatitis C virus replicons escape RNA interference induced by a short interfering RNA directed against the NS5b coding region. J Virol 79: 7050–7058 [CrossRef] [PubMed].
    [Google Scholar]
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