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Asymmetric RNA duplexes mediate RNA interference in mammalian cells

Nature Biotechnology volume 26, pages 1379–1382 (2008)Cite this article

An Erratum to this article was published on 01 February 2009

This article has been updated

Abstract

RNA interference (RNAi) has become an indispensable technology for biomedical research and has demonstrated the potential to become a new class of therapeutic1,2,3. Current RNAi technology in mammalian cells relies on short interfering RNA (siRNA) consisting of symmetrical duplexes of 19–21 base pairs (bp) with 3′ overhangs4,5. Here we report that asymmetric RNA duplexes with 3′ and 5′ antisense overhangs silence mammalian genes effectively. An asymmetric interfering RNA (aiRNA) of 15 bp was incorporated into the RNA-induced silencing complex (RISC) and mediated sequence-specific cleavage of the target mRNA between base 10 and 11 relative to the 5′ end of the antisense strand. The gene silencing mediated by aiRNA was efficacious, durable and correlated with reduced off-target silencing by the sense strand. These results establish aiRNA as a scaffold structure for designing RNA duplexes to induce RNAi in mammalian cells.

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Figure 1: aiRNA induces efficacious and rapid gene silencing in mammalian cells.
Figure 2: The mechanism of gene silencing triggered by aiRNA.
Figure 3: aiRNA induces efficacious and durable gene silencing against multiple mammalian genes.
Figure 4: Off-target silencing by aiRNA and siRNA.

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  • 09 February 2009

    In the version of this article initially published, on page 1379, column 2, line 7, a parenthesis was missplaced. “…the passenger (often the sense strand)” should have read, “…the passenger (often the sense) strand”. The error has been corrected in the HTML and PDF versions of the article.

References

  1. de Fougerolles, A., Vornlocher, H.P., Maraganore, J. & Lieberman, J. Interfering with disease: a progress report on siRNA-based therapeutics. Nat. Rev. Drug Discov. 6, 443–453 (2007).

    Article  CAS  Google Scholar 

  2. Fire, A. et al. Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature 391, 806–811 (1998).

    Article  CAS  Google Scholar 

  3. Grimm, D. & Kay, M.A. Therapeutic application of RNAi: is mRNA targeting finally ready for prime time? J. Clin. Invest. 117, 3633–3641 (2007).

    Article  CAS  Google Scholar 

  4. Elbashir, S.M. et al. Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells. Nature 411, 494–498 (2001).

    Article  CAS  Google Scholar 

  5. Zamore, P.D., Tuschl, T., Sharp, P.A. & Bartel, D.P. RNAi: double-stranded RNA directs the ATP-dependent cleavage of mRNA at 21 to 23 nucleotide intervals. Cell 101, 25–33 (2000).

    Article  CAS  Google Scholar 

  6. Elbashir, S.M., Martinez, J., Patkaniowska, A., Lendeckel, W. & Tuschl, T. Functional anatomy of siRNAs for mediating efficient RNAi in Drosophila melanogaster embryo lysate. EMBO J. 20, 6877–6888 (2001).

    Article  CAS  Google Scholar 

  7. Matranga, C., Tomari, Y., Shin, C., Bartel, D.P. & Zamore, P.D. Passenger-strand cleavage facilitates assembly of siRNA into Ago2-containing RNAi enzyme complexes. Cell 123, 607–620 (2005).

    Article  CAS  Google Scholar 

  8. Rand, T.A., Petersen, S., Du, F. & Wang, X. Argonaute2 cleaves the anti-guide strand of siRNA during RISC activation. Cell 123, 621–629 (2005).

    Article  CAS  Google Scholar 

  9. Kim, D.H. et al. Synthetic dsRNA Dicer substrates enhance RNAi potency and efficacy. Nat. Biotechnol. 23, 222–226 (2005).

    Article  CAS  Google Scholar 

  10. Siolas, D. et al. Synthetic shRNAs as potent RNAi triggers. Nat. Biotechnol. 23, 227–231 (2005).

    Article  CAS  Google Scholar 

  11. Hohjoh, H. Enhancement of RNAi activity by improved siRNA duplexes. FEBS Lett. 557, 193–198 (2004).

    Article  CAS  Google Scholar 

  12. Vermeulen, A. et al. The contributions of dsRNA structure to Dicer specificity and efficiency. RNA 11, 674–682 (2005).

    Article  CAS  Google Scholar 

  13. Sano, M. et al. Effect of asymmetric terminal structures of short RNA duplexes on the RNA interference activity and strand selection. Nucleic Acids Res. 36, 5812–5821 (2008).

    Article  CAS  Google Scholar 

  14. Bolcato-Bellemin, A.L., Bonnet, M.E., Creusat, G., Erbacher, P. & Behr, J.P. Sticky overhangs enhance siRNA-mediated gene silencing. Proc. Natl. Acad. Sci. USA 104, 16050–16055 (2007).

    Article  CAS  Google Scholar 

  15. Ui-Tei, K. et al. Functional dissection of siRNA sequence by systematic DNA substitution: modified siRNA with a DNA seed arm is a powerful tool for mammalian gene silencing with significantly reduced off-target effect. Nucleic Acids Res. 36, 2136–2151 (2008).

    Article  CAS  Google Scholar 

  16. Chen, P.Y. et al. Strand-specific 5′-O-methylation of siRNA duplexes controls guide strand selection and targeting specificity. RNA 14, 263–274 (2008).

    Article  CAS  Google Scholar 

  17. Clark, P.R., Pober, J.S. & Kluger, M.S. Knockdown of TNFR1 by the sense strand of an ICAM-1 siRNA: dissection of an off-target effect. Nucleic Acids Res. 36, 1081–1097 (2008).

    Article  CAS  Google Scholar 

  18. Jackson, A.L. et al. Expression profiling reveals off-target gene regulation by RNAi. Nat. Biotechnol. 21, 635–637 (2003).

    Article  CAS  Google Scholar 

  19. Scacheri, P.C. et al. Short interfering RNAs can induce unexpected and divergent changes in the levels of untargeted proteins in mammalian cells. Proc. Natl. Acad. Sci. USA 101, 1892–1897 (2004).

    Article  CAS  Google Scholar 

  20. Tschuch, C. et al. Off-target effects of siRNA specific for GFP. BMC Mol. Biol. 9, 60 (2008).

    Article  Google Scholar 

  21. Sledz, C.A., Holko, M., de Veer, M.J., Silverman, R.H. & Williams, B.R. Activation of the interferon system by short-interfering RNAs. Nat. Cell Biol. 5, 834–839 (2003).

    Article  CAS  Google Scholar 

  22. Xiang, S., Fruehauf, J. & Li, C.J. Short hairpin RNA-expressing bacteria elicit RNA interference in mammals. Nat. Biotechnol. 24, 697–702 (2006).

    Article  CAS  Google Scholar 

  23. Khvorova, A., Reynolds, A. & Jayasena, S.D. Functional siRNAs and miRNAs exhibit strand bias. Cell 115, 209–216 (2003).

    Article  CAS  Google Scholar 

  24. Liu, J. et al. Argonaute2 is the catalytic engine of mammalian RNAi. Science 305, 1437–1441 (2004).

    Article  CAS  Google Scholar 

  25. Schwarz, D.S. et al. Asymmetry in the assembly of the RNAi enzyme complex. Cell 115, 199–208 (2003).

    Article  CAS  Google Scholar 

  26. Lin, X. et al. siRNA-mediated off-target gene silencing triggered by a 7 nt complementation. Nucleic Acids Res. 33, 4527–4535 (2005).

    Article  CAS  Google Scholar 

  27. Chu, C.Y. & Rana, T.M. Potent RNAi by short RNA triggers. RNA 14, 1714–1719 (2008).

    Article  CAS  Google Scholar 

  28. Iorns, E., Lord, C.J., Turner, N. & Ashworth, A. Utilizing RNA interference to enhance cancer drug discovery. Nat. Rev. Drug Discov. 6, 556–568 (2007).

    Article  CAS  Google Scholar 

  29. Corey, D.R. Chemical modification: the key to clinical application of RNA interference? J. Clin. Invest. 117, 3615–3622 (2007).

    Article  CAS  Google Scholar 

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Acknowledgements

We thank David Leggett, Youzhi Li, Zhiwei Jiang, Wei Li, Keith Mikule and other members of the research group at Boston Biomedical, Inc. for their advice and discussion, and Arthur B. Pardee, Judy Lieberman and Andrew Keates for critical reading of the manuscript.

Author information

Author notes

  1. Xiangao Sun and Harry A Rogoff: These authors contributed equally to this work.

  2. cli@bostonbiomedical.com

Authors and Affiliations

  1. Department of Therapeutic Biology, Boston Biomedical, Inc., 333 Providence Highway, Norwood, 02062, Massachusetts, USA

    Xiangao Sun, Harry A Rogoff & Chiang J Li

  2. Division of Gastroenterology, Skip Ackerman Center for Molecular Therapeutics, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, 02215, Massachusetts, USA

    Chiang J Li

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  1. Xiangao Sun
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  2. Harry A Rogoff
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Contributions

C.J.L. contributed to the idea, conception and overall experimental design. X.S. and H.A.R. contributed equally to the design and execution of all experiments.

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Competing interests

X.S., H.A.R. and C.J.L. are shareholders of Boston Biomedical, Inc.

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Sun, X., Rogoff, H. & Li, C. Asymmetric RNA duplexes mediate RNA interference in mammalian cells. Nat Biotechnol 26, 1379–1382 (2008). https://doi.org/10.1038/nbt.1512

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