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Argonaute-1 directs siRNA-mediated transcriptional gene silencing in human cells

Abstract

Argonaute proteins are the core components of effector complexes that facilitate RNA interference (RNAi). Small interfering RNAs (siRNAs) targeted to promoter regions mediate transcriptional gene silencing (TGS) in human cells through heterochromatin formation. RNAi effector complexes have yet to be implicated in the mechanism of mammalian TGS. Here we describe the role of the human Argonaute-1 homolog (AGO1) in directing TGS at the promoters for human immunodeficiency virus-1 coreceptor CCR5 and tumor suppressor RASSF1A. AGO1 associates with RNA polymerase II (RNAPII) and is required for histone H3 Lys9 dimethylation and TGS. AGO1, TAR RNA-binding protein-2 (7TRBP2) and Polycomb protein EZH2 colocalize to the siRNA-targeted RASSF1A promoter, implicating Polycomb silencing in the mechanism of mammalian TGS. These results establish a connection between RNAi components AGO1 and TRBP2, RNAPII transcription and Polycomb-regulated control of gene expression.

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Figure 1: Transcriptional gene silencing of CCR5-GFP and RASSF1A.
Figure 2: H3K9me2 and DNA methylation at targeted promoters.
Figure 3: AGO1 recruitment to siRNA-targeted promoters.
Figure 4: AGO1 is required for transcriptional gene silencing.
Figure 5: TRBP2 is required for transcriptional gene silencing.
Figure 6: RNAPII with unphosphorylated CTD associates with AGO1.
Figure 7: Polycomb connection to transcriptional gene silencing.

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References

  1. Zamore, P.D. & Haley, B. Ribo-gnome: the big world of small RNAs. Science 309, 1519–1524 (2005).

    Article  CAS  Google Scholar 

  2. Zilberman, D., Cao, X. & Jacobsen, S.E. ARGONAUTE4 control of locus-specific siRNA accumulation and DNA and histone methylation. Science 299, 716–719 (2003).

    Article  CAS  Google Scholar 

  3. Volpe, T.A. et al. Regulation of heterchromatic silencing and histone H3 lysine-9 methylation by RNAi. Science 297, 1833–1837 (2002).

    Article  CAS  Google Scholar 

  4. Verdel, A. et al. RNAi-mediated targeting of heterochromatin by the RITS complex. Science 303, 672–676 (2004).

    Article  CAS  Google Scholar 

  5. Morris, K.V., Chan, S.W., Jacobsen, S.E. & Looney, D.J. Small interfering RNA-induced transcriptional gene silencing in human cells. Science 305, 1289–1292 (2004).

    Article  CAS  Google Scholar 

  6. Weinberg, M.S. et al. The antisense strand of small interfering RNAs directs histone methylation and transcriptional gene silencing in human cells. RNA 12, 256–262 (2006).

    Article  CAS  Google Scholar 

  7. Chow, J.C., Yen, Z., Ziesche, S.M. & Brown, C.J. Silencing of the mammalian X chromosome. Annu. Rev. Genomics Hum. Genet. 6, 69–92 (2005).

    Article  CAS  Google Scholar 

  8. Moriuchi, H., Moriuchi, M. & Fauci, A.S. Cloning and analysis of the promoter region of CCR5, a coreceptor for HIV-1 entry. J. Immunol. 159, 5441–5449 (1997).

    CAS  PubMed  Google Scholar 

  9. Dammann, R. et al. Epigenetic inactivation of a RAS association domain family protein from the lung tumour suppressor locus 3p21.3. Nat. Genet. 25, 315–319 (2000).

    Article  CAS  Google Scholar 

  10. Ting, A.H., Schuebel, K.E., Herman, J.G. & Baylin, S.B. Short double-stranded RNA induces transcriptional gene silencing in human cancer cells in the absence of DNA methylation. Nat. Genet. 37, 906–910 (2005).

    Article  CAS  Google Scholar 

  11. Castanotto, D. et al. Short hairpin RNA-directed cytosine (CpG) methylation of the RASSF1A gene promoter in HeLa cells. Mol. Ther. 12, 179–183 (2005).

    Article  CAS  Google Scholar 

  12. Meister, G. et al. Human Argonaute2 mediates RNA cleavage targeted by miRNAs and siRNAs. Mol. Cell 15, 185–197 (2004).

    Article  CAS  Google Scholar 

  13. Gatignol, A., Buckler-White, A., Berkhout, B. & Jeang, K.T. Characterization of a human TAR RNA-binding protein that activates the HIV-1 LTR. Science 251, 1597–1600 (1991).

    Article  CAS  Google Scholar 

  14. Chendrimada, T.P. et al. TRBP recruits the Dicer complex to Ago2 for microRNA processing and gene silencing. Nature 436, 740–744 (2005).

    Article  CAS  Google Scholar 

  15. Kato, H. et al. RNA polymerase II is required for RNAi-dependent heterochromatin assembly. Science 309, 467–469 (2005).

    Article  CAS  Google Scholar 

  16. Pirrotta, V. Polycombing the genome: PcG, trxG, and chromatin silencing. Cell 93, 333–336 (1998).

    Article  CAS  Google Scholar 

  17. Cao, R. et al. Role of histone H3 lysine 27 methylation in Polycomb-group silencing. Science 298, 1039–1043 (2002).

    Article  CAS  Google Scholar 

  18. Kuzmichev, A., Nishioka, K., Erdjument-Bromage, H., Tempst, P. & Reinberg, D. Histone methyltransferase activity associated with a human multiprotein complex containing the Enhancer of Zeste protein. Genes Dev. 16, 2893–2905 (2002).

    Article  CAS  Google Scholar 

  19. Kirmizis, A. et al. Silencing of human polycomb target genes is associated with methylation of histone H3 Lys 27. Genes Dev. 18, 1592–1605 (2004).

    Article  CAS  Google Scholar 

  20. Bracken, A.P., Dietrich, N., Pasini, D., Hansen, K.H. & Helin, K. Genome-wide mapping of Polycomb target genes unravels their roles in cell fate transitions. Genes Dev. 20, 1123–1136 (2006).

    Article  CAS  Google Scholar 

  21. Dellino, G.I. et al. Polycomb silencing blocks transcription initiation. Mol. Cell 13, 887–893 (2004).

    Article  CAS  Google Scholar 

  22. Buhler, M., Verdel, A. & Moazed, D. Tethering RITS to a nascent transcript initiates RNAi- and heterochromatin-dependent gene silencing. Cell 125, 873–886 (2006).

    Article  CAS  Google Scholar 

  23. Daviet, L. et al. Analysis of a binding difference between the two dsRNA-binding domains in TRBP reveals the modular function of a KR-helix motif. Eur. J. Biochem. 267, 2419–2431 (2000).

    Article  CAS  Google Scholar 

  24. Grimaud, C. et al. RNAi components are required for nuclear clustering of Polycomb group response elements. Cell 124, 957–971 (2006).

    Article  CAS  Google Scholar 

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Acknowledgements

We thank S. Jacobsen, S. Grewal and R. Martienssen for discussions on TGS, J. Zhang, T. Hong, L. Aagaard and K. Sakurai for technical assistance, D. Castanotto and M. Kalkum for reagents, A. Gatignol (McGill University) for anti-TRBP, G. Pavlakis (National Cancer Institute) for pR5-GFPsg143, members of the Rossi laboratory for technical advice and support and H. Soifer, R. Lin, R. Natarajan and G. Pfeifer for helpful discussions. This work was supported by grants from the US National Institutes of Health to J.J.R. (HL07470 and AI42552) and K.V.M. (HL83473) and from the National Cancer Institute to J.J.R. and K.V.M. (CA33572).

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D.H.K., K.V.M. and J.J.R. designed the study, D.H.K. carried out most of the experiments, K.V.M. and L.M.V. generated 293T CCR5-GFP cells and performed siRNA screening and D.H.K, K.V.M. and J.J.R. interpreted data and wrote the manuscript.

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Correspondence to John J Rossi.

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The authors declare no competing financial interests.

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Kim, D., Villeneuve, L., Morris, K. et al. Argonaute-1 directs siRNA-mediated transcriptional gene silencing in human cells. Nat Struct Mol Biol 13, 793–797 (2006). https://doi.org/10.1038/nsmb1142

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