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

Nature Structural & Molecular Biology volume 13, pages 793–797 (2006)Cite this article

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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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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Authors and Affiliations

  1. Graduate School of Biological Sciences, Beckman Research Institute of the City of Hope, 1450 E. Duarte Road, Duarte, 91010, California, USA

    Daniel H Kim, Louisa M Villeneuve & John J Rossi

  2. Department of Molecular and Experimental Medicine, The Scripps Research Institute, 10550 N. Torrey Pines Road, La Jolla, 92037, California, USA

    Kevin V Morris

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  1. Daniel H Kim
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  2. Louisa M Villeneuve
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Contributions

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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