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Transcriptional Gene Silencing Using Small RNAs

  • Daniel H. Kim
  • John J. Rossi
Protocol
Part of the Methods in Molecular Biology™ book series (MIMB, volume 555)

Abstract

RNA interference is a potent gene silencing pathway initiated by short molecules of double-stranded RNA. Small interfering RNAs (siRNAs) with full sequence complementarity to mRNAs induce cleavage of their target transcripts in the cytoplasm. Recent evidence has shown, however, that siRNAs can also function in the nucleus of mammalian cells to affect changes in chromatin structure. When targeted to promoter regions, siRNAs load into the effector protein Argonaute-1 (AGO1) and direct the formation of silent chromatin domains. This mechanism is known as transcriptional gene silencing (TGS), and the development of TGS as a novel therapeutic modality would be applicable to chronic diseases where long-term, heritable silencing of target genes is warranted. Here we discuss how small RNAs can be used to direct TGS in mammalian cells.

Key words

RNAi siRNA microRNA TGS chromatin epigenetic methylation transcription 

Notes

Acknowledgments

This work was supported by a City of Hope Graduate School Internal Fellowship to D.H.K. and National Institutes of Health grants R37-AI29329 and R01-AI42552 to J.J.R.

References

  1. 1.
    Kim, D. H. and Rossi, J. J. (2007) Strategies for silencing human disease using RNA interference. Nat. Rev. Genet. 8, 173–184.PubMedCrossRefGoogle Scholar
  2. 2.
    Kim, D. H., Villeneuve, L. M., Morris, K. V., and Rossi, J. J. (2006) Argonaute-1 directs siRNA-mediated transcriptional gene silencing in human cells. Nat. Struct. Mol. Biol. 13, 793–797.PubMedCrossRefGoogle Scholar
  3. 3.
    Weinberg, M. S., Villeneuve, L. M., Ehsani, A., Amarzguioui, M., Aagaard, L., Chen, Z. X., Riggs, A. D., Rossi, J. J., and Morris, K. V. (2006) The antisense strand of small interfering RNAs directs histone methylation and transcriptional gene silencing in human cells. RNA 12, 256–262.PubMedCrossRefGoogle Scholar
  4. 4.
    Ting, A. H., Schuebel, K. E., Herman, J. G., and Baylin, S. B. (2005) Short double-stranded RNA induces transcriptional gene silencing in human cancer cells in the absence of DNA methylation. Nat. Genet. 37, 906–910.PubMedCrossRefGoogle Scholar
  5. 5.
    Janowski, B. A., Huffman, K. E., Schwartz, J. C., Ram, R., Hardy, D., Shames, D. S., Minna, J. D., and Corey, D. R. (2005) Inhibiting gene expression at transcription start sites in chromosomal DNA with antigene RNAs. Nat. Chem. Biol. 1, 216–222.PubMedCrossRefGoogle Scholar
  6. 6.
    Pulukuri, S. M. and Rao, J. S. (2007) Small interfering RNA directed reversal of urokinase plasminogen activator demethylation inhibits prostate tumor growth and metastasis. Cancer Res. 67, 6637–6646.PubMedCrossRefGoogle Scholar
  7. 7.
    Han, J., Kim, D., and Morris, K. V. (2007) Promoter-associated RNA is required for RNA-directed transcriptional gene silencing in human cells. Proc. Natl. Acad. Sci. USA 104, 12422–12427.PubMedCrossRefGoogle Scholar
  8. 8.
    Zhang, M. X., et al. (2005) Regulation of endothelial nitric oxide synthase by small RNA. Proc. Natl. Acad. Sci. USA 102, 16967–16972.PubMedCrossRefGoogle Scholar

Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2009

Authors and Affiliations

  • Daniel H. Kim
    • 1
  • John J. Rossi
    • 2
  1. 1.Howard Hughes Medical Institute, Department of Molecular Biology, Massachusetts General Hospital and Department of GeneticsHarvard Medical SchoolBostonUSA
  2. 2.Graduate School of Biological Sciences and Division of Molecular BiologyBeckman Research Institute of the City of HopeDuarteUSA

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