Small dsRNAs induce transcriptional activation in human cells.
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Journal: 2007/January - Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424
Abstract:
Recent studies have shown that small noncoding RNAs, such as microRNAs and siRNAs, regulate gene expression at multiple levels including chromatin architecture, transcription, RNA editing, RNA stability, and translation. Each form of RNA-dependent regulation has been generally found to silence homologous sequences and collectively called RNAi. To further study the regulatory role of small RNAs at the transcriptional level, we designed and synthesized 21-nt dsRNAs targeting selected promoter regions of human genes E-cadherin, p21(WAF1/CIP1) (p21), and VEGF. Surprisingly, transfection of these dsRNAs into human cell lines caused long-lasting and sequence-specific induction of targeted genes. dsRNA mutation studies reveal that the 5' end of the antisense strand, or "seed" sequence, is critical for activity. Mechanistically, the dsRNA-induced gene activation requires the Argonaute 2 (Ago2) protein and is associated with a loss of lysine-9 methylation on histone 3 at dsRNA-target sites. In conclusion, we have identified several dsRNAs that activate gene expression by targeting noncoding regulatory regions in gene promoters. These findings reveal a more diverse role for small RNA molecules in the regulation of gene expression than previously recognized and identify a potential therapeutic use for dsRNA in targeted gene activation.
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Proc Natl Acad Sci U S A 103(46): 17337-17342
PMID: 17085592

Small dsRNAs induce transcriptional activation in human cells

Department of Urology, Veterans Affairs Medical Center and University of California, San Francisco, CA 94121
*To whom correspondence may be addressed at: Urology Research Center, Veterans Affairs Medical Center and University of California, 4150 Clement Street, San Francisco, CA 94121., E-mail: ude.fscu@il.gnehcgnol or ude.fscu.ygoloru@ayihadr
Edited by Mark T. Groudine, Fred Hutchinson Cancer Research Center, Seattle, WA, and approved September 28, 2006

Author contributions: L.-C.L. designed research; L.-C.L., S.T.O., H.Z., D.P., R.F.P., S.U., and H.E. performed research; L.-C.L. and R.F.P. analyzed data; and L.-C.L., R.F.P., and R.D. wrote the paper.

Edited by Mark T. Groudine, Fred Hutchinson Cancer Research Center, Seattle, WA, and approved September 28, 2006
Received 2006 Aug 15
Copyright © 2006 by The National Academy of Sciences of the USA

Abstract

Recent studies have shown that small noncoding RNAs, such as microRNAs and siRNAs, regulate gene expression at multiple levels including chromatin architecture, transcription, RNA editing, RNA stability, and translation. Each form of RNA-dependent regulation has been generally found to silence homologous sequences and collectively called RNAi. To further study the regulatory role of small RNAs at the transcriptional level, we designed and synthesized 21-nt dsRNAs targeting selected promoter regions of human genes E-cadherin, p21 (p21), and VEGF. Surprisingly, transfection of these dsRNAs into human cell lines caused long-lasting and sequence-specific induction of targeted genes. dsRNA mutation studies reveal that the 5′ end of the antisense strand, or “seed” sequence, is critical for activity. Mechanistically, the dsRNA-induced gene activation requires the Argonaute 2 (Ago2) protein and is associated with a loss of lysine-9 methylation on histone 3 at dsRNA-target sites. In conclusion, we have identified several dsRNAs that activate gene expression by targeting noncoding regulatory regions in gene promoters. These findings reveal a more diverse role for small RNA molecules in the regulation of gene expression than previously recognized and identify a potential therapeutic use for dsRNA in targeted gene activation.

Keywords: gene regulation, promoter, Argonaute 2
Abstract

Small dsRNAs were initially discovered as the trigger of RNAi, a mechanism by which homologous mRNA is degraded to result in posttranscriptional gene silencing (1, 2). dsRNA is also involved in transcriptional gene silencing by directing DNA methylation in plants (3, 4) and heterochromatin formation in fission yeast (5) and Drosophila (6). Only recently has transcriptional gene silencing been discovered to occur in mammals (7, 8). There are, however, a few cases in which small RNAs positively regulate cognate sequences. For example, a small RNA isolated from neural stem cells can activate the transcription of genes containing NRSE/RE1 sequences to stimulate neuronal differentiation in adult stem cells (9). In addition, a liver-specific microRNA was found to enhance viral replication by targeting the 5′ noncoding region of the viral genome (10). More than three decades ago, Britten and Davidson (11) proposed a theory in which so-called “activator” RNAs, transcribed from redundant genomic regions, activate a battery of protein coding genes. To further study the potential role that small RNAs have in gene transcription, we selectively targeted promoter regions with synthetic dsRNAs and identified several dsRNAs that readily activate gene expression at a transcriptional level.

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Acknowledgments

We thank G. R. Deng, R. Erickson, and K. Greene for critical reading of the manuscript and Y. Shi for helpful discussion. This work was supported by National Institutes of Health Grants R01 AG21418 and R01 CA1018447 and Veterans Affairs Research Enhancement Award Program and Merit Review grants.

Acknowledgments

Abbreviations

AgoArgonaute
Aza-C5-azacytidine
RNAadsRNA-induced gene activation.
Abbreviations

Footnotes

The authors declare no conflict of interest.

This article is a PNAS direct submission.

Footnotes

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