Abstract
MicroRNAs (miRNAs) are an abundant class of non-coding RNAs that are believed to be important in many biological processes through regulation of gene expression1,2,3. The precise molecular function of miRNAs in mammals is largely unknown and a better understanding will require loss-of-function studies in vivo. Here we show that a novel class of chemically engineered oligonucleotides, termed ‘antagomirs’, are efficient and specific silencers of endogenous miRNAs in mice. Intravenous administration of antagomirs against miR-16, miR-122, miR-192 and miR-194 resulted in a marked reduction of corresponding miRNA levels in liver, lung, kidney, heart, intestine, fat, skin, bone marrow, muscle, ovaries and adrenals. The silencing of endogenous miRNAs by this novel method is specific, efficient and long-lasting. The biological significance of silencing miRNAs with the use of antagomirs was studied for miR-122, an abundant liver-specific miRNA. Gene expression and bioinformatic analysis of messenger RNA from antagomir-treated animals revealed that the 3′ untranslated regions of upregulated genes are strongly enriched in miR-122 recognition motifs, whereas downregulated genes are depleted in these motifs. Analysis of the functional annotation of downregulated genes specifically predicted that cholesterol biosynthesis genes would be affected by miR-122, and plasma cholesterol measurements showed reduced levels in antagomir-122-treated mice. Our findings show that antagomirs are powerful tools to silence specific miRNAs in vivo and may represent a therapeutic strategy for silencing miRNAs in disease.
This is a preview of subscription content, access via your institution
Access options
Subscribe to this journal
Receive 51 print issues and online access
$199.00 per year
only $3.90 per issue
Buy this article
- Purchase on Springer Link
- Instant access to full article PDF
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
References
Ambros, V. The functions of animal microRNAs. Nature 431, 350–355 (2004)
Bartel, D. P. MicroRNAs: Genomics, biogenesis, mechanism, and function. Cell 116, 281–297 (2004)
He, L. & Hannon, G. J. MicroRNAs: Small RNAs with a big role in gene regulation. Nature Rev. Genet. 5, 522–531 (2004)
Zhao, Y., Samal, E. & Srivastava, D. Serum response factor regulates a muscle-specific microRNA that targets Hand2 during cardiogenesis. Nature 436, 214–220 (2005)
Chen, C. Z. & Lodish, H. F. MicroRNAs as regulators of mammalian hematopoiesis. Semin. Immunol. 17, 155–165 (2005)
Poy, M. N. et al. Pancreatic islet-specific microRNA regulates insulin secretion. Nature 432, 226–230 (2004)
Meister, G., Landthaler, M., Dorsett, Y. & Tuschl, T. Sequence-specific inhibition of microRNA- and siRNA-induced RNA silencing. RNA 10, 544–550 (2004)
Hutvagner, G., Simard, M. J., Mello, C. C. & Zamore, P. D. Sequence-specific inhibition of small RNA function. PLoS Biol. 2, E98 (2004)
Eis, P. S. et al. Accumulation of miR-155 and BIC RNA in human B cell lymphomas. Proc. Natl Acad. Sci. USA 102, 3627–3632 (2005)
Metzler, M., Wilda, M., Busch, K., Viehmann, S. & Borkhardt, A. High expression of precursor microRNA-155/BIC RNA in children with Burkitt lymphoma. Genes Chromosom. Cancer 39, 167–169 (2004)
He, L. et al. A microRNA polycistron as a potential human oncogene. Nature 435, 828–833 (2005)
Lu, J. et al. MicroRNA expression profiles classify human cancers. Nature 435, 834–838 (2005)
Soutschek, J. et al. Therapeutic silencing of an endogenous gene by systemic administration of modified siRNAs. Nature 432, 173–178 (2004)
Manoharan, M., Kesavan, V., & Rajeev, K. G. SiRNA's containing ribose substitutes to which lipophilic moieties may be attached. U.S. Pat. Appl. Publ. (2005), US 2005107325.
Chang, J. et al. miRNA-122, a mammalian liver-specific microRNA, is processed from hcr mRNA and may downregulate the high affinity cationic amino acid transporter CAT1. RNA Biol. 1, 106–113 (2004)
Tomari, Y. & Zamore, P. D. MicroRNA biogenesis: drosha can't cut it without a partner. Curr. Biol. 15, R61–R64 (2005)
Lim, L. P. et al. Microarray analysis shows that some microRNAs downregulate large numbers of target mRNAs. Nature 433, 769–773 (2005)
Bagga, S. et al. Regulation by let-7 and lin-4 miRNAs results in target mRNA degradation. Cell 122, 553–563 (2005)
Krek, A. et al. Combinatorial microRNA target prediction. Nature Genet. 37, 495–500 (2005)
Lewis, B. P., Burge, C. B. & Bartel, D. P. Conserved seed pairing, often flanked by adenosines, indicates that thousands of human genes are microRNA targets. Cell 120, 15–20 (2005)
John, B. et al. Human MicroRNA targets. PLoS Biol. 2, e363 (2004)
Cheng, A. M., Byrom, M. W., Shelton, J. & Ford, L. P. Antisense inhibition of human miRNAs and indications for an involvement of miRNA in cell growth and apoptosis. Nucleic Acids Res. 33, 1290–1297 (2005)
Jopling, C. L., Yi, M., Lancaster, A. M., Lemon, S. M. & Sarnow, P. Modulation of hepatitis C virus RNA abundance by a liver-specific microRNA. Science 309, 1577–1580 (2005)
Weiler, J., Hunziker, J. & Hall, J. Anti-miRNA oligonucleotides (AMOs): ammunition to target miRNAs implicated in human disease? Gene Therapy Advance online publication 29 September 2005; doi:10.1038/sj.gt.3302654.
Pruitt, K. D., Tatusova, T. & Maglott, D. R. NCBI Reference Sequence (RefSeq): a curated non-redundant sequence database of genomes, transcripts and proteins. Nucleic Acids Res. 33, D501–D504 (2005)
Acknowledgements
We thank J. Maraganore, V. Kotelianski and P. Sharp for discussion and suggestions. These studies were supported by NIH grants (to M.S., T.T. and N.R.), and an unrestricted grant from Bristol Myers Squibb (M.S.).
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Competing interests
M.S. serves on the Scientific Advisory Board and T.T. is a co-founder of Alnylam Pharmaceuticals, Inc. Some participating authors are employed by Alnylam Pharmaceuticals, Inc.
Supplementary information
Supplementary Figure 1
MiR-122 is quantitatively recovered from a duplex formed with antagomir-122. (PDF 64 kb)
Supplementary Figure 2
Annealing antagomir-122 to liver RNA in vitro does not alter miR-122 detection in formamide-containing gels. (PDF 126 kb)
Supplementary Figure 3
Sustained silencing of miR-122 after injection of antagomir-122 into mice. (PDF 217 kb)
Supplementary Figure 4
Antagomirs specifically target miRNAs that derive from the same primary transcript. (PDF 234 kb)
Supplementary Table 1
Affymetrix gene expression analysis of genes up-regulated ≥1.4-fold in livers of mice treated with antagomir-122 compared to control mm-antagomir-122. (XLS 123 kb)
Supplementary Table 2
Affymetrix gene expression analysis of genes down-regulated ≥1.4-fold in livers of mice treated with antagomir-122 compared to control mm-antagomir-122. Probe sets with "absent (A)" to "absent" changes were excluded. (XLS 98 kb)
Supplementary Notes
This contains Supplementary Figure Legends and Supplementary Notes. (DOC 24 kb)
Supplementary Methods
This file contains additional details of the methods used in this study. (DOC 33 kb)
Rights and permissions
About this article
Cite this article
Krützfeldt, J., Rajewsky, N., Braich, R. et al. Silencing of microRNAs in vivo with ‘antagomirs’. Nature 438, 685–689 (2005). https://doi.org/10.1038/nature04303
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1038/nature04303
This article is cited by
MicroRNA-218-5p-Ddx41 axis restrains microglia-mediated neuroinflammation through downregulating type I interferon response in a mouse model of Parkinson’s disease
Journal of Translational Medicine (2024)
MiR-181a protects the heart against myocardial infarction by regulating mitochondrial fission via targeting programmed cell death protein 4
Scientific Reports (2024)
Understanding molecular mechanisms and miRNA-based targets in diabetes foot ulcers
Molecular Biology Reports (2024)
Oligonucleotide therapeutics and their chemical modification strategies for clinical applications
Journal of Pharmaceutical Investigation (2024)
An engineered miRNA PS-OMe miR130 inhibits acute lung injury by targeting eCIRP in sepsis
Molecular Medicine (2023)
Comments
By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.