Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Perspective
  • Published:

Desperately seeking microRNA targets

Abstract

MicroRNAs (miRNAs) suppress gene expression by inhibiting translation, promoting mRNA decay or both. Each miRNA may regulate hundreds of genes to control the cell's response to developmental and other environmental cues. The best way to understand the function of a miRNA is to identify the genes that it regulates. Target gene identification is challenging because miRNAs bind to their target mRNAs by partial complementarity over a short sequence, suppression of an individual target gene is often small, and the rules of targeting are not completely understood. Here we review computational and experimental approaches to the identification of miRNA-regulated genes. The examination of changes in gene expression that occur when miRNA expression is altered and biochemical isolation of miRNA-associated transcripts complement target prediction algorithms. Bioinformatic analysis of over-represented pathways and nodes in protein-DNA interactomes formed from experimental candidate miRNA gene target lists can focus attention on biologically significant target genes.

This is a preview of subscription content, access via your institution

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Methods for identifying miRNA targets.

Similar content being viewed by others

References

  1. Ghildiyal, M. & Zamore, P.D. Small silencing RNAs: an expanding universe. Nat. Rev. Genet. 10, 94–108 (2009).

    Article  CAS  Google Scholar 

  2. Davis, B. & Hata, A. Regulation of microRNA biogenesis: a miRiad of mechanisms. Cell Commun. Signal. 7, 18 (2009).

    Article  Google Scholar 

  3. Winter, J., Jung, S., Keller, S., Gregory, R.I. & Diederichs, S. Many roads to maturity: microRNA biogenesis pathways and their regulation. Nat. Cell Biol. 11, 228–234 (2009).

    Article  CAS  Google Scholar 

  4. Kim, V.N., Han, J. & Siomi, M.C. Biogenesis of small RNAs in animals. Nat. Rev. Mol. Cell Biol. 10, 126–139 (2009).

    Article  CAS  Google Scholar 

  5. Bartel, D.P. MicroRNAs: target recognition and regulatory functions. Cell 136, 215–233 (2009).

    Article  CAS  Google Scholar 

  6. Hendrickson, D.G. et al. Concordant regulation of translation and mRNA abundance for hundreds of targets of a human microRNA. PLoS Biol. 7, e1000238 (2009).

    Article  Google Scholar 

  7. Brodersen, P. & Voinnet, O. Revisiting the principles of microRNA target recognition and mode of action. Nat. Rev. Mol. Cell Biol. 10, 141–148 (2009).

    Article  CAS  Google Scholar 

  8. Friedman, R.C., Farh, K.K., Burge, C.B. & Bartel, D.P. Most mammalian mRNAs are conserved targets of microRNAs. Genome Res. 19, 92–105 (2009).

    Article  CAS  Google Scholar 

  9. Krek, A. et al. Combinatorial microRNA target predictions. Nat. Genet. 37, 495–500 (2005).

    Article  CAS  Google Scholar 

  10. 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).

    Article  CAS  Google Scholar 

  11. Lewis, B.P., Shih, I., Jones-Rhoades, M.W., Bartel, D.P. & Burge, C.B. Prediction of mammalian microRNA targets. Cell 115, 787–798 (2003).

    Article  CAS  Google Scholar 

  12. John, B. et al. Human microRNA targets. PLoS Biol. 2, e363 (2004).

    Article  Google Scholar 

  13. Miranda, K.C. et al. A pattern-based method for the identification of microRNA binding sites and their corresponding heteroduplexes. Cell 126, 1203–1217 (2006).

    Article  CAS  Google Scholar 

  14. Rajewsky, N. microRNA target predictions in animals. Nat. Genet. 38 (suppl.), S8–S13 (2006).

    Article  CAS  Google Scholar 

  15. Alexiou, P., Maragkakis, M., Papadopoulos, G.L., Reczko, M. & Hatzigeorgiou, A.G. Lost in translation: an assessment and perspective for computational microRNA target identification. Bioinformatics 25, 3049–3055 (2009).

    Article  CAS  Google Scholar 

  16. Johnson, S.M. et al. RAS is regulated by the let-7 microRNA family. Cell 120, 635–647 (2005).

    Article  CAS  Google Scholar 

  17. Lal, A. et al. miR-24 inhibits cell proliferation by targeting E2F2, MYC, and other cell-cycle genes via binding to “seedless” 3′UTR microRNA recognition elements. Mol. Cell 35, 610–625 (2009).

    Article  CAS  Google Scholar 

  18. Shin, C. et al. Expanding the microRNA targeting code: functional sites with centered pairing. Mol. Cell 38, 789–802 (2010).

    Article  CAS  Google Scholar 

  19. Tay, Y.M. et al. MicroRNA-134 modulates the differentiation of mouse embryonic stem cells, where it causes post-transcriptional attenuation of Nanog and LRH1. Stem Cells 26, 17–29 (2008).

    Article  CAS  Google Scholar 

  20. Tay, Y., Zhang, J., Thomson, A.M., Lim, B. & Rigoutsos, I. MicroRNAs to Nanog, Oct4 and Sox2 coding regions modulate embryonic stem cell differentiation. Nature 455, 1124–1128 (2008).

    Article  CAS  Google Scholar 

  21. Baek, D. et al. The impact of microRNAs on protein output. Nature 455, 64–71 (2008).

    Article  CAS  Google Scholar 

  22. Selbach, M. et al. Widespread changes in protein synthesis induced by microRNAs. Nature 455, 58–63 (2008).

    Article  CAS  Google Scholar 

  23. Lim, L.P. et al. Microarray analysis shows that some microRNAs downregulate large numbers of target mRNAs. Nature 433, 769–773 (2005).

    Article  CAS  Google Scholar 

  24. Johnson, C.D. et al. The let-7 MicroRNA represses cell proliferation pathways in human cells. Cancer Res. 67, 7713–7722 (2007).

    Article  CAS  Google Scholar 

  25. Chang, T. et al. Transactivation of miR-34a by p53 broadly influences gene expression and promotes apoptosis. Mol. Cell 26, 745–752 (2007).

    Article  CAS  Google Scholar 

  26. Zhang, L. et al. Systematic identification of C. elegans miRISC proteins, miRNAs, and mRNA targets by their interactions with GW182 proteins AIN-1 and AIN-2. Mol. Cell 28, 598–613 (2007).

    Article  CAS  Google Scholar 

  27. Easow, G., Teleman, A.A. & Cohen, S.M. Isolation of microRNA targets by miRNP immunopurification. RNA 13, 1198–1204 (2007).

    Article  CAS  Google Scholar 

  28. Hong, X., Hammell, M., Ambros, V. & Cohen, S.M. Immunopurification of Ago1 miRNPs selects for a distinct class of microRNA targets. Proc. Natl. Acad. Sci. USA 106, 15085–15090 (2009).

    Article  CAS  Google Scholar 

  29. Karginov, F.V. et al. A biochemical approach to identifying microRNA targets. Proc. Natl. Acad. Sci. USA 104, 19291–19296 (2007).

    Article  CAS  Google Scholar 

  30. Hendrickson, D.G., Hogan, D.J., Herschlag, D., Ferrell, J.E. & Brown, P.O. Systematic identification of mRNAs recruited to argonaute 2 by specific microRNAs and corresponding changes in transcript abundance. PLoS ONE 3, e2126 (2008).

    Article  Google Scholar 

  31. Beitzinger, M., Peters, L., Zhu, J.Y., Kremmer, E. & Meister, G. Identification of human microRNA targets from isolated argonaute protein complexes. RNA Biol. 4, 76–84 (2007).

    Article  CAS  Google Scholar 

  32. Zisoulis, D.G. et al. Comprehensive discovery of endogenous Argonaute binding sites in Caenorhabditis elegans. Nat. Struct. Mol. Biol. 17, 173–179 (2010).

    Article  CAS  Google Scholar 

  33. Chi, S.W., Zang, J.B., Mele, A. & Darnell, R.B. Argonaute HITS-CLIP decodes microRNA-mRNA interaction maps. Nature 460, 479–486 (2009).

    Article  CAS  Google Scholar 

  34. Hafner, M. et al. Transcriptome-wide identification of RNA-binding protein and microRNA target sites by PAR-CLIP. Cell 141, 129–141 (2010).

    Article  CAS  Google Scholar 

  35. Hammell, M. Computational methods to identify miRNA targets. Semin. Cell Dev. Biol. 21, 738–744 (2010).

    Article  CAS  Google Scholar 

  36. Mayr, C., Hemann, M.T. & Bartel, D.P. Disrupting the pairing between let-7 and Hmga2 enhances oncogenic transformation. Science 315, 1576–1579 (2007).

    Article  CAS  Google Scholar 

  37. Meng, F. et al. The microRNA let-7a modulates interleukin-6-dependent STAT-3 survival signaling in malignant human cholangiocytes. J. Biol. Chem. 282, 8256–8264 (2007).

    Article  CAS  Google Scholar 

  38. Duursma, A.M., Kedde, M., Schrier, M., le Sage, C. & Agami, R. miR-148 targets human DNMT3b protein coding region. RNA 14, 872–877 (2008).

    Article  CAS  Google Scholar 

  39. Grimson, A. et al. MicroRNA targeting specificity in mammals: determinants beyond seed pairing. Mol. Cell 27, 91–105 (2007).

    Article  CAS  Google Scholar 

  40. Betel, D., Wilson, M., Gabow, A., Marks, D.S. & Sander, C. The microRNA.org resource: targets and expression. Nucleic Acids Res. 36, D149–D153 (2008).

    Article  CAS  Google Scholar 

  41. Vella, M.C., Choi, E., Lin, S., Reinert, K. & Slack, F.J. The C. elegans microRNA let-7 binds to imperfect let-7 complementary sites from the lin-41 3′UTR. Genes Dev. 18, 132–137 (2004).

    Article  CAS  Google Scholar 

  42. Kertesz, M., Iovino, N., Unnerstall, U., Gaul, U. & Segal, E. The role of site accessibility in microRNA target recognition. Nat. Genet. 39, 1278–1284 (2007).

    Article  CAS  Google Scholar 

  43. Hammell, M. et al. mirWIP: microRNA target prediction based on microRNA-containing ribonucleoprotein-enriched transcripts. Nat. Methods 5, 813–819 (2008).

    Article  CAS  Google Scholar 

  44. Navarro, F. et al. miR-34a contributes to megakaryocytic differentiation of K562 cells independently of p53. Blood 114, 2181–2192 (2009).

    Article  CAS  Google Scholar 

  45. Sethupathy, P., Megraw, M. & Hatzigeorgiou, A.G. A guide through present computational approaches for the identification of mammalian microRNA targets. Nat. Methods 3, 881–886 (2006).

    Article  CAS  Google Scholar 

  46. Ritchie, W., Flamant, S. & Rasko, J.E.J. Predicting microRNA targets and functions: traps for the unwary. Nat. Methods 6, 397–398 (2009).

    Article  CAS  Google Scholar 

  47. Sandberg, R., Neilson, J.R., Sarma, A., Sharp, P.A. & Burge, C.B. Proliferating cells express mRNAs with shortened 3′ untranslated regions and fewer microRNA target sites. Science 320, 1643–1647 (2008).

    Article  CAS  Google Scholar 

  48. Mayr, C. & Bartel, D.P. Widespread shortening of 3′UTRs by alternative cleavage and polyadenylation activates oncogenes in cancer cells. Cell 138, 673–684 (2009).

    Article  CAS  Google Scholar 

  49. Ji, Z., Lee, J.Y., Pan, Z., Jiang, B. & Tian, B. Progressive lengthening of 3′ untranslated regions of mRNAs by alternative polyadenylation during mouse embryonic development. Proc. Natl. Acad. Sci. USA 106, 7028–7033 (2009).

    Article  CAS  Google Scholar 

  50. Wang, E.T. et al. Alternative isoform regulation in human tissue transcriptomes. Nature 456, 470–476 (2008).

    Article  CAS  Google Scholar 

  51. Ambros, V. The functions of animal microRNAs. Nature 431, 350–355 (2004).

    Article  CAS  Google Scholar 

  52. Wightman, B., Ha, I. & Ruvkun, G. Posttranscriptional regulation of the heterochronic gene lin-14 by lin-4 mediates temporal pattern formation in C. elegans. Cell 75, 855–862 (1993).

    Article  CAS  Google Scholar 

  53. Lee, R.C., Feinbaum, R.L. & Ambros, V. The C. elegans heterochronic gene lin-4 encodes small RNAs with antisense complementarity to lin-14. Cell 75, 843–854 (1993).

    Article  CAS  Google Scholar 

  54. Mavrakis, K.J. et al. Genome-wide RNA-mediated interference screen identifies miR-19 targets in Notch-induced T-cell acute lymphoblastic leukaemia. Nat. Cell Biol. 12, 372–379 (2010).

    Article  CAS  Google Scholar 

  55. Mu, P. et al. Genetic dissection of the miR-17:92 cluster of microRNAs in Myc-induced B-cell lymphomas. Genes Dev. 23, 2806–2811 (2009).

    Article  CAS  Google Scholar 

  56. Olive, V. et al. miR-19 is a key oncogenic component of mir-17–92. Genes Dev. 23, 2839–2849 (2009).

    Article  CAS  Google Scholar 

  57. Xin, M. et al. MicroRNAs miR-143 and miR-145 modulate cytoskeletal dynamics and responsiveness of smooth muscle cells to injury. Genes Dev. 23, 2166–2178 (2009).

    Article  CAS  Google Scholar 

  58. Elia, L. et al. The knockout of miR-143 and -145 alters smooth muscle cell maintenance and vascular homeostasis in mice: correlates with human disease. Cell Death Differ. 16, 1590–1598 (2009).

    Article  CAS  Google Scholar 

  59. Linsley, P.S. et al. Transcripts targeted by the microRNA-16 family cooperatively regulate cell cycle progression. Mol. Cell. Biol. 27, 2240–2252 (2007).

    Article  CAS  Google Scholar 

  60. Krützfeldt, J. et al. Silencing of microRNAs in vivo with 'antagomirs'. Nature 438, 685–689 (2005).

    Article  Google Scholar 

  61. Khan, A.A. et al. Transfection of small RNAs globally perturbs gene regulation by endogenous microRNAs. Nat. Biotechnol. 27, 549–555 (2009).

    Article  CAS  Google Scholar 

  62. Landthaler, M. et al. Molecular characterization of human Argonaute-containing ribonucleoprotein complexes and their bound target mRNAs. RNA 14, 2580–2596 (2008).

    Article  CAS  Google Scholar 

  63. Ding, L. & Han, M. GW182 family proteins are crucial for microRNA-mediated gene silencing. Trends Cell Biol. 17, 411–416 (2007).

    Article  CAS  Google Scholar 

  64. Su, H., Trombly, M.I., Chen, J. & Wang, X. Essential and overlapping functions for mammalian Argonautes in microRNA silencing. Genes Dev. 23, 304–317 (2009).

    Article  CAS  Google Scholar 

  65. Maniataki, E. & Mourelatos, Z. Human mitochondrial tRNAMet is exported to the cytoplasm and associates with the Argonaute 2 protein. RNA 11, 849–852 (2005).

    Article  CAS  Google Scholar 

  66. Zhang, X., Graves, P.R. & Zeng, Y. Stable Argonaute2 overexpression differentially regulates microRNA production. Biochim. Biophys. Acta 1789, 153–159 (2009).

    Article  CAS  Google Scholar 

  67. Diederichs, S. & Haber, D.A. Dual role for Argonautes in microRNA processing and posttranscriptional regulation of microRNA expression. Cell 131, 1097–1108 (2007).

    Article  CAS  Google Scholar 

  68. Ørom, U.A. & Lund, A.H. Isolation of microRNA targets using biotinylated synthetic microRNAs. Methods 43, 162–165 (2007).

    Article  Google Scholar 

  69. Ørom, U.A., Nielsen, F.C. & Lund, A.H. MicroRNA-10a binds the 5′UTR of ribosomal protein mRNAs and enhances their translation. Mol. Cell 30, 460–471 (2008).

    Article  Google Scholar 

  70. Didiano, D. & Hobert, O. Perfect seed pairing is not a generally reliable predictor for miRNA-target interactions. Nat. Struct. Mol. Biol. 13, 849–851 (2006).

    Article  CAS  Google Scholar 

  71. Lal, A. et al. p16INK4a translation suppressed by miR-24. PLoS ONE 3, e1864 (2008).

    Article  Google Scholar 

  72. Lal, A. et al. miR-24-mediated downregulation of H2AX suppresses DNA repair in terminally differentiated blood cells. Nat. Struct. Mol. Biol. 16, 492–498 (2009).

    Article  CAS  Google Scholar 

  73. Neilson, J.R., Zheng, G.X., Burge, C.B. & Sharp, P.A. Dynamic regulation of miRNA expression in ordered stages of cellular development. Genes Dev. 21, 578–589 (2007).

    Article  CAS  Google Scholar 

  74. Gu, S., Jin, L., Zhang, F., Sarnow, P. & Kay, M.A. Biological basis for restriction of microRNA targets to the 3′ untranslated region in mammalian mRNAs. Nat. Struct. Mol. Biol. 16, 144–150 (2009).

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding authors

Correspondence to Judy Lieberman or Ashish Lal.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Thomas, M., Lieberman, J. & Lal, A. Desperately seeking microRNA targets. Nat Struct Mol Biol 17, 1169–1174 (2010). https://doi.org/10.1038/nsmb.1921

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nsmb.1921

This article is cited by

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing