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.

  • Article
  • Published:

Lin28 recruits the TUTase Zcchc11 to inhibit let-7 maturation in mouse embryonic stem cells

Abstract

Lin28 and Lin28B, two developmentally regulated RNA-binding proteins and likely proto-oncogenes, selectively inhibit the maturation of let-7 family microRNAs (miRNAs) in embryonic stem cells and certain cancer cell lines. Moreover, let-7 precursors (pre–let-7) were previously found to be terminally uridylated in a Lin28-dependent fashion. Here we identify Zcchc11 (zinc finger, CCHC domain containing 11) as the 3′ terminal uridylyl transferase (TUTase) responsible for Lin28-mediated pre–let-7 uridylation and subsequent blockade of let-7 processing in mouse embryonic stem cells. We demonstrate that Zcchc11 activity is UTP-dependent, selective for let-7 and recruited by Lin28. Furthermore, knockdown of either Zcchc11 or Lin28, or overexpression of a catalytically inactive TUTase, relieves the selective inhibition of let-7 processing and leads to the accumulation of mature let-7 miRNAs and repression of let-7 target reporter genes. Our results establish a role for Zcchc11-catalyzed pre–let-7 uridylation in the control of miRNA biogenesis.

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: Identification of Zcchc11 as a candidate pre–let-7 TU Tase.
Figure 2: Zcchc11 TUTase activity is Lin28-dependent.
Figure 3: Inhibition of Zcchc11 selectively elevates mature let-7 levels in embryonic cells.
Figure 4: Zcchc11 inhibition exclusively affects let-7 biogenesis and leads to repression of let-7 target gene expression.

Similar content being viewed by others

Accession codes

Accessions

Protein Data Bank

References

  1. 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 

  2. Blakaj, A. & Lin, H. Piecing together the mosaic of early mammalian development through microRNAs. J. Biol. Chem. 283, 9505–9508 (2008).

    Article  CAS  Google Scholar 

  3. Esquela-Kerscher, A. & Slack, F.J. Oncomirs—microRNAs with a role in cancer. Nat. Rev. Cancer 6, 259–269 (2006).

    Article  CAS  Google Scholar 

  4. Hagan, J.P. & Croce, C.M. MicroRNAs in carcinogenesis. Cytogenet. Genome Res. 118, 252–259 (2007).

    Article  CAS  Google Scholar 

  5. Büssing, I., Slack, F.J. & Grosshans, H. let-7 microRNAs in development, stem cells and cancer. Trends Mol. Med. 14, 400–409 (2008).

    Article  Google Scholar 

  6. Viswanathan, S.R., Daley, G.Q. & Gregory, R.I. Selective blockade of microRNA processing by Lin28. Science 320, 97–100 (2008).

    Article  CAS  Google Scholar 

  7. Rybak, A. et al. A feedback loop comprising lin-28 and let-7 controls pre–let-7 maturation during neural stem-cell commitment. Nat. Cell Biol. 10, 987–993 (2008).

    Article  CAS  Google Scholar 

  8. Newman, M.A., Thomson, J.M. & Hammond, S.M. Lin-28 interaction with the let-7 precursor loop mediates regulated microRNA processing. RNA 14, 1539–1549 (2008).

    Article  CAS  Google Scholar 

  9. Heo, I. et al. Lin28 mediates the terminal uridylation of let-7 precursor microRNA. Mol. Cell 32, 276–284 (2008).

    Article  CAS  Google Scholar 

  10. Piskounova, E. et al. Determinants of microRNA processing inhibition by the developmentally regulated RNA-binding protein Lin28. J. Biol. Chem. 283, 21310–21314 (2008).

    Article  CAS  Google Scholar 

  11. Ambros, V. & Horvitz, H.R. Heterochronic mutants of the nematode Caenorhabditis elegans. Science 226, 409–416 (1984).

    Article  CAS  Google Scholar 

  12. Yu, J. et al. Induced pluripotent stem cell lines derived from human somatic cells. Science 318, 1917–1920 (2007).

    Article  CAS  Google Scholar 

  13. West, J.A. et al. A role for Lin28 in primordial germ-cell development and germ-cell malignancy. Nature 460, 909–913 (2009).

    Article  CAS  Google Scholar 

  14. Lettre, G. et al. Identification of ten loci associated with height highlights new biological pathways in human growth. Nat. Genet. 40, 584–591 (2008).

    Article  CAS  Google Scholar 

  15. Sulem, P. et al. Genome-wide association study identifies sequence variants on 6q21 associated with age at menarche. Nat. Genet. 41, 734–738 (2009).

    Article  CAS  Google Scholar 

  16. Ong, K.K. et al. Genetic variation in LIN28B is associated with the timing of puberty. Nat. Genet. 41, 729–733 (2009).

    Article  CAS  Google Scholar 

  17. Stolk, L. et al. Loci at chromosomes 13, 19 and 20 influence age at natural menopause. Nat. Genet. 41, 645–647 (2009).

    Article  CAS  Google Scholar 

  18. He, C. et al. Genome-wide association studies identify loci associated with age at menarche and age at natural menopause. Nat. Genet. 41, 724–728 (2009).

    Article  CAS  Google Scholar 

  19. Perry, J.R. et al. Meta-analysis of genome-wide association data identifies two loci influencing age at menarche. Nat. Genet. 41, 648–650 (2009).

    Article  CAS  Google Scholar 

  20. Dangi-Garimella, S. et al. Raf kinase inhibitory protein suppresses a metastasis signalling cascade involving LIN28 and let-7. EMBO J. 28, 347–358 (2009).

    Article  CAS  Google Scholar 

  21. Guo, Y. et al. Identification and characterization of lin-28 homolog B (LIN28B) in human hepatocellular carcinoma. Gene 384, 51–61 (2006).

    Article  CAS  Google Scholar 

  22. Chang, T.C. et al. Lin-28B transactivation is necessary for Myc-mediated let-7 repression and proliferation. Proc. Natl. Acad. Sci. USA 106, 3384–3389 (2009).

    Article  CAS  Google Scholar 

  23. Viswanathan, S.R. et al. LIN28 promotes transformation and is associated with advanced human malignancies. Nat. Genet. 41, 843–848 (2009).

    Article  CAS  Google Scholar 

  24. Martin, G. & Keller, W. RNA-specific ribonucleotidyl transferases. RNA 13, 1834–1849 (2007).

    Article  CAS  Google Scholar 

  25. Wilusz, C.J. & Wilusz, J. New ways to meet your (3′) end oligouridylation as a step on the path to destruction. Genes Dev. 22, 1–7 (2008).

    Article  CAS  Google Scholar 

  26. Trippe, R. et al. Identification, cloning, and functional analysis of the human U6 snRNA-specific terminal uridylyl transferase. RNA 12, 1494–1504 (2006).

    Article  CAS  Google Scholar 

  27. Mullen, T.E. & Marzluff, W.F. Degradation of histone mRNA requires oligouridylation followed by decapping and simultaneous degradation of the mRNA both 5′ to 3′ and 3′ to 5′. Genes Dev. 22, 50–65 (2008).

    Article  CAS  Google Scholar 

  28. Kwak, J.E. & Wickens, M. A family of poly(U) polymerases. RNA 13, 860–867 (2007).

    Article  CAS  Google Scholar 

  29. Tomecki, R., Dmochowska, A., Gewartowski, K., Dziembowski, A. & Stepien, P.P. Identification of a novel human nuclear-encoded mitochondrial poly(A) polymerase. Nucleic Acids Res. 32, 6001–6014 (2004).

    Article  CAS  Google Scholar 

  30. Lehrbach N.J. et al. LIN-28 and the poly(U) polymerase PUP-2 regulate let-7 microRNA processing in Caenorhabditis elegans. Nat. Struct. Mol. Biol. advance online publication, doi:10.1038/nsmb.1675 (27 August 2009).

  31. Rissland, O.S. & Norbury, C.J. Decapping is preceded by 3′ uridylation in a novel pathway of bulk mRNA turnover. Nat. Struct. Mol. Biol. 16, 616–623 (2009).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We are grateful to S. Viswanathan and G. Daley (Children's Hospital Boston) for providing Flag-Lin28–expressing ES cells, H. Rusk and R. LaPierre for technical assistance and the Proteomics Center at Children's Hospital Boston for expertise in the microcapillary HPLC and MS. We thank E. Miska (University of Cambridge) for helpful discussions. R.I.G. is supported by laboratory start-up funds from The Children's Hospital Boston and grants from the US National Institute of General Medical Sciences (NIGMS) (1R01GM086386-01A1), The Harvard Stem Cell Institute, The March of Dimes Basil O'Conner award and the Emerald Foundation. R.I.G. is supported by The Pew Scholars Program in the Biomedical Sciences.

Author information

Authors and Affiliations

Authors

Contributions

J.P.H. and E.P. performed all experiments; J.P.H., E.P. and R.I.G. designed all experiments, analyzed data and wrote the manuscript.

Corresponding author

Correspondence to Richard I Gregory.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–3 and Supplementary Tables 1–4 (PDF 1178 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Hagan, J., Piskounova, E. & Gregory, R. Lin28 recruits the TUTase Zcchc11 to inhibit let-7 maturation in mouse embryonic stem cells. Nat Struct Mol Biol 16, 1021–1025 (2009). https://doi.org/10.1038/nsmb.1676

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nsmb.1676

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