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Embryonic stem cell–specific microRNAs regulate the G1-S transition and promote rapid proliferation

Nature Genetics volume 40, pages 1478–1483 (2008)Cite this article

Abstract

Dgcr8 knockout embryonic stem (ES) cells lack microprocessor activity and hence all canonical microRNAs (miRNAs). These cells proliferate slowly and accumulate in G1 phase of the cell cycle1. Here, by screening a comprehensive library of individual miRNAs in the background of the Dgcr8 knockout ES cells, we report that multiple ES cell–specific miRNAs, members of the miR-290 family, rescue the ES cell proliferation defect. Furthermore, rescued cells no longer accumulate in the G1 phase of the cell cycle. These miRNAs function by suppressing several key regulators of the G1-S transition. These results show that post-transcriptional regulation by miRNAs promotes the G1-S transition of the ES cell cycle, enabling rapid proliferation of these cells. Our screening strategy provides an alternative and powerful approach for uncovering the role of individual miRNAs in biological processes, as it overcomes the common problem of redundancy and saturation in the miRNA system.

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Figure 1: Screening for miRNAs that rescue the proliferation defects of Dgcr8 knockout (Dgcr8Δ/Δ) ES cells.
Figure 2: Rescue of proliferation and G1 accumulation defects by representative miRNAs.
Figure 3: Expression of Cdkn1a mRNA and protein upon introduction of ESCC miRNAs.
Figure 4: Luciferase reporter assay indicates that ESCC miRNAs directly interact with the 3′ UTR of Cdkn1a.
Figure 5: Additional inhibitors of the cyclin E–Cdk2 pathway are regulated by miRNAs in ES cells.

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References

  1. Wang, Y., Medvid, R., Melton, C., Jaenisch, R. & Blelloch, R. DGCR8 is essential for microRNA biogenesis and silencing of embryonic stem cell self-renewal. Nat. Genet. 39, 380–385 (2007).

    Article  CAS  Google Scholar 

  2. Keller, G. Embryonic stem cell differentiation: emergence of a new era in biology and medicine. Genes Dev. 19, 1129–1155 (2005).

    Article  CAS  Google Scholar 

  3. White, J. & Dalton, S. Cell cycle control of embryonic stem cells. Stem Cell Rev. 1, 131–138 (2005).

    Article  CAS  Google Scholar 

  4. Burdon, T., Smith, A. & Savatier, P. Signalling, cell cycle and pluripotency in embryonic stem cells. Trends Cell Biol. 12, 432–438 (2002).

    Article  CAS  Google Scholar 

  5. Sabapathy, K., Klemm, M., Jaenisch, R. & Wagner, E.F. Regulation of ES cell differentiation by functional and conformational modulation of p53. EMBO J. 16, 6217–6229 (1997).

    Article  CAS  Google Scholar 

  6. Sorrentino, E., Nazzicone, V., Farini, D., Campagnolo, L. & De Felici, M. Comparative transcript profiles of cell cycle-related genes in mouse primordial germ cells, embryonic stem cells and embryonic germ cells. Gene Expr. Patterns 7, 714–721 (2007).

    Article  CAS  Google Scholar 

  7. Stead, E. et al. Pluripotent cell division cycles are driven by ectopic Cdk2, cyclin A/E and E2F activities. Oncogene 21, 8320–8333 (2002).

    Article  CAS  Google Scholar 

  8. White, J. et al. Developmental activation of the Rb-E2F pathway and establishment of cell cycle-regulated cyclin-dependent kinase activity during embryonic stem cell differentiation. Mol. Biol. Cell 16, 2018–2027 (2005).

    Article  CAS  Google Scholar 

  9. Murchison, E.P., Partridge, J.F., Tam, O.H., Cheloufi, S. & Hannon, G.J. Characterization of Dicer-deficient murine embryonic stem cells. Proc. Natl. Acad. Sci. USA 102, 12135–12140 (2005).

    Article  CAS  Google Scholar 

  10. Shcherbata, H.R. et al. The MicroRNA pathway plays a regulatory role in stem cell division. Cell Cycle 5, 172–175 (2006).

    Article  CAS  Google Scholar 

  11. Chen, C. et al. Defining embryonic stem cell identity using differentiation-related microRNAs and their potential targets. Mamm. Genome 18, 316–327 (2007).

    Article  CAS  Google Scholar 

  12. Houbaviy, H.B., Murray, M.F. & Sharp, P.A. Embryonic stem cell-specific microRNAs. Dev. Cell 5, 351–358 (2003).

    Article  CAS  Google Scholar 

  13. Houbaviy, H.B., Dennis, L., Jaenisch, R. & Sharp, P.A. Characterization of a highly variable eutherian microRNA gene. RNA 11, 1245–1257 (2005).

    Article  CAS  Google Scholar 

  14. Suh, M.R. et al. Human embryonic stem cells express a unique set of microRNAs. Dev. Biol. 270, 488–498 (2004).

    Article  CAS  Google Scholar 

  15. Calabrese, J.M., Seila, A.C., Yeo, G.W. & Sharp, P.A. RNA sequence analysis defines Dicer's role in mouse embryonic stem cells. Proc. Natl. Acad. Sci. USA 104, 18097–18102 (2007).

    Article  CAS  Google Scholar 

  16. Kato, J. Induction of S phase by G1 regulatory factors. Front. Biosci. 4, D787–D792 (1999).

    Article  CAS  Google Scholar 

  17. Lee, M.H. & Yang, H.Y. Regulators of G1 cyclin-dependent kinases and cancers. Cancer Metastasis Rev. 22, 435–449 (2003).

    Article  CAS  Google Scholar 

  18. Savatier, P., Lapillonne, H., van Grunsven, L.A., Rudkin, B.B. & Samarut, J. Withdrawal of differentiation inhibitory activity/leukemia inhibitory factor up-regulates D-type cyclins and cyclin-dependent kinase inhibitors in mouse embryonic stem cells. Oncogene 12, 309–322 (1996).

    CAS  PubMed  Google Scholar 

  19. Ambros, V. microRNAs: tiny regulators with great potential. Cell 107, 823–826 (2001).

    Article  CAS  Google Scholar 

  20. Bartel, D.P. MicroRNAs: genomics, biogenesis, mechanism, and function. Cell 116, 281–297 (2004).

    Article  CAS  Google Scholar 

  21. He, L. & Hannon, G.J. MicroRNAs: small RNAs with a big role in gene regulation. Nat. Rev. Genet. 5, 522–531 (2004).

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  23. 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 

  24. Li, Y. et al. Lats2, a putative tumor suppressor, inhibits G1/S transition. Oncogene 22, 4398–4405 (2003).

    Article  CAS  Google Scholar 

  25. Voorhoeve, P.M. et al. A genetic screen implicates miRNA-372 and miRNA-373 as oncogenes in testicular germ cell tumors. Cell 124, 1169–1181 (2006).

    Article  CAS  Google Scholar 

  26. Ivanovska, I. et al. MicroRNAs in the miR-106b family regulate p21/CDKN1A and promote cell cycle progression. Mol. Cell. Biol. 28, 2167–2174 (2008).

    Article  CAS  Google Scholar 

  27. Benetti, R. et al. A mammalian microRNA cluster controls DNA methylation and telomere recombination via Rbl2-dependent regulation of DNA methyltransferases. Nat. Struct. Mol. Biol. 15, 268–279 (2008).

    Article  CAS  Google Scholar 

  28. Sinkkonen, L. et al. MicroRNAs control de novo DNA methylation through regulation of transcriptional repressors in mouse embryonic stem cells. Nat. Struct. Mol. Biol. 15, 259–267 (2008).

    Article  CAS  Google Scholar 

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Acknowledgements

We would like to thank A. D. Sharma and K. Okamura for their technical assistance with the immunoblot assay; M. Grskovic and C. Chaivorapol for their technical assistance with the luciferase reporter assay; M. Sachs for help in developing the overexpression construct; and M. Conti, M. Ramalho-Santos, J. Reiter and members of the Blelloch laboratory for critical reading of the manuscript. This work was supported by funds to R.B. from the US National Institutes of Health (K08 NS48118 and R01 NS057221), the Stem Cell Research Foundation and the Pew Charitable Trust. Y.W. is supported by a California Institute of Regenerative Medicine Training Grant.

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Authors and Affiliations

  1. Center for Reproductive Sciences and Department of Urology, Institute for Regeneration Medicine, University of California – San Francisco, San Francisco, 94143, California, USA

    Yangming Wang, Archana Shenoy, Joshua E Babiarz, Lauren Baehner & Robert Blelloch

  2. Dharmacon Technologies, Thermo Fisher Scientific, Lafayette, 80026, Colorado, USA

    Scott Baskerville

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  1. Yangming Wang
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Contributions

Y.W. performed all experiments except those shown in Figure 3a, which were done by J.E.B. S.B. developed and provided the miRNA mimic library. A.S. built mutant reporters for Figure 4. L.B. provided technical support. Y.W. and R.B. designed all experiments, analyzed data and wrote the manuscript.

Note: Supplementary information is available on the Nature Genetics website.

Corresponding author

Correspondence to Robert Blelloch.

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Competing interests

Scott Baskerville was an employee of Dharmacon/Thermo Fisher at the time of this study. He, in the form of a collaboration, supplied the microRNA mimic library used in experiments described in the manuscript.

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Supplementary Figures 1–6 and Supplementary Tables 1 and 2 (PDF 2120 kb)

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Wang, Y., Baskerville, S., Shenoy, A. et al. Embryonic stem cell–specific microRNAs regulate the G1-S transition and promote rapid proliferation. Nat Genet 40, 1478–1483 (2008). https://doi.org/10.1038/ng.250

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