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:

A feedback regulatory loop involving microRNA-9 and nuclear receptor TLX in neural stem cell fate determination

Nature Structural & Molecular Biology volume 16, pages 365–371 (2009)Cite this article

Abstract

MicroRNAs have been implicated as having important roles in stem cell biology. MicroRNA-9 (miR-9) is expressed specifically in neurogenic areas of the brain and may be involved in neural stem cell self-renewal and differentiation. We showed previously that the nuclear receptor TLX is an essential regulator of neural stem cell self-renewal. Here we show that miR-9 suppresses TLX expression to negatively regulate neural stem cell proliferation and accelerate neural differentiation. Introducing a TLX expression vector that is not prone to miR-9 regulation rescued miR-9–induced proliferation deficiency and inhibited precocious differentiation. In utero electroporation of miR-9 in embryonic brains led to premature differentiation and outward migration of the transfected neural stem cells. Moreover, TLX represses expression of the miR-9 pri-miRNA. By forming a negative regulatory loop with TLX, miR-9 provides a model for controlling the balance between neural stem cell proliferation and differentiation.

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

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

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

Figure 1: miR-9–directed repression of TLX expression.
Figure 2: Overexpression of miR-9 regulates neural stem cell proliferation and differentiation.
Figure 3: TLXΔ3′ UTR rescues miR-9–induced neural stem cell proliferation deficiency.
Figure 4: miR-9 antisense RNA promotes neural stem cell proliferation.
Figure 5: In utero electroporation of miR-9 in embryonic neural stem cells.
Figure 6: Regulation of miR-9 pri-miRNA expression by TLX.

Similar content being viewed by others

References

  1. McKay, R. Stem cells in the central nervous system. Science 276, 66–71 (1997).

    Article  CAS  Google Scholar 

  2. Alvarez-Buylla, A. & Temple, S. Stem cells in the developing and adult nervous system. J. Neurobiol. 36, 105–110 (1998).

    Article  CAS  Google Scholar 

  3. Gage, F.H., Kempermann, G., Palmer, T.D., Peterson, D.A. & Ray, J. Multipotent progenitor cells in the adult dentate gyrus. J. Neurobiol. 36, 249–266 (1998).

    Article  CAS  Google Scholar 

  4. Weiss, S. & van der Kooy, D. CNS stem cells: where's the biology (a.k.a. beef)? J. Neurobiol. 36, 307–314 (1998).

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  8. 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 

  9. Bernstein, E. et al. Dicer is essential for mouse development. Nat. Genet. 35, 215–217 (2003).

    Article  CAS  Google Scholar 

  10. Wienholds, E., Koudijs, M.J., van Eeden, F.J., Cuppen, E. & Plasterk, R.H. The microRNA-producing enzyme Dicer1 is essential for zebrafish development. Nat. Genet. 35, 217–218 (2003).

    Article  CAS  Google Scholar 

  11. Carmell, M.A., Xuan, Z., Zhang, M.Q. & Hannon, G.J. The Argonaute family: tentacles that reach into RNAi, developmental control, stem cell maintenance, and tumorigenesis. Genes Dev. 16, 2733–2742 (2002).

    Article  CAS  Google Scholar 

  12. Lagos-Quintana, M. et al. Identification of tissue-specific microRNAs from mouse. Curr. Biol. 12, 735–739 (2002).

    Article  CAS  Google Scholar 

  13. Krichevsky, A.M., King, K.S., Donahue, C.P., Khrapko, K. & Kosik, K.S. A microRNA array reveals extensive regulation of microRNAs during brain development. RNA 9, 1274–1281 (2003).

    Article  CAS  Google Scholar 

  14. Deo, M., Yu, J.Y., Chung, K.H., Tippens, M. & Turner, D.L. Detection of mammalian microRNA expression by in situ hybridization with RNA oligonucleotides. Dev. Dyn. 235, 2538–2548 (2006).

    Article  CAS  Google Scholar 

  15. Kapsimali, M. et al. MicroRNAs show a wide diversity of expression profiles in the developing and mature central nervous system. Genome Biol. 8, R173 (2007).

    Article  Google Scholar 

  16. Shi, Y. et al. Expression and function of orphan nuclear receptor TLX in adult neural stem cells. Nature 427, 78–83 (2004).

    Article  CAS  Google Scholar 

  17. Sun, G., Yu, R.T., Evans, R.M. & Shi, Y. Orphan nuclear receptor TLX recruits histone deacetylases to repress transcription and regulate neural stem cell proliferation. Proc. Natl. Acad. Sci. USA 104, 15282–15287 (2007).

    Article  CAS  Google Scholar 

  18. Zhang, C.L., Zou, Y., He, W., Gage, F.H. & Evans, R.M. A role for adult TLX-positive neural stem cells in learning and behaviour. Nature 451, 1004–1007 (2008).

    Article  CAS  Google Scholar 

  19. Roy, K. et al. The Tlx gene regulates the timing of neurogenesis in the cortex. J. Neurosci. 24, 8333–8345 (2004).

    Article  CAS  Google Scholar 

  20. Stenman, J.M., Wang, B. & Campbell, K. Tlx controls proliferation and patterning of lateral telencephalic progenitor domains. J. Neurosci. 23, 10568–10576 (2003).

    Article  CAS  Google Scholar 

  21. Li, W. et al. Nuclear receptor TLX regulates cell cycle progression in neural stem cells of the developing brain. Mol. Endocrinol. 22, 56–64 (2008).

    Article  CAS  Google Scholar 

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

    Article  Google Scholar 

  23. 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 

  24. Sempere, L.F. et al. Expression profiling of mammalian microRNAs uncovers a subset of brain-expressed microRNAs with possible roles in murine and human neuronal differentiation. Genome Biol. 5, R13 (2004).

    Article  Google Scholar 

  25. Chen, C.Z., Li, L., Lodish, H.F. & Bartel, D.P. MicroRNAs modulate hematopoietic lineage differentiation. Science 303, 83–86 (2004).

    Article  CAS  Google Scholar 

  26. 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 

  27. Brennecke, J., Stark, A., Russell, R.B. & Cohen, S.M. Principles of microRNA-target recognition. PLoS Biol. 3, e85 (2005).

    Article  Google Scholar 

  28. Meister, G., Landthaler, M., Dorsett, Y. & Tuschl, T. Sequence-specific inhibition of microRNA- and siRNA-induced RNA silencing. RNA 10, 544–550 (2004).

    Article  CAS  Google Scholar 

  29. Hutvagner, G., Simard, M.J., Mello, C.C. & Zamore, P.D. Sequence-specific inhibition of small RNA function. PLoS Biol. 2, e98 (2004).

    Article  Google Scholar 

  30. Qian, X. et al. Timing of CNS cell generation: a programmed sequence of neuron and glial cell production from isolated murine cortical stem cells. Neuron 28, 69–80 (2000).

    Article  CAS  Google Scholar 

  31. Kim, J. et al. A microRNA feedback circuit in midbrain dopamine neurons. Science 317, 1220–1224 (2007).

    Article  CAS  Google Scholar 

  32. Conaco, C., Otto, S., Han, J.J. & Mandel, G. Reciprocal actions of REST and a microRNA promote neuronal identity. Proc. Natl. Acad. Sci. USA 103, 2422–2427 (2006).

    Article  CAS  Google Scholar 

  33. Wu, J. & Xie, X. Comparative sequence analysis reveals an intricate network among REST, CREB and miRNA in mediating neuronal gene expression. Genome Biol. 7, R85 (2006).

    Article  Google Scholar 

  34. Li, B., Carey, M. & Workman, J.L. The role of chromatin during transcription. Cell 128, 707–719 (2007).

    Article  CAS  Google Scholar 

  35. Kouzarides, T. Chromatin modifications and their function. Cell 128, 693–705 (2007).

    Article  CAS  Google Scholar 

  36. Krichevsky, A.M., Sonntag, K.C., Isacson, O. & Kosik, K.S. Specific microRNAs modulate embryonic stem cell-derived neurogenesis. Stem Cells 24, 857–864 (2006).

    Article  CAS  Google Scholar 

  37. Cao, X., Pfaff, S.L. & Gage, F.H. A functional study of miR-124 in the developing neural tube. Genes Dev. 21, 531–536 (2007).

    Article  CAS  Google Scholar 

  38. Visvanathan, J., Lee, S., Lee, B., Lee, J.W. & Lee, S.K. The microRNA miR-124 antagonizes the anti-neural REST/SCP1 pathway during embryonic CNS development. Genes Dev. 21, 744–749 (2007).

    Article  CAS  Google Scholar 

  39. Farh, K.K. et al. The widespread impact of mammalian microRNAs on mRNA repression and evolution. Science 310, 1817–1821 (2005).

    Article  CAS  Google Scholar 

  40. Bredenkamp, N., Seoighe, C. & Illing, N. Comparative evolutionary analysis of the FoxG1 transcription factor from diverse vertebrates identifies conserved recognition sites for microRNA regulation. Dev. Genes Evol. 217, 227–233 (2007).

    Article  CAS  Google Scholar 

  41. Li, Y., Wang, F., Lee, J.A. & Gao, F.B. MicroRNA-9a ensures the precise specification of sensory organ precursors in Drosophila. Genes Dev. 20, 2793–2805 (2006).

    Article  CAS  Google Scholar 

  42. Leucht, C. et al. MicroRNA-9 directs late organizer activity of the midbrain-hindbrain boundary. Nat. Neurosci. 11, 641–648 (2008).

    Article  CAS  Google Scholar 

  43. Hornstein, E. & Shomron, N. Canalization of development by microRNAs. Nat. Genet. 38, S20–S24 (2006).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank J. Rossi and J. Zaia for their critical comments on the manuscript, C.-Z. Chen (Stanford University), H.F. Lodish and D.P. Bartel (Massachusetts Institute of Technology) for providing the microRNA expression vector MDH1-PGK-GFP2, and Q. Lu (City of Hope) for providing the pEF-pUb-EGFP plasmid. This work was supported by the US National Institutes of Health, National Institute of Neurological Disorders and Stroke grant R01 NS059546 (to Y.S.).

Author information

Authors and Affiliations

  1. Department of Neurosciences, Center for Gene Expression and Drug Discovery, Beckman Research Institute of City of Hope, Duarte, California, USA

    Chunnian Zhao, GuoQiang Sun, Shengxiu Li & Yanhong Shi

Authors
  1. Chunnian Zhao
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. GuoQiang Sun
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Shengxiu Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Yanhong Shi
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

Y.S., C.Z. and G.S. designed the project; C.Z., G.S. and S.L. performed the experiments; Y.S. and C.Z. analyzed the data and wrote the manuscript.

Corresponding author

Correspondence to Yanhong Shi.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–4 and Supplementary Methods (PDF 440 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Zhao, C., Sun, G., Li, S. et al. A feedback regulatory loop involving microRNA-9 and nuclear receptor TLX in neural stem cell fate determination. Nat Struct Mol Biol 16, 365–371 (2009). https://doi.org/10.1038/nsmb.1576

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nsmb.1576

This article is cited by

Search

Advanced 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