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:

Cyclical DNA methylation of a transcriptionally active promoter

Nature volume 452, pages 45–50 (2008)Cite this article

A Corrigendum to this article was published on 21 January 2010

Abstract

Processes that regulate gene transcription are directly under the influence of the genome organization. The epigenome contains additional information that is not brought by DNA sequence, and generates spatial and functional constraints that complement genetic instructions. DNA methylation on CpGs constitutes an epigenetic mark generally correlated with transcriptionally silent condensed chromatin. Replication of methylation patterns by DNA methyltransferases maintains genome stability through cell division. Here we present evidence of an unanticipated dynamic role for DNA methylation in gene regulation in human cells. Periodic, strand-specific methylation/demethylation occurs during transcriptional cycling of the pS2/TFF1 gene promoter on activation by oestrogens. DNA methyltransferases exhibit dual actions during these cycles, being involved in CpG methylation and active demethylation of 5mCpGs through deamination. Inhibition of this process precludes demethylation of the pS2 gene promoter and its subsequent transcriptional activation. Cyclical changes in the methylation status of promoter CpGs may thus represent a critical event in transcriptional achievement.

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: Cyclical methylation of the pS2 promoter.
Figure 2: Kinetic association of Dnmts with transcriptionally active pS2 promoter in MDA::ERα cells.
Figure 3: Inhibition of Dnmt activity modifies pS2 transcriptional cycling.
Figure 4: Mobilization of TDG, p68, Dnmt3a/b and BER proteins during pS2 promoter demethylation.
Figure 5: Dnmt3a/b deaminate CpGs and 5m CpGs.

Similar content being viewed by others

References

  1. Spector, D. L. The dynamics of chromosome organization and gene regulation. Annu. Rev. Biochem. 72, 573–608 (2003)

    Article  CAS  Google Scholar 

  2. Casolari, J. M., Brown, C. R., Drubin, D. A., Rando, O. J. & Silver, P. A. Developmentally induced changes in transcriptional program alter spatial organization across chromosomes. Genes Dev. 19, 1188–1198 (2005)

    Article  CAS  Google Scholar 

  3. Khorasanizadeh, S. The nucleosome: from genomic organization to genomic regulation. Cell 116, 259–272 (2004)

    Article  CAS  Google Scholar 

  4. Dillon, N. Gene regulation and large-scale chromatin organization in the nucleus. Chromosome Res. 14, 117–126 (2006)

    Article  CAS  Google Scholar 

  5. Klose, R. J. & Bird, A. P. Genomic DNA methylation: the mark and its mediators. Trends Biochem. Sci. 31, 89–97 (2006)

    Article  CAS  Google Scholar 

  6. Oswald, J. et al. Active demethylation of the paternal genome in the mouse zygote. Curr. Biol. 10, 475–478 (2000)

    Article  CAS  Google Scholar 

  7. Murayama, A. et al. A specific CpG site demethylation in the human interleukin 2 gene promoter is an epigenetic memory. EMBO J. 25, 1081–1092 (2006)

    Article  CAS  Google Scholar 

  8. Rougier, N. et al. Chromosome methylation patterns during mammalian preimplantation development. Genes Dev. 12, 2108–2113 (1998)

    Article  CAS  Google Scholar 

  9. Matsuo, K. et al. An embryonic demethylation mechanism involving binding of transcription factors to replicating DNA. EMBO J. 17, 1446–1453 (1998)

    Article  CAS  Google Scholar 

  10. Morales-Ruiz, T. et al. Demeter and Repressor of Silencing 1 encode 5-methylcytosine DNA glycosylases. Proc. Natl Acad. Sci. USA 103, 6853–6858 (2006)

    Article  ADS  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  12. Becker, P. B. & Horz, W. ATP-dependent nucleosome remodeling. Annu. Rev. Biochem. 71, 247–273 (2002)

    Article  CAS  Google Scholar 

  13. Shang, Y., Hu, X., DiRenzo, J., Lazar, M. A. & Brown, M. Cofactor dynamics and sufficiency in estrogen receptor-regulated transcription. Cell 103, 843–852 (2000)

    Article  CAS  Google Scholar 

  14. Metivier, R. et al. Estrogen receptor-alpha directs ordered, cyclical, and combinatorial recruitment of cofactors on a natural target promoter. Cell 115, 751–763 (2003)

    Article  CAS  Google Scholar 

  15. Robinson-Rechavi, M., Escriva Garcia, H. & Laudet, V. The nuclear receptor superfamily. J. Cell Sci. 116, 585–586 (2003)

    Article  Google Scholar 

  16. Brzozowski, A. M. et al. Molecular basis of agonism and antagonism in the oestrogen receptor. Nature 389, 753–758 (1997)

    Article  ADS  CAS  Google Scholar 

  17. Zhang, Y. et al. Analysis of the NuRD subunits reveals a histone deacetylase core complex and a connection with DNA methylation. Genes Dev. 13, 1924–1935 (1999)

    Article  CAS  Google Scholar 

  18. Harikrishnan, K. N. et al. Brahma links the SWI/SNF chromatin-remodeling complex with MeCP2-dependent transcriptional silencing. Nature Genet. 37, 254–264 (2005)

    Article  CAS  Google Scholar 

  19. Christman, J. K. 5-Azacytidine and 5-aza-2′-deoxycytidine as inhibitors of DNA methylation: mechanistic studies and their implications for cancer therapy. Oncogene 21, 5483–5495 (2002)

    Article  CAS  Google Scholar 

  20. Brueckner, B. et al. Epigenetic reactivation of tumor suppressor genes by a novel small-molecule inhibitor of human DNA methyltransferases. Cancer Res. 65, 6305–6311 (2005)

    Article  CAS  Google Scholar 

  21. Sharma, D., Saxena, N. K., Davidson, N. E. & Vertino, P. M. Restoration of tamoxifen sensitivity in estrogen receptor-negative breast cancer cells: tamoxifen-bound reactivated ER recruits distinctive corepressor complexes. Cancer Res. 66, 6370–6378 (2006)

    Article  CAS  Google Scholar 

  22. Zingg, J. M., Shen, J. C., Yang, A. S., Rapoport, H. & Jones, P. A. Methylation inhibitors can increase the rate of cytosine deamination by (cytosine-5)-DNA methyltransferase. Nucleic Acids Res. 24, 3267–3275 (1996)

    Article  CAS  Google Scholar 

  23. Sharath, A. N., Weinhold, E. & Bhagwat, A. S. Reviving a dead enzyme: cytosine deaminations promoted by an inactive DNA methyltransferase and an S-adenosylmethionine analogue. Biochemistry 39, 14611–14616 (2000)

    Article  CAS  Google Scholar 

  24. Waters, T. R., Gallinari, P., Jiricny, J. & Swann, P. F. Human thymine DNA glycosylase binds to apurinic sites in DNA but is displaced by human apurinic endonuclease 1. J. Biol. Chem. 274, 67–74 (1999)

    Article  CAS  Google Scholar 

  25. Li, Y. Q., Zhou, P. Z., Zheng, X. D., Walsh, C. P. & Xu, G. L. Association of Dnmt3a and thymine DNA glycosylase links DNA methylation with base-excision repair. Nucleic Acids Res. 35, 390–400 (2007)

    Article  CAS  Google Scholar 

  26. Gallais, R. et al. Deoxyribonucleic acid methyl transferases 3a and 3b associate with the nuclear orphan receptor COUP-TFI during gene activation. Mol. Endocrinol. 21, 2085–2098 (2007)

    Article  CAS  Google Scholar 

  27. Endoh, H. et al. Purification and identification of p68 RNA helicase acting as a transcriptional coactivator specific for the activation function 1 of human estrogen receptor alpha. Mol. Cell. Biol. 19, 5363–5372 (1999)

    Article  CAS  Google Scholar 

  28. Jost, J. P. et al. A chicken embryo protein related to the mammalian DEAD box protein p68 is tightly associated with the highly purified protein-RNA complex of 5-MeC-DNA glycosylase. Nucleic Acids Res. 27, 3245–3252 (1999)

    Article  CAS  Google Scholar 

  29. Suetake, I., Miyazaki, J., Murakami, C., Takeshima, H. & Tajima, S. Distinct enzymatic properties of recombinant mouse DNA methyltransferases Dnmt3a and Dnmt3b. J. Biochem. 133, 737–744 (2003)

    Article  CAS  Google Scholar 

  30. Fritah, A., Redeuilh, G. & Sabbah, M. Molecular cloning and characterization of the human WISP-2/CCN5 gene promoter reveal its upregulation by oestrogens. J. Endocrinol. 191, 613–624 (2006)

    Article  CAS  Google Scholar 

  31. Fatemi, M., Hermann, A., Gowher, H. & Jeltsch, A. Dnmt3a and Dnmt1 functionally cooperate during de novo methylation of DNA. Eur. J. Biochem. 269, 4981–4984 (2002)

    Article  CAS  Google Scholar 

  32. Chen, D. et al. T:G mismatch-specific thymine-DNA glycosylase potentiates transcription of estrogen-regulated genes through direct interaction with estrogen receptor alpha. J. Biol. Chem. 278, 38586–38592 (2003)

    Article  CAS  Google Scholar 

  33. Jost, J. P., Thiry, S. & Siegmann, M. Estradiol receptor potentiates, in vitro, the activity of 5-methylcytosine DNA glycosylase. FEBS Lett. 527, 63–66 (2002)

    Article  CAS  Google Scholar 

  34. Zhu, B. et al. 5-methylcytosine-DNA glycosylase activity is present in a cloned G/T mismatch DNA glycosylase associated with the chicken embryo DNA demethylation complex. Proc. Natl Acad. Sci. USA 97, 5135–5139 (2000)

    Article  ADS  CAS  Google Scholar 

  35. Fuks, F. DNA methylation and histone modifications: teaming up to silence genes. Curr. Opin. Genet. Dev. 15, 490–495 (2005)

    Article  CAS  Google Scholar 

  36. Oka, M., Rodic, N., Graddy, J., Chang, L. J. & Terada, N. CpG sites preferentially methylated by Dnmt3a in vivo. J. Biol. Chem. 281, 9901–9908 (2006)

    Article  CAS  Google Scholar 

  37. Li, F. et al. Chimeric DNA methyltransferases target DNA methylation to specific DNA sequences and repress expression of target genes. Nucleic Acids Res. 35, 100–112 (2007)

    Article  Google Scholar 

  38. Lucarelli, M., Fuso, A., Strom, R. & Scarpa, S. The dynamics of myogenin site-specific demethylation is strongly correlated with its expression and with muscle differentiation. J. Biol. Chem. 276, 7500–7506 (2001)

    Article  CAS  Google Scholar 

  39. Kress, C., Thomassin, H. & Grange, T. Active cytosine demethylation triggered by a nuclear receptor involves DNA strand breaks. Proc. Natl Acad. Sci. USA 103, 11112–11117 (2006)

    Article  ADS  CAS  Google Scholar 

  40. Barreto, G. et al. Gadd45a promotes epigenetic gene activation by repair-mediated DNA demethylation. Nature 445, 671–675 (2007)

    Article  CAS  Google Scholar 

  41. Leteurtre, F. et al. Effects of DNA methylation on topoisomerase I and II cleavage activities. J. Biol. Chem. 269, 7893–7900 (1994)

    CAS  PubMed  Google Scholar 

  42. Ju, B. G. et al. A topoisomerase IIbeta-mediated dsDNA break required for regulated transcription. Science 312, 1798–1802 (2006)

    Article  ADS  CAS  Google Scholar 

  43. Lamm, G. M., Nicol, S. M., Fuller-Pace, F. V. & Lamond, A. I. p72: a human nuclear DEAD box protein highly related to p68. Nucleic Acids Res. 24, 3739–3947 (1996)

    Article  CAS  Google Scholar 

  44. Gowher, H., Liebert, K., Hermann, A., Xu, G. & Jeltsch, A. Mechanism of stimulation of catalytic activity of Dnmt3A and Dnmt3B DNA-(cytosine-C5)-methyltransferases by Dnmt3L. J. Biol. Chem. 280, 13341–13348 (2005)

    Article  CAS  Google Scholar 

  45. Gowher, H. et al. Mutational analysis of the catalytic domain of the murine Dnmt3a DNA-(cytosine C5)-methyltransferase. J. Mol. Biol. 357, 928–941 (2006)

    Article  CAS  Google Scholar 

  46. Metivier, R. et al. Transcriptional complexes engaged by apo-oestrogen receptor-alpha isoforms have divergent outcomes. EMBO J. 23, 3653–3666 (2004)

    Article  CAS  Google Scholar 

  47. Nelson, J. E. & Krawetz, S. A. Purification of cloned and genomic DNA by guanidine thiocyanate/isobutyl alcohol fractionation. Anal. Biochem. 207, 197–201 (1992)

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank B. Brueckner for the gift of the RG108 compound, P. Chambon for the pSG5-TDGwt plasmid, F. Fuller-Pace for the anti-p68 RNA helicase antibody, and V. Legagneux and U. K. Laemmli for the anti-CAPD2 and anti-CAPH antibodies, respectively. This work was supported by funds from the Ministère de l’Education Nationale de l’Enseignement Supérieur et de la Recherche (MENESR), the Centre National de la Recherche Scientifique (CNRS), the University of Rennes I, the Association pour la Recherche contre le Cancer (ARC), the Ligue contre le Cancer, and by funding from EMBO and EMBL. R.G. was supported by a grant from the MENESR and a fellowship from the Ligue Nationale Contre le Cancer. We also thank C. Ralliere for DNA sequencing, C. Martin, F. Percevault and C. Tascon for their technical assistance, and B.Stride and S. Kangaspeska for their comments during the writing of the manuscript.

Authors Contributions ChIP, sequential-ChIP experiments, bisulphite and run-on assays were performed by R.M. Methylation/deamination assays were set up by R.G. and performed by R.G. and R.M. C.T. ran all RT-PCR experiments on siRNA-treated cells, and set up the analysis on the Wisp-2 gene. Co-immunoprecipitations and proteomic controls were performed by C.L.P., R.G. and R.M. Synthesis and purification of TDG protein were performed by P.B. and F.D., as the preparation of the anti-TDG antibody. R.Z.J. purified all the Dnmt3 catalytic domains. R.P.C. and D.I. were involved, under the supervision of V.B., in the mass sequencing of the clones generated during the bisulphite-mediated analysis of CpG methylation and through the in vitro deamination assays. G.R. introduced R.M. to FACS analysis and bisulphite-modification of DNA. R.M., G.R., A.J., F.G. and G.S. were responsible for the overall project management, strategy and data interpretation. R.M., G.R. and G.S. prepared the manuscript. All authors discussed the results and commented on the manuscript.

Author information

Author notes

  1. Peter Barath & Frank Gannon

    Present address: Present addresses: Cancer Research Institute, Slovak Academy of Sciences, Vlarska 7, SK-833 91 Bratislava, Slovak Republic (P.B.); Science Foundation Ireland, Wilton Park House, Wilton Place, Dublin 2, Ireland (F.G.).,

Authors and Affiliations

  1. Université de Rennes I, CNRS, UMR 6026 Equipe SPARTE, IFR 140 GFAS, Campus de Beaulieu, 35042 Rennes cedex, France

    Raphaël Métivier, Rozenn Gallais, Christophe Tiffoche, Christine Le Péron, Peter Barath, Florence Demay & Gilles Salbert

  2. Jacobs University Bremen, Campus Ring 1, 28759 Bremen, Germany

    Renata Z. Jurkowska & Albert Jeltsch

  3. EMBL, Meyerhofstrasse 1, D-69117 Heidelberg, Germany,

    Richard P. Carmouche, David Ibberson, George Reid, Vladimir Benes & Frank Gannon

Authors
  1. Raphaël Métivier
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Rozenn Gallais
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Christophe Tiffoche
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Christine Le Péron
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Renata Z. Jurkowska
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Richard P. Carmouche
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. David Ibberson
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Peter Barath
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Florence Demay
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. George Reid
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Vladimir Benes
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Albert Jeltsch
    View author publications

    You can also search for this author in PubMed Google Scholar

  13. Frank Gannon
    View author publications

    You can also search for this author in PubMed Google Scholar

  14. Gilles Salbert
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Raphaël Métivier.

Supplementary information

Supplementary Information

The file contains Supplementary Discussion with additional references, Supplementary Figures S1 to S10 with Legends and Supplementary Tables 1 to 4. Supplementary Figures and Supplementary Tables present additional controls and data that support both the main text and supplementary discussion. Supplementary Figure S10 depicts a scheme summarizing the proposed mechanism. Supplementary Figures 1a, 3d, 7c, e and 8a-d were replaced on 21 Jan 2010. (PDF 15174 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Métivier, R., Gallais, R., Tiffoche, C. et al. Cyclical DNA methylation of a transcriptionally active promoter. Nature 452, 45–50 (2008). https://doi.org/10.1038/nature06544

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature06544

This article is cited by

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.

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