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:

Human DNA methylomes at base resolution show widespread epigenomic differences

Abstract

DNA cytosine methylation is a central epigenetic modification that has essential roles in cellular processes including genome regulation, development and disease. Here we present the first genome-wide, single-base-resolution maps of methylated cytosines in a mammalian genome, from both human embryonic stem cells and fetal fibroblasts, along with comparative analysis of messenger RNA and small RNA components of the transcriptome, several histone modifications, and sites of DNA–protein interaction for several key regulatory factors. Widespread differences were identified in the composition and patterning of cytosine methylation between the two genomes. Nearly one-quarter of all methylation identified in embryonic stem cells was in a non-CG context, suggesting that embryonic stem cells may use different methylation mechanisms to affect gene regulation. Methylation in non-CG contexts showed enrichment in gene bodies and depletion in protein binding sites and enhancers. Non-CG methylation disappeared upon induced differentiation of the embryonic stem cells, and was restored in induced pluripotent stem cells. We identified hundreds of differentially methylated regions proximal to genes involved in pluripotency and differentiation, and widespread reduced methylation levels in fibroblasts associated with lower transcriptional activity. These reference epigenomes provide a foundation for future studies exploring this key epigenetic modification in human disease and development.

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: Global trends of human DNA methylomes.
Figure 2: Bisulphite-PCR validation of non-CG DNA methylation in differentiated and stem cells.
Figure 3: Non-CG DNA methylation in H1 embryonic stem cells.
Figure 4: Density of DNA methylation at sites of DNA–protein interaction.
Figure 5: Cell-type variation in DNA methylation.
Figure 6: Clustering of genomic, epigenetic and transcriptional features at differentially methylated regions.

Similar content being viewed by others

Accession codes

Primary accessions

Gene Expression Omnibus

Data deposits

Sequence data is available under the GEO accessions GSM429321-23, GSM432685-92, GSM438361-64, GSE17917, GSE18292 and GSE16256, and the SRA accessions SRX006782-89, SRX006239-41, SRX007165.1-68.1 and SRP000941. Analysed data sets can be obtained from http://neomorph.salk.edu/human_methylome.

References

  1. Holliday, R. & Pugh, J. E. DNA modification mechanisms and gene activity during development. Science 187, 226–232 (1975)

    Article  ADS  CAS  Google Scholar 

  2. Riggs, A. D. X inactivation, differentiation, and DNA methylation. Cytogenet. Cell Genet. 14, 9–25 (1975)

    Article  CAS  Google Scholar 

  3. Bestor, T. H. The DNA methyltransferases of mammals. Hum. Mol. Genet. 9, 2395–2402 (2000)

    Article  CAS  Google Scholar 

  4. Li, E., Bestor, T. H. & Jaenisch, R. Targeted mutation of the DNA methyltransferase gene results in embryonic lethality. Cell 69, 915–926 (1992)

    Article  CAS  Google Scholar 

  5. Lippman, Z. et al. Role of transposable elements in heterochromatin and epigenetic control. Nature 430, 471–476 (2004)

    Article  ADS  CAS  Google Scholar 

  6. Okano, M., Bell, D. W., Haber, D. A. & Li, E. DNA methyltransferases Dnmt3a and Dnmt3b are essential for de novo methylation and mammalian development. Cell 99, 247–257 (1999)

    Article  CAS  Google Scholar 

  7. Reik, W. Stability and flexibility of epigenetic gene regulation in mammalian development. Nature 447, 425–432 (2007)

    Article  ADS  CAS  Google Scholar 

  8. Straussman, R. et al. Developmental programming of CpG island methylation profiles in the human genome. Nature Struct. Mol. Biol. 16, 564–571 (2009)

    Article  CAS  Google Scholar 

  9. Weber, M. & Schübeler, D. Genomic patterns of DNA methylation: targets and function of an epigenetic mark. Curr. Opin. Cell Biol. 19, 273–280 (2007)

    Article  CAS  Google Scholar 

  10. Cedar, H. & Bergman, Y. Linking DNA methylation and histone modification: patterns and paradigms. Nature Rev. Genet. 10, 295–304 (2009)

    Article  CAS  Google Scholar 

  11. Rauch, T. A., Wu, X., Zhong, X., Riggs, A. D. & Pfeifer, G. P. A human B cell methylome at 100-base pair resolution. Proc. Natl Acad. Sci. USA 106, 671–678 (2009)

    Article  ADS  CAS  Google Scholar 

  12. Ball, M. et al. Targeted and genome-scale strategies reveal gene-body methylation signatures in human cells. Nature Biotechnol. 27, 361–368 (2009)

    Article  ADS  CAS  Google Scholar 

  13. Deng, J. et al. Targeted bisulfite sequencing reveals changes in DNA methylation associated with nuclear reprogramming. Nature Biotechnol. 27, 353–360 (2009)

    Article  CAS  Google Scholar 

  14. Meissner, A. et al. Genome-scale DNA methylation maps of pluripotent and differentiated cells. Nature 454, 766–770 (2008)

    Article  ADS  CAS  Google Scholar 

  15. Lister, R. et al. Highly integrated single-base resolution maps of the epigenome in Arabidopsis . Cell 133, 523–536 (2008)

    Article  CAS  Google Scholar 

  16. Cokus, S. J. et al. Shotgun bisulphite sequencing of the Arabidopsis genome reveals DNA methylation patterning. Nature 452, 215–219 (2008)

    Article  ADS  CAS  Google Scholar 

  17. Thomson, J. A. et al. Embryonic stem cell lines derived from human blastocysts. Science 282, 1145–1147 (1998)

    Article  ADS  CAS  Google Scholar 

  18. Nichols, W. W. et al. Characterization of a new human diploid cell strain, IMR-90. Science 196, 60–63 (1977)

    Article  ADS  CAS  Google Scholar 

  19. Ramsahoye, B. H. et al. Non-CpG methylation is prevalent in embryonic stem cells and may be mediated by DNA methyltransferase 3a. Proc. Natl Acad. Sci. USA 97, 5237–5242 (2000)

    Article  ADS  CAS  Google Scholar 

  20. Woodcock, D. M., Crowther, P. J. & Diver, W. P. The majority of methylated deoxycytidines in human DNA are not in the CpG dinucleotide. Biochem. Biophys. Res. Commun. 145, 888–894 (1987)

    Article  CAS  Google Scholar 

  21. Aoki, A. et al. Enzymatic properties of de novo-type mouse DNA (cytosine-5) methyltransferases. Nucleic Acids Res. 29, 3506–3512 (2001)

    Article  CAS  Google Scholar 

  22. Gowher, H. & Jeltsch, A. Enzymatic properties of recombinant Dnmt3a DNA methyltransferase from mouse: the enzyme modifies DNA in a non-processive manner and also methylates non-CpA sites. J. Mol. Biol. 309, 1201–1208 (2001)

    Article  CAS  Google Scholar 

  23. Gonzalo, S. et al. DNA methyltransferases control telomere length and telomere recombination in mammalian cells. Nature Cell Biol. 8, 416–424 (2006)

    Article  CAS  Google Scholar 

  24. Steinert, S., Shay, J. W. & Wright, W. E. Modification of subtelomeric DNA. Mol. Cell. Biol. 24, 4571–4580 (2004)

    Article  CAS  Google Scholar 

  25. Brunner, A. L. et al. Distinct DNA methylation patterns characterize differentiated human embryonic stem cells and developing human fetal liver. Genome Res. 19, 1044–1056 (2009)

    Article  CAS  Google Scholar 

  26. Ferguson-Smith, A. C. & Greally, J. Epigenetics: perceptive enzymes. Nature 449, 148–149 (2007)

    Article  ADS  CAS  Google Scholar 

  27. Jia, D., Jurkowska, R. Z., Zhang, X., Jeltsch, A. & Cheng, X. Structure of Dnmt3a bound to Dnmt3L suggests a model for de novo DNA methylation. Nature 449, 248–251 (2007)

    Article  ADS  CAS  Google Scholar 

  28. Bell, A. C. & Felsenfeld, G. Methylation of a CTCF-dependent boundary controls imprinted expression of the Igf2 gene. Nature 405, 482–485 (2000)

    Article  ADS  CAS  Google Scholar 

  29. Clark, S. J., Harrison, J. & Molloy, P. L. Sp1 binding is inhibited by (m)Cp(m)CpG methylation. Gene 195, 67–71 (1997)

    Article  CAS  Google Scholar 

  30. Hark, A. T. et al. CTCF mediates methylation-sensitive enhancer-blocking activity at the H19/Igf2 locus. Nature 405, 486–489 (2000)

    Article  ADS  CAS  Google Scholar 

  31. Kitazawa, S., Kitazawa, R. & Maeda, S. Transcriptional regulation of rat cyclin D1 gene by CpG methylation status in promoter region. J. Biol. Chem. 274, 28787–28793 (1999)

    Article  CAS  Google Scholar 

  32. Mancini, D. N., Singh, S. M., Archer, T. K. & Rodenhiser, D. I. Site-specific DNA methylation in the neurofibromatosis (NF1) promoter interferes with binding of CREB and SP1 transcription factors. Oncogene 18, 4108–4119 (1999)

    Article  CAS  Google Scholar 

  33. Johnson, D. S., Mortazavi, A., Myers, R. M. & Wold, B. Genome-wide mapping of in vivo protein-DNA interactions. Science 316, 1497–1502 (2007)

    Article  ADS  CAS  Google Scholar 

  34. Heintzman, N. et al. Histone modifications at human enhancers reflect global cell-type-specific gene expression. Nature 459, 108–112 (2009)

    Article  ADS  CAS  Google Scholar 

  35. Schmidl, C. et al. Lineage-specific DNA methylation in T cells correlates with histone methylation and enhancer activity. Genome Res. 19, 1165–1174 (2009)

    Article  CAS  Google Scholar 

  36. Johnson, L. M. et al. The SRA methyl-cytosine-binding domain links DNA and histone methylation. Curr. Biol. 17, 379–384 (2007)

    Article  CAS  Google Scholar 

  37. Jones, P. A. & Baylin, S. B. The epigenomics of cancer. Cell 128, 683–692 (2007)

    Article  CAS  Google Scholar 

  38. Hellman, A. & Chess, A. Gene body-specific methylation on the active X chromosome. Science 315, 1141–1143 (2007)

    Article  ADS  CAS  Google Scholar 

  39. The International Stem Cell Initiative Characterization of human embryonic stem cell lines by the International Stem Cell Initiative. Nature Biotechnol. 25, 803–816 (2007)

    Article  Google Scholar 

  40. Villesen, P., Aagaard, L., Wiuf, C. & Pedersen, F. S. Identification of endogenous retroviral reading frames in the human genome. Retrovirology 1, 32 (2004)

    Article  Google Scholar 

  41. Chan, S. W. Inputs and outputs for chromatin-targeted RNAi. Trends Plant Sci. 13, 383–389 (2008)

    Article  CAS  Google Scholar 

  42. Azuara, V. et al. Chromatin signatures of pluripotent cell lines. Nature Cell Biol. 8, 532–538 (2006)

    Article  CAS  Google Scholar 

  43. Bernstein, B. E. et al. A bivalent chromatin structure marks key developmental genes in embryonic stem cells. Cell 125, 315–326 (2006)

    Article  CAS  Google Scholar 

  44. Ludwig, T. et al. Feeder-independent culture of human embryonic stem cells. Nature Methods 3, 637–646 (2006)

    Article  CAS  Google Scholar 

  45. Ludwig, T. et al. Derivation of human embryonic stem cells in defined conditions. Nature Biotechnol. 24, 185–187 (2006)

    Article  CAS  Google Scholar 

  46. Langmead, B., Trapnell, C., Pop, M. & Salzberg, S. L. Ultrafast and memory-efficient alignment of short DNA sequences to the human genome. Genome Biol. 10, R25 (2009)

    Article  Google Scholar 

Download references

Acknowledgements

We thank A. Elwell and A. Hernandez for assistance with sequence library preparation and Illumina sequencing. R.L. is supported by a Human Frontier Science Program Long-term Fellowship. R.D.H. is supported by an American Cancer Society Postdoctoral Fellowship. This work was supported by grants from the following: Mary K. Chapman Foundation, The National Institutes of Health (U01 ES017166 and U01 1U01ES017166-01), the California Institute for Regenerative Medicine (RS1-00292-1), the Australian Research Council Centre of Excellence Program (CE0561495, DP0771156) and Morgridge Institute for Research, Madison, Wisconsin. We thank the NIH Roadmap Reference Epigenome Consortium (http://nihroadmap.nih.gov/epigenomics/referenceepigenomeconsortium.asp) and C. Gunter (Hudson-Alpha Institute) for assistance. This study was carried out as part of the NIH Roadmap Epigenomics Program.

Author Contributions Experiments were designed by J.R.E., B.R., R.L., J.A.T. and R.D.H. Cells were grown by J.A.-B. and Q.-M.N. MethylC-Seq, RNA-Seq and smRNA-Seq experiments were conducted by R.L. and J.R.N. ChIP-Seq experiments were conducted by R.D.H., L.L. and Z.Y. ChIP-Seq data analysis was performed by G.H., R.D.H. and L.E. BS-PCR validation was performed by R.H.D. Sequencing data processing was performed by R.L., J.T.-F., L.E., V.R. and G.H. Bioinformatic and statistical analyses were conducted by M.P., R.L., G.H., J.T.-F., R.H.D., R.S. and A.H.M. AnnoJ development was performed by J.T.F and A.H.M. The manuscript was prepared by R.L., M.P., R.H.D., A.H.M. and J.R.E.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Joseph R. Ecker.

Supplementary information

Supplementary Information

This file contains Supplementary Figures 1-16 with Legends, Supplementary Methods, Supplementary Data and Supplementary References. (PDF 5433 kb)

Supplementary Tables

This file contains Supplementary Tables 1-12 which provide additional material that is referred to in the main article. (XLS 3523 kb)

PowerPoint slides

Rights and permissions

Reprints and permissions

About this article

Cite this article

Lister, R., Pelizzola, M., Dowen, R. et al. Human DNA methylomes at base resolution show widespread epigenomic differences . Nature 462, 315–322 (2009). https://doi.org/10.1038/nature08514

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature08514

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

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