Dynamic CpG island methylation landscape in oocytes and preimplantation embryos


Elucidating how and to what extent CpG islands (CGIs) are methylated in germ cells is essential to understand genomic imprinting and epigenetic reprogramming1,2,3. Here we present, to our knowledge, the first integrated epigenomic analysis of mammalian oocytes, identifying over a thousand CGIs methylated in mature oocytes. We show that these CGIs depend on DNMT3A and DNMT3L4,5 but are not distinct at the sequence level, including in CpG periodicity6. They are preferentially located within active transcription units and are relatively depleted in H3K4me3, supporting a general transcription-dependent mechanism of methylation. Very few methylated CGIs are fully protected from post-fertilization reprogramming but, notably, the majority show incomplete demethylation in embryonic day (E) 3.5 blastocysts. Our study shows that CGI methylation in gametes is not entirely related to genomic imprinting but is a strong factor in determining methylation status in preimplantation embryos, suggesting a need to reassess mechanisms of post-fertilization demethylation.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Figure 1: DNA methylation landscape in oocytes and sperm determined by RRBS.
Figure 2: Mechanism of DNA methylation establishment in oocytes.
Figure 3: Biological significance and fate of CGI methylation in oocytes.


  1. 1

    Bartolomei, M.S. Genomic imprinting: employing and avoiding epigenetic processes. Genes Dev. 23, 2124–2133 (2009).

    CAS  Article  Google Scholar 

  2. 2

    Morgan, H.D., Santos, F., Green, K., Dean, W. & Reik, W. Epigenetic reprogramming in mammals. Hum. Mol. Genet. 14, R47–R58 (2005).

    CAS  Article  Google Scholar 

  3. 3

    Sasaki, H. & Matsui, Y. Epigenetic events in mammalian germ-cell development: reprogramming and beyond. Nat. Rev. Genet. 9, 129–140 (2008).

    CAS  Article  Google Scholar 

  4. 4

    Bourc'his, D., Xu, G.-L., Lin, C.-S., Bollman, B. & Bestor, T.H. Dnmt3L and the establishment of maternal genomic imprints. Science 294, 2536–2539 (2001).

    CAS  Article  Google Scholar 

  5. 5

    Kaneda, M. et al. Essential role for de novo DNA methyltransferase Dnmt3a in paternal and maternal imprinting. Nature 429, 900–903 (2004).

    CAS  Article  Google Scholar 

  6. 6

    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).

    CAS  Article  Google Scholar 

  7. 7

    Schaefer, C.B., Ooi, S.K.T., Bestor, T.H. & Bourc'his, D. Epigenetic decisions in mammalian germ cells. Science 316, 398–399 (2007).

    CAS  Article  Google Scholar 

  8. 8

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

    CAS  Article  Google Scholar 

  9. 9

    Smith, Z.D., Gu, H., Bock, C., Gnirke, A. & Meissner, A. High-throughput bisulfite sequencing in mammalian genomes. Methods 48, 226–232 (2009).

    CAS  Article  Google Scholar 

  10. 10

    Howlett, S.K. & Reik, W. Methylation levels of maternal and paternal genomes during preimplantation development. Development 113, 119–127 (1991).

    CAS  PubMed  Google Scholar 

  11. 11

    Illingworth, R.S. et al. Orphan CpG islands identify numerous conserved promoters in the mammalian genome. PLoS Genet. 6, e1001134 (2010).

    Article  Google Scholar 

  12. 12

    Chotalia, M. et al. Transcription is required for establishment of germline methylation marks at imprinted genes. Genes Dev. 23, 105–117 (2009).

    CAS  Article  Google Scholar 

  13. 13

    Lucifero, D., Mann, M.R.W., Bartolomei, M.S. & Trasler, J.M. Gene-specific timing and epigenetic memory in oocyte imprinting. Hum. Mol. Genet. 13, 839–849 (2004).

    CAS  Article  Google Scholar 

  14. 14

    Illingworth, R. et al. A novel CpG island set identifies tissue-specific methylation at developmental gene loci. PLoS Biol. 6, e22 (2008).

    Article  Google Scholar 

  15. 15

    Maunakea, A.K. et al. Conserved role of intragenic DNA methylation in regulating alternative promoters. Nature 466, 253–257 (2010).

    CAS  Article  Google Scholar 

  16. 16

    Reinhart, B., Paoloni-Giacobino, A. & Chaillet, J.R. Specific differentially methylated domain sequences direct the maintenance of methylation at imprinted genes. Mol. Cell. Biol. 26, 8347–8356 (2006).

    CAS  Article  Google Scholar 

  17. 17

    Bock, C., Halachev, K., Buch, J. & Lengauer, T. EpiGRAPH: user-friendly software for statistical analysis and prediction of (epi)genomic data. Genome Biol. 10, R14 (2009).

    Article  Google Scholar 

  18. 18

    Hata, K., Okano, M., Lei, H. & Li, E. Dnmt3L cooperates with the Dnmt3 family of de novo DNA methyltransferases to establish maternal imprints in mice. Development 129, 1983–1993 (2002).

    CAS  PubMed  Google Scholar 

  19. 19

    Popp, C. et al. Genome-wide erasure of DNA methylation in mouse primordial germ cells is affected by AID deficiency. Nature 463, 1101–1105 (2010).

    CAS  Article  Google Scholar 

  20. 20

    Ciccone, D.N. et al. KDM1B is a histone H3K4 demethylase required to establish maternal genomic imprints. Nature 461, 415–418 (2009).

    CAS  Article  Google Scholar 

  21. 21

    Fang, R. et al. Human LSD2/KDM1b/AOF1 regulates gene transcription by modulating intragenic H3K4me2 methylation. Mol. Cell 39, 222–233 (2010).

    CAS  Article  Google Scholar 

  22. 22

    Ooi, S.K.T. et al. DNMT3L connects unmethylated lysine 4 of histone H3 to de novo methylation of DNA. Nature 448, 714–717 (2007).

    CAS  Article  Google Scholar 

  23. 23

    Zhang, Y. et al. Chromatin methylation activity of Dnmt3a and Dnmt3a/3L is guided by interaction of the ADD domain with the histone H3 tail. Nucleic Acids Res. 38, 4246–4253 (2010).

    CAS  Article  Google Scholar 

  24. 24

    Dhayalan, A. et al. The Dnmt3a PWWP domain reads histone 3 lysine 36 trimethylation and guides DNA methylation. J. Biol. Chem. 285, 26114–26120 (2010).

    CAS  Article  Google Scholar 

  25. 25

    Borgel, J. et al. Targets and dynamics of promoter DNA methylation during early mouse development. Nat. Genet. 42, 1093–1100 (2010).

    CAS  Article  Google Scholar 

  26. 26

    Tartakover-Matalon, S. et al. Impaired migration of trophoblast cells caused by simvastatin is associated with decreased membrane IGF-I receptor, MMP2 activity and HSP27 expression. Hum. Reprod. 22, 1161–1167 (2007).

    CAS  Article  Google Scholar 

  27. 27

    Hemberger, M., Dean, W. & Reik, W. Epigenetic dynamics of stem cells and cell lineage commitment: digging Waddington's canal. Nat. Rev. Mol. Cell Biol. 10, 526–537 (2009).

    CAS  Article  Google Scholar 

  28. 28

    Krueger, F. & Andrews, S.R. Bismark: A flexible aligner and methylation caller for Bisulfite-Seq applications. Bioinformatics 27, 1571–1572 (2011).

    CAS  Article  Google Scholar 

  29. 29

    Dahl, J.A. & Collas, P. A rapid micro chromatin immunoprecipitation assay (ChIP). Nat. Protoc. 3, 1032–1045 (2008).

    CAS  Article  Google Scholar 

  30. 30

    Creyghton, M.P. et al. Histone H3K27ac separates active from poised enhancers and predicts developmental state. Proc. Natl. Acad. Sci. USA 107, 21931–21936 (2010).

    CAS  Article  Google Scholar 

  31. 31

    Tomizawa, S. et al. Dynamic stage-specific changes in imprinted differentially methylated regions during early mammalian development and prevalence of non-CpG methylation in oocytes. Development 138, 811–820 (2011).

    CAS  Article  Google Scholar 

Download references


We thank K. Tabbada for technical assistance with Illumina sequencing, H. Mertani, P. Mollard, W. Dean and W. Reik for input and discussions and M. Branco and W. Reik for making available the DNMT3A conditional knockout line. This work was supported by grants G0800013 and G0801156 from the Medical Research Council to G.K. and by the Biotechnology and Biological Sciences Research Council. S.A.S. was supported by the Babraham Institute and the Centre for Trophoblast Research.

Author information




S.A.S. designed the study, performed RRBS, mRNA-Seq, direct BS-PCR experiments, data analysis and wrote the manuscript. S.-i.T. contributed to direct bisulphite sequencing PCR experiments and performed oocyte collections. F.K. and S.R.A. performed CpG methylation calls, general Illumina sequence alignments and data analysis. N.R. performed ChIP-Seq experiments. N.C. analyzed data. A.S.-P. performed statistical analysis. S.S. and K.H. provided Dnmt3L wild-type and knockout oocytes. G.K. designed and supervised the study and wrote the manuscript.

Corresponding author

Correspondence to Gavin Kelsey.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–9 and Supplementary Tables 2–4. (PDF 4729 kb)

Supplementary Table 1

CpG Island methylation calls (separate Excel file). (XLSX 13157 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Smallwood, S., Tomizawa, S., Krueger, F. et al. Dynamic CpG island methylation landscape in oocytes and preimplantation embryos. Nat Genet 43, 811–814 (2011). https://doi.org/10.1038/ng.864

Download citation

Further reading


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