Sequential ChIP-bisulfite sequencing enables direct genome-scale investigation of chromatin and DNA methylation cross-talk
Open accessAbstract
Cross-talk between DNA methylation and histone modifications drives the establishment of composite epigenetic signatures and is traditionally studied using correlative rather than direct approaches. Here, we present sequential ChIP-bisulfite-sequencing (ChIP-BS-seq) as an approach to quantitatively assess DNA methylation patterns associated with chromatin modifications or chromatin-associated factors directly. A chromatin-immunoprecipitation (ChIP)-capturing step is used to obtain a restricted representation of the genome occupied by the epigenetic feature of interest, for which a single-base resolution DNA methylation map is then generated. When applied to H3 lysine 27 trimethylation (H3K27me3), we found that H3K27me3 and DNA methylation are compatible throughout most of the genome, except for CpG islands, where these two marks are mutually exclusive. Further ChIP-BS-seq-based analysis in Dnmt triple-knockout (TKO) embryonic stem cells revealed that total loss of CpG methylation is associated with alteration of H3K27me3 levels throughout the genome: H3K27me3 in localized peaks is decreased while broad local enrichments (BLOCs) of H3K27me3 are formed. At an even broader scale, these BLOCs correspond to regions of high DNA methylation in wild-type ES cells, suggesting that DNA methylation prevents H3K27me3 deposition locally and at a megabase scale. Our strategy provides a unique way of investigating global interdependencies between DNA methylation and other chromatin features.
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The enrichment in EZH2 and SUZ12 binding sites is consistent with the observed enrichment of methylation differences in PRC2 repressed regions and previous observations that DNA methylation often interact with PRC2 binding.
“consistent with the observed enrichment of methylation differences in PRC2 repressed regions (Additional file 1: Fig. S3c) and previous observations that DNA methylation often interact with PRC2 binding [52, 53].”
From Sex-specific DNA methylation differences in Alzheimer’s disease pathology (2021) · cites this paper as [53]
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Fast and accurate short read alignment with Burrows-Wheeler transform
Li, H., & Durbin, R. (2009) · Bioinformatics · doi:10.1093/bioinformatics/btp324 · Cited by 58,607 · see citing works
High-Resolution Profiling of Histone Methylations in the Human Genome
Barski, A., Cuddapah, S., Cui, K., Roh, T.-Y., Schones, D. E., Wang, Z., Wei, G., Chepelev, I., & Zhao, K. (2007) · Cell · doi:10.1016/j.cell.2007.05.009 · Cited by 6,845 · see citing works
Genome-scale DNA methylation maps of pluripotent and differentiated cells
Meissner, A., Mikkelsen, T. S., Gu, H., Wernig, M., Hanna, J., Sivachenko, A., Zhang, X., Bernstein, B. E., Nusbaum, C., Jaffe, D. B., et al. (2008) · Nature · doi:10.1038/nature07107 · Cited by 2,536 · see citing works
Self-citationLinking DNA methylation and histone modification: patterns and paradigms
Cedar, H., & Bergman, Y. (2009) · Nature Reviews Genetics · doi:10.1038/nrg2540 · Cited by 2,267 · see citing works
Schlesinger, Y., Straussman, R., Keshet, I., Farkash, S., Hecht, M., Zimmerman, J., Eden, E., Yakhini, Z., Ben-Shushan, E., Reubinoff, B. E., et al. (2007) · Nature Genetics · doi:10.1038/ng1950 · Cited by 1,226 · see citing works
Ohm, J. E., McGarvey, K. M., Yu, X., Cheng, L., Schuebel, K. E., Cope, L., Mohammad, H. P., Chen, W., Daniel, V. C., Yu, W., et al. (2007) · Nature Genetics · doi:10.1038/ng1972 · Cited by 1,130 · see citing works
Karimi, M. M., Goyal, P., Maksakova, I. A., Bilenky, M., Leung, D., Tang, J. X., Shinkai, Y., Mager, D. L., Jones, S., Hirst, M., et al. (2011) · Cell Stem Cell · doi:10.1016/j.stem.2011.04.004 · Cited by 535 · cited 3× · see citing works
“As shown before (Karimi et al. 2011), loss of DNA methylation did not cause massive deregulation, and the expression levels of most genes were”
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Research works printed but not linked: 37
- Bartke T, Vermeulen M, Xhemalce B, Robson SC, Mann M, Kouzarides T 2010 Nucleosome-interacting proteins regulated by DNA and histone methylation. Cell 143: 470– 484 21029866 PMC3640253
- Bell AC, Felsenfeld G 2000 Methylation of a CTCF-dependent boundary controls imprinted expression of the Igf2 gene. Nature 405: 482– 485 10839546
- Bock C, Tomazou EM, Brinkman AB, Müller F, Simmer F, Gu H, Jäger N, Gnirke A, Stunnenberg HG, Meissner A 2010 Quantitative comparison of genome-wide DNA methylation mapping technologies. Nat Biotechnol 28: 1106– 1114 20852634 PMC3066564
- Brinkman AB, Simmer F, Ma K, Kaan A, Zhu J, Stunnenberg HG 2010 Whole-genome DNA methylation profiling using MethylCap-seq. Methods 52: 232– 236 20542119
- Denissov S, van Driel M, Voit R, Hekkelman M, Hulsen T, Hernandez N, Grummt I, Wehrens R, Stunnenberg H 2007 Identification of novel functional TBP-binding sites and general factor repertoires. EMBO J 26: 944– 954 17268553 PMC1852848
- Fischle W 2008 Talk is cheap–cross-talk in establishment, maintenance, and readout of chromatin modifications. Genes Dev 22: 3375– 3382 19141469
- Gal-Yam EN, Egger G, Iniguez L, Holster H, Einarsson S, Zhang X, Lin JC, Liang G, Jones PA, Tanay A 2008 Frequent switching of Polycomb repressive marks and DNA hypermethylation in the PC3 prostate cancer cell line. Proc Natl Acad Sci 105: 12979– 12984 18753622 PMC2529074
- Gu H, Bock C, Mikkelsen TS, Jäger N, Smith ZD, Tomazou E, Gnirke A, Lander ES, Meissner A 2010 Genome-scale DNA methylation mapping of clinical samples at single-nucleotide resolution. Nat Methods 7: 133– 136 20062050 PMC2860480
- Gu H, Smith ZD, Bock C, Boyle P, Gnirke A, Meissner A 2011 Preparation of reduced representation bisulfite sequencing libraries for genome-scale DNA methylation profiling. Nat Protoc 6: 468– 481 21412275
- Guelen L, Pagie L, Brasset E, Meuleman W, Faza MB, Talhout W, Eussen BH, de Klein A, Wessels L, de Laat W, 2008 Domain organization of human chromosomes revealed by mapping of nuclear lamina interactions. Nature 453: 948– 951 18463634
- Hark AT, Schoenherr CJ, Katz DJ, Ingram RS, Levorse JM, Tilghman SM 2000 CTCF mediates methylation-sensitive enhancer-blocking activity at the H19/Igf2 locus. Nature 405: 486– 489 10839547
- Harris RA, Wang T, Coarfa C, Nagarajan RP, Hong C, Downey SL, Johnson BE, Fouse SD, Delaney A, Zhao Y, 2010 Comparison of sequencing-based methods to profile DNA methylation and identification of monoallelic epigenetic modifications. Nat Biotechnol 28: 1097– 1105 20852635 PMC2955169
- Hawkins RD, Hon GC, Lee LK, Ngo Q, Lister R, Pelizzola M, Edsall LE, Kuan S, Luu Y, Klugman S, 2010 Distinct epigenomic landscapes of pluripotent and lineage-committed human cells. Cell Stem Cell 6: 479– 491 20452322 PMC2867844
- Kagey JD, Kapoor-Vazirani P, McCabe MT, Powell DR, Vertino PM 2010 Long-term stability of demethylation after transient exposure to 5-aza-2′-deoxycytidine correlates with sustained RNA polymerase II occupancy. Mol Cancer Res 8: 1048– 1059 20587535 PMC3086892
- Kanduri C, Pant V, Loukinov D, Pugacheva E, Qi CF, Wolffe A, Ohlsson R, Lobanenkov VV 2000 Functional association of CTCF with the insulator upstream of the H19 gene is parent of origin-specific and methylation-sensitive. Curr Biol 10: 853– 856 10899010
- Kondo Y, Shen L, Cheng AS, Ahmed S, Boumber Y, Charo C, Yamochi T, Urano T, Furukawa K, Kwabi-Addo B, 2008 Gene silencing in cancer by histone H3 lysine 27 trimethylation independent of promoter DNA methylation. Nat Genet 40: 741– 750 18488029
- Lee J-S, Smith E, Shilatifard A 2010 The language of histone crosstalk. Cell 142: 682– 685 20813257 PMC3711869
- Lindroth AM, Park YJ, McLean CM, Dokshin GA, Persson JM, Herman H, Pasini D, Miró X, Donohoe ME, Lee JT, 2008 Antagonism between DNA and H3K27 methylation at the imprinted Rasgrf1 locus. PLoS Genet 4: e1000145 doi: 10.1371/journal.pgen.1000145 18670629 PMC2475503 · DOI
- Lister R, Pelizzola M, Dowen RH, Hawkins RD, Hon G, Tonti-Filippini J, Nery JR, Lee L, Ye Z, Ngo Q-M, 2009 Human DNA methylomes at base resolution show widespread epigenomic differences. Nature 462: 315– 322 19829295 PMC2857523
- Marks H, Chow JC, Denissov S, Françoijs K-J, Brockdorff N, Heard E, Stunnenberg HG 2009 High-resolution analysis of epigenetic changes associated with X inactivation. Genome Res 19: 1361– 1373 19581487 PMC2720191
- McGarvey KM, Fahrner JA, Greene E, Martens J, Jenuwein T, Baylin SB 2006 Silenced tumor suppressor genes reactivated by DNA demethylation do not return to a fully euchromatic chromatin state. Cancer Res 66: 3541– 3549 16585178
- Meissner A, Gnirke A, Bell GW, Ramsahoye B, Lander ES, Jaenisch R 2005 Reduced representation bisulfite sequencing for comparative high-resolution DNA methylation analysis. Nucleic Acids Res 33: 5868– 5877 16224102 PMC1258174
- Mikkelsen TS, Ku M, Jaffe DB, Issac B, Lieberman E, Giannoukos G, Alvarez P, Brockman W, Kim T-K, Koche RP, 2007 Genome-wide maps of chromatin state in pluripotent and lineage-committed cells. Nature 448: 553– 560 17603471 PMC2921165
- Mohn F, Weber M, Rebhan M, Roloff TC, Richter J, Stadler MB, Bibel M, Schübeler D 2008 Lineage-specific polycomb targets and de novo DNA methylation define restriction and potential of neuronal progenitors. Mol Cell 30: 755– 766 18514006
- Pan G, Tian S, Nie J, Yang C, Ruotti V, Wei H, Jonsdottir GA, Stewart R, Thomson JA 2007 Whole-genome analysis of histone H3 lysine 4 and lysine 27 methylation in human embryonic stem cells. Cell Stem Cell 1: 299– 312 18371364
- Pauler FM, Sloane MA, Huang R, Regha K, Koerner MV, Tamir I, Sommer A, Aszodi A, Jenuwein T, Barlow DP 2009 H3K27me3 forms BLOCs over silent genes and intergenic regions and specifies a histone banding pattern on a mouse autosomal chromosome. Genome Res 19: 221– 233 19047520 PMC2652204
- Peters AHFM, Kubicek S, Mechtler K, O'Sullivan RJ, Derijck AAHA, Perez-Burgos L, Kohlmaier A, Opravil S, Tachibana M, Shinkai Y, 2003 Partitioning and plasticity of repressive histone methylation states in mammalian chromatin. Mol Cell 12: 1577– 1589 14690609
- Simon JA, Kingston RE 2009 Mechanisms of polycomb gene silencing: Knowns and unknowns. Nat Rev Mol Cell Biol 10: 697– 708 19738629
- Smith ZD, Gu H, Bock C, Gnirke A, Meissner A 2009 High-throughput bisulfite sequencing in mammalian genomes. Methods 48: 226– 232 19442738 PMC2864123
- Song Q, Smith AD 2011 Identifying dispersed epigenomic domains from ChIP-Seq data. Bioinformatics 27: 870– 871 21325299 PMC3051331
- Stadler MB, Murr R, Burger L, Ivanek R, Lienert F, Schöler A, Wirbelauer C, Oakeley EJ, Gaidatzis D, Tiwari VK, 2011 DNA-binding factors shape the mouse methylome at distal regulatory regions. Nature 480: 490– 495 22170606
- Thomson JP, Skene PJ, Selfridge J, Clouaire T, Guy J, Webb S, Kerr ARW, Deaton A, Andrews R, James KD, 2010 CpG islands influence chromatin structure via the CpG-binding protein Cfp1. Nature 464: 1082– 1086 20393567 PMC3730110
- Viré E, Brenner C, Deplus R, Blanchon L, Fraga M, Didelot C, Morey L, Van Eynde A, Bernard D, Vanderwinden J-M, 2006 The Polycomb group protein EZH2 directly controls DNA methylation. Nature 439: 871– 874 16357870
- Widschwendter M, Fiegl H, Egle D, Mueller-Holzner E, Spizzo G, Marth C, Weisenberger DJ, Campan M, Young J, Jacobs I, 2007 Epigenetic stem cell signature in cancer. Nat Genet 39: 157– 158 17200673
- Wu H, Coskun V, Tao J, Xie W, Ge W, Yoshikawa K, Li E, Zhang Y, Sun YE 2010 Dnmt3a-dependent nonpromoter DNA methylation facilitates transcription of neurogenic genes. Science 329: 444– 448 20651149 PMC3539760
- Zhang Y, Liu T, Meyer CA, Eeckhoute J, Johnson DS, Bernstein BE, Nusbaum C, Myers RM, Brown M, Li W, 2008 Model-based analysis of ChIP-Seq (MACS). Genome Biol 9: R137 doi: 10.1186/gb-2008-9-9-r137 18798982 PMC2592715 · DOI · Paper page
- Zhao XD, Han X, Chew JL, Liu J, Chiu KP, Choo A, Orlov YL, Sung W-K, Shahab A, Kuznetsov VA, 2007 Whole-genome mapping of histone H3 Lys4 and 27 trimethylations reveals distinct genomic compartments in human embryonic stem cells. Cell Stem Cell 1: 286– 298 18371363
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Feichtinger, J., Hernández, I., Fischer, C., Hanscho, M., Auer, N., Hackl, M., Jadhav, V., Baumann, M., Krempl, P. M., Schmidl, C., et al. (2016) · Biotechnology and Bioengineering · doi:10.1002/bit.25990 · Cited by 148 · see citing works
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“S3c) and previous observations that DNA methylation often interact with PRC2 binding [52, 53].”
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- Authors
- Brinkman, A. B., Gu, H., Bartels, S. J. J., Zhang, Y., Matarese, F., Simmer, F., Marks, H., Bock, C., Gnirke, A., Meissner, A., & Stunnenberg, H. G.
- Affiliations
- Radboud University Nijmegen (NL)
- Broad Institute (US)
- Corresponding author
- Stunnenberg, Hendrik G.
- ORCID
- Venue
- Genome Research
- Publisher
- Cold Spring Harbor Laboratory
- Published
- 2012-03-30
- Volume
- 22
- Issue
- 6
- Pages
- 1128-1138
- ISSN
- 1088-9051 (ISSN-L)
- PMC
- PMC3371717
- OpenAlex
- W2116816603
- Cited by
- 413
- References
- 7
- Citations by year
- 2026: 18 · 2025: 19 · 2024: 19 · 2023: 17 · 2022: 32 · 2021: 23 · 2020: 23 · 2019: 34 · 2018: 22 · 2017: 31 · 2016: 37 · 2015: 33 · 2014: 41 · 2013: 44 · 2012: 19
- Type
- posted content
- Language
- en
- Open access
- bronze
- doi.org
- Locations
- Genome Research (publishedVersion)
- Europe PMC (PubMed Central) (publishedVersion, cc-by-nc)
- PubMed Central (publishedVersion, cc-by-nc)
- PubMed (publishedVersion)
- Digital Access to Scholarship at Harvard (DASH) (Harvard University) (submittedVersion, cc-by)
- HighWire Press Open Archive (submittedVersion)
- Radboud Repository (Radboud University) (submittedVersion)
- Radboud Repository (Radboud University) (submittedVersion, other-oa)
- Funders
- European Commission (200620)
- Nederlandse Organisatie voor Wetenschappelijk Onderzoek (864-05-002)
- National Institutes of Health
- Harvard Stem Cell Institute
- National Institute of Environmental Health Sciences (U01ES017155)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Epigenetics and DNA Methylation
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Genomics and Chromatin Dynamics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Chromatin Remodeling and Cancer
- Keywords
- DNA methylation, histone modifications, H3K27me3, chromatin modification, CpG islands, DNA demethylation, epigenetic signatures
- MeSH terms
- Animals, Cell Line, Chromatin, Colonic Neoplasms, Histones, Humans, Lysine, Sulfites, Sequence Analysis, DNA, CpG Islands, DNA Methylation, Genomics, Epigenesis, Genetic, Cell Line, Tumor, Chromatin Immunoprecipitation, Mice, Embryonic Stem Cells, Gene Knockout Techniques
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