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The pluripotent genome in three dimensions is shaped around pluripotency factors


It is becoming increasingly clear that the shape of the genome importantly influences transcription regulation. Pluripotent stem cells such as embryonic stem cells were recently shown to organize their chromosomes into topological domains that are largely invariant between cell types1,2. Here we combine chromatin conformation capture technologies with chromatin factor binding data to demonstrate that inactive chromatin is unusually disorganized in pluripotent stem-cell nuclei. We show that gene promoters engage in contacts between topological domains in a largely tissue-independent manner, whereas enhancers have a more tissue-restricted interaction profile. Notably, genomic clusters of pluripotency factor binding sites find each other very efficiently, in a manner that is strictly pluripotent-stem-cell-specific, dependent on the presence of Oct4 and Nanog protein and inducible after artificial recruitment of Nanog to a selected chromosomal site. We conclude that pluripotent stem cells have a unique higher-order genome structure shaped by pluripotency factors. We speculate that this interactome enhances the robustness of the pluripotent state.

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Figure 1: Inactive regions lack specific long-range interactions in embryonic stem cells.
Figure 2: Expressed Nanog gene shows preferential interaction with other pluripotency genes.
Figure 3: Spatial interactome of chromatin factors is revealed by PE-SCAn.
Figure 4: Pluripotency factors influence the 3D organization of the genome.

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Gene Expression Omnibus

Data deposits

4C sequencing data andmapped wig files have been submitted to the Gene Expression Omnibus (GEO) under accession number GSE37275.


  1. Dixon, J. R. et al. Topological domains in mammalian genomes identified by analysis of chromatin interactions. Nature 485, 376–380 (2012)

    CAS  ADS  Article  Google Scholar 

  2. Nora, E. P. et al. Spatial partitioning of the regulatory landscape of the X-inactivation centre. Nature 485, 381–385 (2012)

    CAS  ADS  Article  Google Scholar 

  3. de Wit, E. & de Laat, W. A decade of 3C technologies: insights into nuclear organization. Genes Dev. 26, 11–24 (2012)

    CAS  Article  Google Scholar 

  4. van Steensel, B. & Dekker, J. Genomics tools for unraveling chromosome architecture. Nature Biotechnol. 28, 1089–1095 (2010)

    CAS  Article  Google Scholar 

  5. Splinter, E. & de Laat, W. The complex transcription regulatory landscape of our genome: control in three dimensions. EMBO J. 30, 4345–4355 (2011)

    CAS  Article  Google Scholar 

  6. Simonis, M. et al. Nuclear organization of active and inactive chromatin domains uncovered by chromosome conformation capture-on-chip (4C). Nature Genet. 38, 1348–1354 (2006)

    CAS  Article  Google Scholar 

  7. Lieberman-Aiden, E. et al. Comprehensive mapping of long-range interactions reveals folding principles of the human genome. Science 326, 289–293 (2009)

    CAS  ADS  Article  Google Scholar 

  8. Dekker, J., Rippe, K., Dekker, M. & Kleckner, N. Capturing chromosome conformation. Science 295, 1306–1311 (2002)

    CAS  ADS  Article  Google Scholar 

  9. Splinter, E. et al. The inactive X chromosome adopts a unique three-dimensional conformation that is dependent on Xist RNA. Genes Dev. 25, 1371–1383 (2011)

    CAS  Article  Google Scholar 

  10. Splinter, E., de Wit, E., van de Werken, H. J., Klous, P. & de Laat, W. Determining long-range chromatin interactions for selected genomic sites using 4C-seq technology: from fixation to computation. Methods 58, 221–230 (2012)

    CAS  Article  Google Scholar 

  11. Wutz, A., Rasmussen, T. P. & Jaenisch, R. Chromosomal silencing and localization are mediated by different domains of Xist RNA. Nature Genet. 30, 167–174 (2002)

    CAS  Article  Google Scholar 

  12. McLean, C. Y. et al. GREAT improves functional interpretation of cis-regulatory regions. Nature Biotechnol. 28, 495–501 (2010)

    CAS  Article  Google Scholar 

  13. Apostolou, E. et al. Genome-wide chromatin interactions of the Nanog locus in pluripotency, differentiation, and reprogramming. Cell Stem Cell 12, 699–712 (2013)

    CAS  Article  Google Scholar 

  14. Handoko, L. et al. CTCF-mediated functional chromatin interactome in pluripotent cells. Nature Genet. 43, 630–638 (2011)

    CAS  Article  Google Scholar 

  15. Lin, Y. C. et al. Global changes in the nuclear positioning of genes and intra- and interdomain genomic interactions that orchestrate B cell fate. Nature Immunol. 13, 1196–1204 (2012)

    CAS  Article  Google Scholar 

  16. Li, G. et al. Extensive promoter-centered chromatin interactions provide a topological basis for transcription regulation. Cell 148, 84–98 (2012)

    CAS  Article  Google Scholar 

  17. Shen, Y. et al. A map of the cis-regulatory sequences in the mouse genome. Nature 488, 116–120 (2012)

    CAS  ADS  Article  Google Scholar 

  18. Niwa, H., Miyazaki, J. & Smith, A. G. Quantitative expression of Oct-3/4 defines differentiation, dedifferentiation or self-renewal of ES cells. Nature Genet. 24, 372–376 (2000)

    CAS  Article  Google Scholar 

  19. Chambers, I. et al. Nanog safeguards pluripotency and mediates germline development. Nature 450, 1230–1234 (2007)

    CAS  ADS  Article  Google Scholar 

  20. Holwerda, S. J. et al. Allelic exclusion of the immunoglobulin heavy chain locus is independent of its nuclear localization in mature B cells. Nucleic Acids Res. (7 June 2013)

  21. Schoenfelder, S. et al. Preferential associations between co-regulated genes reveal a transcriptional interactome in erythroid cells. Nature Genet. 42, 53–61 (2010)

    CAS  Article  Google Scholar 

  22. Xu, M. & Cook, P. R. Similar active genes cluster in specialized transcription factories. J. Cell Biol. 181, 615–623 (2008)

    CAS  Article  Google Scholar 

  23. Dhar, S. S. & Wong-Riley, M. T. Chromosome conformation capture of transcriptional interactions between cytochrome c oxidase genes and genes of glutamatergic synaptic transmission in neurons. J. Neurochem. 115, 676–683 (2010)

    CAS  Article  Google Scholar 

  24. Krijger, P. H. & de Laat, W. Identical cells with different 3D genomes; cause and consequences? Curr. Opin. Genet. Dev. 23, 191–196 (2013)

    CAS  Article  Google Scholar 

  25. Noordermeer, D. et al. Variegated gene expression caused by cell-specific long-range DNA interactions. Nature Cell Biol. 13, 944–951 (2011)

    CAS  Article  Google Scholar 

  26. Kind, J. et al. Single-cell dynamics of genome-nuclear lamina interactions. Cell 153, 178–192 (2013)

    CAS  Article  Google Scholar 

  27. Feng, B. et al. Reprogramming of fibroblasts into induced pluripotent stem cells with orphan nuclear receptor Esrrb. Nature Cell Biol. 11, 197–203 (2009)

    CAS  Article  Google Scholar 

  28. Karantzali, E. et al. Sall1 regulates embryonic stem cell differentiation in association with nanog. J. Biol. Chem. 286, 1037–1045 (2011)

    CAS  Article  Google Scholar 

  29. Parisi, S. et al. Klf5 is involved in self-renewal of mouse embryonic stem cells. J. Cell Sci. 121, 2629–2634 (2008)

    CAS  Article  Google Scholar 

  30. Smith, A. G. Culture and differentiation of embryonic stem cells. J. Tissue Cult. Methods 13, 89–94 (1991)

    Article  Google Scholar 

  31. Peric-Hupkes, D. et al. Molecular maps of the reorganization of genome-nuclear lamina interactions during differentiation. Mol. Cell 38, 603–613 (2010)

    CAS  Article  Google Scholar 

  32. Warlich, E. et al. Lentiviral vector design and imaging approaches to visualize the early stages of cellular reprogramming. Mol. Ther. 19, 782–789 (2011)

    CAS  Article  Google Scholar 

  33. Andrews, N. C. & Faller, D. V. A rapid micropreparation technique for extraction of DNA-binding proteins from limiting numbers of mammalian cells. Nucleic Acids Res. 19, 2499 (1991)

    CAS  Article  Google Scholar 

  34. Laemmli, U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227, 680–685 (1970)

    CAS  ADS  Article  Google Scholar 

  35. Wilson, A. A. et al. Sustained expression of α1-antitrypsin after transplantation of manipulated hematopoietic stem cells. Am. J. Respir. Cell Mol. Biol. 39, 133–141 (2008)

    CAS  Article  Google Scholar 

  36. Marson, A. et al. Connecting microRNA genes to the core transcriptional regulatory circuitry of embryonic stem cells. Cell 134, 521–533 (2008)

    CAS  Article  Google Scholar 

  37. 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  ADS  Article  Google Scholar 

  38. Kagey, M. H. et al. Mediator and cohesin connect gene expression and chromatin architecture. Nature 467, 430–435 (2010)

    CAS  ADS  Article  Google Scholar 

  39. Mikkelsen, T. S. et al. Genome-wide maps of chromatin state in pluripotent and lineage-committed cells. Nature 448, 553–560 (2007)

    CAS  ADS  Article  Google Scholar 

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

    CAS  ADS  Article  Google Scholar 

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We would like to thank C. Vermeulen and S. Holwerda for counting FISH slides; G. Geeven for the analysis of sequencing data; J. Brandsma for technical assistance; P. Verschure for the LacR–GFP backbone construct; and H. Niwa for providing ZHBTc4 ES cells. We also thank the Netherlands Institute for Regenerative Medicine (NIRM) network for supporting the R.A.P. laboratory and the Medical Research Council UK for supporting the I.C. laboratory. This work was financially supported by grants from the Dutch Scientific Organization (NWO) to E.d.W. (700.10.402, ‘Veni’) and W.d.L. (91204082 and 935170621), InteGeR FP7 Marie Curie ITN (PITN-GA-2007-214902) and a European Research Council Starting Grant (209700, ‘4C’) to W.d.L.

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Authors and Affiliations



E.d.W. conceived the study, analysed the data and wrote the manuscript. B.A.M.B. designed and performed reprogramming and knockout experiments, and helped to write the manuscript. Y.Z. and P.H.L.K. designed and performed LacR–Nanog experiments. E.S. and P.H.L.K. performed cell culture and 4C experiments. M.J.A.M.V., E.P.N. and E.H. designed, performed and analysed FISH experiments. M.W. and N.G. assisted with reprogramming experiments. R.A.P. shared Oct4 conditional knockout cells and assisted with depletion experiments. N.F. and I.C. shared conditional knockout cells and assisted with Nanog depletion experiments. W.d.L. conceived the study and wrote the manuscript

Corresponding author

Correspondence to Wouter de Laat.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Figures

This file contains Supplementary Figures 1-16. (PDF 10504 kb)

Supplementary Table 1

This file contains read distributions and 4C experiment characteristics for ESC, NPC, iPSC and astrocytes. (XLS 34 kb)

Supplementary Table 2

This file contains an overview of interchromosomal interactions with Nanog in ESCs. (XLS 37 kb)

Supplementary Table 3

This file contains GREAT enrichment scores for analyzed viewpoints and tissues. (XLS 85 kb)

Supplementary Table 4

This file contains 3D DNA FISH distance scores. (XLSX 30 kb)

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de Wit, E., Bouwman, B., Zhu, Y. et al. The pluripotent genome in three dimensions is shaped around pluripotency factors. Nature 501, 227–231 (2013).

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