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The pluripotent genome in three dimensions is shaped around pluripotency factors

Abstract

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

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

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Acknowledgements

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

Authors

Contributions

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). https://doi.org/10.1038/nature12420

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