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Abstract

The folding of genomic DNA from the beads-on-a-string-like structure of nucleosomes into higher-order assemblies is crucially linked to nuclear processes. Here we calculate 3D structures of entire mammalian genomes using data from a new chromosome conformation capture procedure that allows us to first image and then process single cells. The technique enables genome folding to be examined at a scale of less than 100 kb, and chromosome structures to be validated. The structures of individual topological-associated domains and loops vary substantially from cell to cell. By contrast, A and B compartments, lamina-associated domains and active enhancers and promoters are organized in a consistent way on a genome-wide basis in every cell, suggesting that they could drive chromosome and genome folding. By studying genes regulated by pluripotency factor and nucleosome remodelling deacetylase (NuRD), we illustrate how the determination of single-cell genome structure provides a new approach for investigating biological processes.

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Change history

  • 05 April 2017

    The graph for chromosome 14 in Extended Data Fig. 6 was corrected.

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Acknowledgements

We thank A. Riddell for cell sorting, P. Humphreys for confocal microscopy, A. Peter Gunnarson for the density mapping software, the CRUK Cambridge Institute for DNA sequencing, T. Nagano and P. Fraser for processing the preliminary haploid mouse ES cells, and W. Dean, S. Schoenfelder and S. Wingett for advice. We thank the Wellcome Trust (082010/Z/07/Z), the EC FP7 4DCellFate project (277899) and the MRC (MR/M010082/1) for financial support.

Author information

Author notes

    • Martin Leeb
    • , Lluis Morey
    •  & Anton Wutz

    Present addresses: Max F. Perutz Laboratories, University of Vienna, Vienna Biocenter, Dr. Bohr-Gasse 9/3, 1030 Vienna, Austria (M.L.); Sylvester Comprehensive Cancer Center, University of Miami Miller School of Medicine, Department of Human Genetics, Miami, Florida 33136, USA (L.M.); Inst. f. Molecular Health Sciences, ETH Zurich, HPL E 12, Otto-Stern-Weg 7, 8093 Zürich, Switzerland (A.W.).

    • Tim J. Stevens
    • , David Lando
    •  & Srinjan Basu

    These authors contributed equally to this work.

Affiliations

  1. Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 1GA, UK

    • Tim J. Stevens
    • , David Lando
    • , Srinjan Basu
    • , Liam P. Atkinson
    • , Yang Cao
    • , Kai J. Wohlfahrt
    • , Wayne Boucher
    • , Aoife O’Shaughnessy-Kirwan
    • , Brian Hendrich
    •  & Ernest D. Laue
  2. MRC Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge Biomedical Campus, Cambridge CB2 0QH, UK

    • Tim J. Stevens
  3. Department of Chemistry, University of Cambridge, Lensfield Road, Cambridge CB2 1EW, UK

    • Steven F. Lee
    • , Matthieu G. S. Palayret
    •  & Dave Klenerman
  4. Wellcome Trust – MRC Stem Cell Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QR, UK

    • Martin Leeb
    • , Aoife O’Shaughnessy-Kirwan
    • , Julie Cramard
    • , Meryem Ralser
    • , Anton Wutz
    •  & Brian Hendrich
  5. EMBL-CRG Systems Biology Unit, Centre for Genomic Regulation (CRG), 08003 Barcelona, Spain

    • Andre J. Faure
    • , Enrique Blanco
    • , Lluis Morey
    • , Miriam Sansó
    • , Ben Lehner
    •  & Luciano Di Croce
  6. Universitat Pompeu Fabra, 08003 Barcelona, Spain

    • Ben Lehner
    •  & Luciano Di Croce
  7. Institució Catalana de Recerca i Estudis Avançats (ICREA), 08010 Barcelona, Spain

    • Ben Lehner
    •  & Luciano Di Croce

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Contributions

D.L., S.B. and Y.C. developed the protocol and carried out imaging/Hi-C processing. T.J.S. developed the software with assistance from L.P.A., W.B. and K.J.W. A.O’S.-K., J.C., M.R. and B.H. carried out the CHD4/MBD3 depletion experiments, associated RNA-seq and ChIP–seq, and created the mEos3.2-Halo tagged ES cell lines. M.L. and A.W. provided the initial samples of haploid mouse ES cells. S.F.L., M.G.S.P. and D.K. designed and built the microscope. L.M., M.S. and L.D.C. carried out ChIP–seq and RNA-seq experiments, while A.J.F., E.B. and B.L. carried out bioinformatics analysis. T.J.S. and E.D.L. designed experiments, analysed the results and wrote the manuscript with contributions from all the other authors.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Ernest D. Laue.

Reviewer Information Nature thanks W. Huber and the other anonymous reviewer(s) for their contribution to the peer review of this work.

Extended data

Supplementary information

PDF files

  1. 1.

    Supplementary Information

    This file contains a description of the protocols for cell culture, imaging and single-cell Hi-C, as well as the procedures used for processing the Hi-C data, and calculating/analysing the 3D structures. Details of how to download the software are also provided.

Videos

  1. 1.

    Five superimposed 3D structures of the intact mouse ES cell genome for Cell 1

    An expanding view of the chromosomes, which are coloured differently, is shown – see Figs. 1c, 2c for more details.

  2. 2.

    Five superimposed 3D structures of chromosome 10 from the intact mouse ES cell genome for Cell 1.

    Five superimposed 3D structures of chromosome 10 from the intact mouse ES cell genome for Cell 1, coloured from red through to purple (centromere to telomere) – see Fig. 1c for more details.

  3. 3.

    Five superimposed 3D structures of the intact mouse ES cell genome for Cell 1

    Five superimposed 3D structures of the intact mouse ES cell genome for Cell 1, with the chromosomes coloured from red through to purple (centromere to telomere) – see Fig. 2a for more details.

  4. 4.

    3D structure of the intact mouse ES genome for Cell 1

    3D structure of the intact mouse ES genome for Cell 1, with expanding views of the spatial distribution of the A (blue) and B (red) compartments – see Figs. 2c,d,e for more details.

  5. 5.

    3D structure of chromosome 10 from the intact mouse ES cell genome for Cell 1

    3D structure of chromosome 10 from the intact mouse ES cell genome for Cell 1, illustrating the way the chromosome contributes to the A/B compartments – see Fig. 2f for more details.

  6. 6.

    3D structure of the intact mouse ES genome for Cell 1

    3D structure of the intact mouse ES genome for Cell 1, with expanding views of the spatial distribution of the constitutive lamin-associated domains (cLADs) (yellow) and regions containing highly expressed genes (blue) – see Fig. 2e for more details.

  7. 7.

    3D structure of chromosome 10 from the intact mouse ES cell genome for Cell 1

    3D structure of chromosome 10 from the intact mouse ES cell genome for Cell 1, illustrating the way the chromosome contributes to the spatial distribution of the constitutive lamin-associated domains (cLADs) (yellow) and regions containing highly expressed genes (blue) – see Fig. 2g for more details.

  8. 8.

    Close up views of the 3D structures of two B compartment TADs (Region 1 in Fig. 5a for Cell 4).

    The TADs are coloured according to whether they are in the A (blue) or B (red) compartments, with white indicating a transitional segment (between A and B).

  9. 9.

    Close up views of the 3D structures of two B compartment TADs (Region 1 in Fig. 5a for Cell 5).

    The TADs are coloured according to whether they are in the A (blue) or B (red) compartments, with white indicating a transitional segment (between A and B).

  10. 10.

    Close up views of the 3D structures of TADs either side of an A/B compartment boundary (Region 2 in Fig. 5a for Cell 4).

    The TADs are coloured according to whether they are in the A (blue) or B (red) compartments, with white indicating a transitional segment (between A and B).

  11. 11.

    Close up views of the 3D structures of TADs either side of an A/B compartment boundary (Region 2 in Fig. 5a for Cell 5).

    The TADs are coloured according to whether they are in the A (blue) or B (red) compartments, with white indicating a transitional segment (between A and B).

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DOI

https://doi.org/10.1038/nature21429

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