Letter | Published:

Spatial partitioning of the regulatory landscape of the X-inactivation centre

Nature volume 485, pages 381385 (17 May 2012) | Download Citation

Abstract

In eukaryotes transcriptional regulation often involves multiple long-range elements and is influenced by the genomic environment1. A prime example of this concerns the mouse X-inactivation centre (Xic), which orchestrates the initiation of X-chromosome inactivation (XCI) by controlling the expression of the non-protein-coding Xist transcript. The extent of Xic sequences required for the proper regulation of Xist remains unknown. Here we use chromosome conformation capture carbon-copy (5C)2 and super-resolution microscopy to analyse the spatial organization of a 4.5-megabases (Mb) region including Xist. We discover a series of discrete 200-kilobase to 1 Mb topologically associating domains (TADs), present both before and after cell differentiation and on the active and inactive X. TADs align with, but do not rely on, several domain-wide features of the epigenome, such as H3K27me3 or H3K9me2 blocks and lamina-associated domains. TADs also align with coordinately regulated gene clusters. Disruption of a TAD boundary causes ectopic chromosomal contacts and long-range transcriptional misregulation. The Xist/Tsix sense/antisense unit illustrates how TADs enable the spatial segregation of oppositely regulated chromosomal neighbourhoods, with the respective promoters of Xist and Tsix lying in adjacent TADs, each containing their known positive regulators. We identify a novel distal regulatory region of Tsix within its TAD, which produces a long intervening RNA, Linx. In addition to uncovering a new principle of cis-regulatory architecture of mammalian chromosomes, our study sets the stage for the full genetic dissection of the X-inactivation centre.

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Accessions

Primary accessions

Gene Expression Omnibus

Data deposits

High-throughput data are deposited in Gene ExpressionOmnibus under accession number GSE35721 for all 5C experiments and GSE34243 for expression microarrays.

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Acknowledgements

We thank T. Pollex and T. Forné for experimental help; the imaging facility PICTIBiSA@BDD for technical assistance, D. Gentien and C. Hego for microarray hybridizations. We thank K. Bernhard, F. Stewart and A. Smith for protocols and material for 2i culture and EpiSC differentiation. We are grateful to members of the E.H. laboratory for critical input. This work was funded by grants from the Ministère de la Recherche et de l’Enseignement Supérieur and the ARC (to E.P.N.); a HFSP Long term fellowship (LT000597/2010-L) (to E.G.S.). EU EpiGeneSys FP7 Network of Excellence no. 257082, the Fondation pour la Recherche Medicale, ANR, ERC Advanced Investigator award no. 250367 and EU FP7 SYBOSS grant no. 242129 (to E.H.). N.B. was supported by BMBF (FORSYS) and EMBO (fellowship ASTF 307-2011). J.D., B.R.L. and N.L.v.B. were supported by NIH (R01 HG003143) and a W. M. Keck Foundation Distinguished Young Scholar Award.

Author information

Author notes

    • Bryan R. Lajoie
    • , Edda G. Schulz
    •  & Luca Giorgetti

    These authors contributed equally to this work.

Affiliations

  1. Institut Curie, 26 rue d'Ulm, Paris F-75248, France

    • Elphège P. Nora
    • , Edda G. Schulz
    • , Luca Giorgetti
    • , Ikuhiro Okamoto
    • , Nicolas Servant
    • , Tristan Piolot
    • , Emmanuel Barillot
    •  & Edith Heard
  2. CNRS UMR3215, Paris F-75248, France

    • Elphège P. Nora
    • , Edda G. Schulz
    • , Luca Giorgetti
    • , Ikuhiro Okamoto
    • , Tristan Piolot
    •  & Edith Heard
  3. INSERM U934, Paris F-75248, France

    • Elphège P. Nora
    • , Edda G. Schulz
    • , Luca Giorgetti
    • , Ikuhiro Okamoto
    • , Tristan Piolot
    •  & Edith Heard
  4. Programs in Systems Biology and Gene Function and Expression, Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts 01605-0103, USA

    • Bryan R. Lajoie
    • , Nynke L. van Berkum
    •  & Job Dekker
  5. INSERM U900, Paris, F-75248 France

    • Nicolas Servant
    •  & Emmanuel Barillot
  6. Mines ParisTech, Fontainebleau, F-77300 France

    • Nicolas Servant
    •  & Emmanuel Barillot
  7. Institute of Pathology, Charité–Universitätsmedizin, 10117 Berlin, and Institute of Theoretical Biology Humboldt Universität, 10115 Berlin, Germany

    • Johannes Meisig
    •  & Nils Blüthgen
  8. Department of Biochemistry and Biophysics, University of California San Francisco, San Francisco, California 94158-2517, USA

    • John Sedat
  9. Department of Reproduction and Development, Erasmus MC, University Medical Center, 3000 CA Rotterdam, The Netherlands

    • Joost Gribnau

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Contributions

E.P.N. performed and analysed 3C, 5C, (RT–)qPCR, immunofluorescence, RNA and DNA FISH. B.R.L. and N.L.v.B. helped in the design and/or the analysis of 3C and 5C. L.G. performed 3C, FISH and 5C analysis. E.G.S. generated the time-course transcriptomic data, which was analysed by J.M. and N.B.; I.O. performed FISH on pre-implantation embryos. J.G. donated the XTX mouse ESC line. N.S. and E.B. helped in the epigenomic and 5C analyses. J.S. and T.P. set up OMX microscopy and analysis and T.P. performed structured illumination microscopy and image analysis. The manuscript was written by E.P.N., J.D. and E.H. with contribution from E.G.S. and input from all authors.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Job Dekker or Edith Heard.

Supplementary information

PDF files

  1. 1.

    Supplementary Information

    This file contains Supplementary Figures 1-15, Supplementary Methods and additional references.

Zip files

  1. 1.

    Supplementary Data 1

    This file contains mapped 5C counts.

  2. 2.

    Supplementary Data 2

    This file contains intraTAD 5C peaks.

Excel files

  1. 1.

    Supplementary Table 1

    This file contains 5C Oligos and restriction Fragment.

  2. 2.

    Supplementary Table 2

    This file contains cDNA microarrays.

  3. 3.

    Supplementary Table 3

    This file contains the sequencing log.

  4. 4.

    Supplementary Table 4

    This file contains qPCR Primers.

About this article

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DOI

https://doi.org/10.1038/nature11049

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