Transient colocalization of X-inactivation centres accompanies the initiation of X inactivation

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Abstract

The initial differential treatment of the two X chromosomes during X-chromosome inactivation is controlled by the X-inactivation centre (Xic). This locus determines how many X chromosomes are present in a cell ('counting') and which X chromosome will be inactivated in female cells ('choice'). Critical control sequences in the Xic include the non-coding RNAs Xist and Tsix, and long-range chromatin elements. However, little is known about the process that ensures that X inactivation is triggered appropriately when more than one Xic is present in a cell. Using three-dimensional fluorescence in situ hybridization (FISH) analysis, we showed that the two Xics transiently colocalize, just before X inactivation, in differentiating female embryonic stem cells. Using Xic transgenes capable of imprinted but not random X inactivation, and Xic deletions that disrupt random X inactivation, we demonstrated that Xic colocalization is linked to Xic function in random X inactivation. Both long-range sequences and the Tsix element, which generates the antisense transcript to Xist, are required for the transient interaction of Xics. We propose that transient colocalization of Xics may be necessary for a cell to determine Xic number and to ensure the correct initiation of X inactivation.

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Figure 1: Nuclear location of the Xic and Xist RNA in differentiating embryonic stem cells.
Figure 2: Xic colocalization in differentiating embryonic stem cells.
Figure 3: Analysis of inter-Xic distance distributions using quantile–quantile plots.

References

  1. 1

    Rastan, S. Non-random X-chromosome inactivation in mouse X-autosome translocation embryos — location of the inactivation centre. J. Embryol. Exp. Morphol. 78, 1–22 (1983).

  2. 2

    Rastan, S. & Robertson, E. J. X-chromosome deletions in embryo-derived (EK) cell lines associated with lack of X-chromosome inactivation. J. Embryol. Exp. Morphol. 90, 379–388 (1985).

  3. 3

    Boumil, R. M. & Lee, J. T. Forty years of decoding the silence in X-chromosome inactivation. Hum. Mol. Genet. 10, 2225–2232 (2001).

  4. 4

    Clerc, P. & Avner, P. Multiple elements within the Xic regulate random X inactivation in mice. Semin. Cell Dev. Biol. 14, 85–92 (2003).

  5. 5

    Clerc, P. & Avner, P. Role of the region 3′ to Xist exon 6 in the counting process of X-chromosome inactivation. Nature Genet. 19, 249–253 (1998).

  6. 6

    Morey, C. et al. The region 3' to Xist mediates X chromosome counting and H3 Lys-4 dimethylation within the Xist gene. EMBO J. 23, 594–604 (2004).

  7. 7

    Heard, E., Mongelard, F., Arnaud, D. & Avner, P. Xist yeast artificial chromosome transgenes function as X-inactivation centers only in multicopy arrays and not as single copies. Mol. Cell Biol. 19, 3156–3166 (1999).

  8. 8

    Marahrens, Y. X-inactivation by chromosomal pairing events. Genes Dev. 13, 2624–2632 (1999).

  9. 9

    Osborne, C. S. et al. Active genes dynamically colocalize to shared sites of ongoing transcription. Nature Genet. 36, 1065–1071 (2004).

  10. 10

    Spilianakis, C. G. et al. Interchromosomal associations between alternatively expressed loci. Nature 435, 637–645 (2005).

  11. 11

    Spector, D. L. The dynamics of chromosome organization and gene regulation. Annu. Rev. Biochem. 72, 573–608 (2003).

  12. 12

    Taddei, A., Hediger, F., Neumann, F. R. & Gasser, S. M. The function of nuclear architecture: a genetic approach. Annu. Rev. Genet. 38, 305–345 (2004).

  13. 13

    Chambeyron, S. & Bickmore, W. A. Does looping and clustering in the nucleus regulate gene expression? Curr. Opin. Cell Biol. 16, 256–262 (2004).

  14. 14

    Comings, D. E. The rationale for an ordered arrangement of chromatin in the interphase nucleus. Am. J. Hum. Genet. (1968).

  15. 15

    Okamoto, I. et al. Evidence for de novo imprinted X-chromosome inactivation independent of meiotic inactivation in mice. Nature 438, 369–373 (2005).

  16. 16

    Morey, C., Arnaud, D., Avner, P. & Clerc, P. Tsix-mediated repression of Xist accumulation is not sufficient for normal random X inactivation. Hum. Mol. Genet. 10, 1403–1411 (2001).

  17. 17

    Navarro, P. et al. Tsix transcription across the Xist gene alters chromatin conformation without affecting Xist transcription: implications for X-chromosome inactivation. Genes Dev. 19, 1474–1484 (2005).

  18. 18

    Sado, T., Hoki, Y. & Sasaki, H. Tsix silences Xist through modification of chromatin structure. Dev. Cell 9, 159–165 (2005).

  19. 19

    Stavropoulos, N., Rowntree, R. K. & Lee, J. T. Identification of developmentally specific enhancers for Tsix in the regulation of X chromosome inactivation. Mol. Cell. Biol. 25, 2757–2769 (2005).

  20. 20

    LaSalle, J. M. & Lalande, M. Homologous association of oppositely imprinted chromosomal domains. Science 272, 725–728 (1996).

  21. 21

    Rougeulle, C. et al. Differential histone H3 Lys-9 and Lys-27 methylation profiles on the X chromosome. Mol. Cell. Biol. 24, 5475–5484 (2004).

  22. 22

    Bacher, C. P. et al. 4-D single particle tracking of synthetic and proteinaceous microspheres reveals preferential movement of nuclear particles along chromatin — poor tracks. BMC Cell Biol. 5, 45 (2004).

  23. 23

    Tvarusko, W. et al. Time-resolved analysis and visualization of dynamic processes in living cells. Proc. Natl Acad. Sci. USA 96, 7950–7955 (1999).

  24. 24

    Hollander, M. & Wolfe, D. A. in Nonparametric statistical inference 27–33 (John Wiley & Sons, New York, 1973).

  25. 25

    Conover, W. J. in Practical nonparametric statistics (ed. Wiley, B.) 295–301 & 309–314 (John Wiley & Sons, New York, 1971).

  26. 26

    Becker, R. A., Chambers, J. M. & Wilks, A. R. The New S Language (Wadsworth & Brooks/Cole, 1988).

  27. 27

    Penny, G. et al. Requirement for Xist in X chromosome inactivation. Nature 379, 131–137 (1996).

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Acknowledgements

We would like to thank A. Belmont, D. Spector and N. Mise for helpful comments on the manuscript and P. Le Baccon for support with image analysis. This project was supported by a Human Frontier Science Program (HFSP) research grant to E.H. and R.E. R.E. also acknowledges support on multi-dimensional image acquisition from Leica Microsystems CMS GmbH, Mannheim, Germany. Support to E.H. was also provided by the Schlumberger Foundation, the Centre National de la Recherche Scientifiques (CNRS) and the Curie Institute (Program Incitatif et Collaboratif). E.H. and P.A. are also supported by the EU Network of Excellence (Epigenome).

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Correspondence to Roland Eils or Edith Heard.

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