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Conditional deletion of Xist disrupts histone macroH2A localization but not maintenance of X inactivation

Nature Genetics volume 22, pages 323–324 (1999)Cite this article

In mammalian females, dosage compensation is achieved by X-chromosome inactivation. Xist has an essential role in the initiation of the inactivation process1,2. Xist is transcribed exclusively from the inactive X (Xi; Refs 3,4), and the RNA stays in the nucleus and coats the chromosome in cis5,6. Continued transcription of Xist and its close association with Xi also implies a role for Xist in maintenance of X inactivation6. Previous studies showed that a fragment of the human Xi lacking XIST remains inactive in mouse-human somatic hybrid cells7 or in leukaemia cells with X-chromosomal translocations8. The cells used in these studies were highly selected, and might not truly represent X inactivation in wild-type cells. Indeed, XIST RNA does not localize to Xi in mouse-human hybrids9. Therefore, we sought to address the role of Xist in primary mouse embryonic fibroblasts of normal karyotype using the Cre-loxP system.

Xist-deficient fibroblasts appeared to proliferate normally and senesced at the same time as controls (data not shown). To determine if loss of Xist led to reactivation of X-linked genes, we performed RNA FISH with probes for the X-linked genes Pgk1 and Hprt as well as Xist. In wild-type fibroblasts and fibroblasts carrying Xist2lox, we detected a Pgk1 signal coming from the active X (Xa) in approximately 50% of nuclei and an Hprt signal in 60%. After Cre-mediated deletion of Xist, we did not see biallelic expression of X-linked genes or a signal in a higher percentage of nuclei, suggesting that the mutant Xi remains transcriptionally silent (Fig. 1c,d).

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Figure 1: Conditional deletion of Xist.
Figure 2: In the absence of Xist RNA, late replication and hypoacetylation on histone H4 are maintained, but preferential association of histone macroH2A1.2 with Xi is not.

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Acknowledgements

We thank J. Dausmann for blastocyst injections; R. Curry for help with maintaining the mouse colony; and A. Wutz, S. Cherry and T. Rasmussen for critical reading of the manuscript and discussions. This work was supported by a grant to R.J. from the National Institute of Health/National Cancer Institute (R35-CA44339).

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Authors and Affiliations

  1. Whitehead Institute for Biomedical Research, Cambridge, 02142, Massachusetts, USA

    Györgyi Csankovszki, Barbara Panning, Brian Bates & Rudolf Jaenisch

  2. Department of Biology, Massachusetts Institute of Technology, Cambridge, 02142, Massachusetts, USA

    Györgyi Csankovszki & Rudolf Jaenisch

  3. Department of Animal Biology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, 19104, Pennsylvania, USA

    John R. Pehrson

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Correspondence to Rudolf Jaenisch.

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Csankovszki, G., Panning, B., Bates, B. et al. Conditional deletion of Xist disrupts histone macroH2A localization but not maintenance of X inactivation. Nat Genet 22, 323–324 (1999). https://doi.org/10.1038/11887

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