Chromatin structure analysis of the mouse
            <i>Xist</i>
            locus

McCabe, Veronica; Formstone, Emma J.; O’Neill, Laura P.; Turner, Bryan M.; Brockdorff, Neil

doi:10.1073/pnas.96.13.7155

Cited by 12 publications

(6 citation statements)

References 46 publications

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“…The differences between alleles on Xi and Xa were again confined to the promoter region, and the overall acetylation levels were comparable between the two alleles outside of the promoter. These results are in agreement with a recent analysis of the acetylation profile of the Xist locus in male and female mouse cells (27).…”

Section: Figsupporting

confidence: 82%

Promoter-specific hypoacetylation of X-inactivated genes

Gilbert

Sharp

1999

Proc. Natl. Acad. Sci. U.S.A.

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The histone H4 acetylation status of the active X (Xa) and inactive X (Xi) chromosomes was investigated at the level of individual genes. A moderate level of acetylation was observed along the lengths of genes on both the Xi and Xa, regardless of their X inactivation status. However, this moderate level of acetylation was modified specifically in promoter regions. Transcriptionally active genes showed elevated levels of acetylation at their promoters on both the Xi and Xa. In contrast, promoters of X-inactivated genes were markedly hypoacetylated, which coincided with the methylation of adjacent CG dinucleotides. This promoterspecific hypoacetylation may be a key component of an X inactivation machinery that operates at the level of individual genes. In mammals, X chromosome dosage is equalized between males and females by X inactivation, the transcriptional silencing of one of the two X chromosomes in female cells. The chromatin of the inactive X (Xi) is distinct from all other chromosomes, including the active X (Xa), in that it is late replicating (1, 2), methylated on CG dinucleotides (3, 4), enriched in the histone H2A variant macroH2A (5), and hypoacetylated on histones H2A, H3, and H4 (6-8). These unique features of the Xi chromatin are associated with, and may contribute to, the silencing of its several thousand resident genes.Although most genes on the Xi are transcriptionally repressed, there are clear exceptions. First, the XIST gene, which initiates X inactivation, is transcribed only from the Xi (9). Second, in both humans and mice, a number of genes escape X inactivation-that is, they are transcribed from the Xi as well as the Xa (for review see refs. 10 and 11). Genes that escape X inactivation typically have homologs on the Y chromosome, which in theory obviates their need for dosage compensation (12).Among the unique properties of the Xi chromatin, CG methylation and late replication have been studied at the level of individual genes and were found to correlate closely with gene expression. Where examined, these traits are associated with particular genes that are silenced, but not with those that escape X inactivation (2,13,14). In contrast, histone acetylation on the Xi has not been examined extensively at the level of individual genes. The prevailing view that the Xi is hypoacetylated comes from immunofluorescence microscopy analysis, which showed that all chromosomes except the Xi stained brightly with antisera against acetylated histones in female cells (6-8). However, these microscopy studies on metaphase chromosomes are very limited in resolution and thus cannot address the acetylation status of individual genes.A chromatin immunoprecipitation (IP) technique has been developed recently that allows the acetylation status of individual nucleosomes to be examined (15-17). We used this method to investigate the acetylation status of histone H4 associated with numerous genes on both the Xi and Xa. Our results indicate that nucleosomes on the Xi are in fact acetylated along the lengths of genes...

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Section: Figsupporting

confidence: 82%

Promoter-specific hypoacetylation of X-inactivated genes

Gilbert

Sharp

1999

Proc. Natl. Acad. Sci. U.S.A.

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“…Conversely, promoters could be shut down after the required period of expression by tether-directed local modification, such as histone deacetylation. One example of such local modification is the hypoacetylation of histone H4 at the inactive Xist promoter in male mice, contrasting with higher average levels of acetylation in female mice, in which one of the two alleles is active (28). However, in both sexes, similar average levels of histone hyperacetylation are observed over the remainder of the locus.…”

Section: Piggybacking: a Mechanism For Coupling Chromatin Modificatiomentioning

confidence: 89%

Chromatin modification by DNA tracking

Travers

1999

Proc. Natl. Acad. Sci. U.S.A.

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In general, the transcriptional competence of a chromatin domain is correlated with increased sensitivity to DNase I cleavage. A recent observation that actively transcribing RNA polymerase II piggybacks a histone acetyltranferase activity [Wittschieben, B., Otero, G., de Bizemont, T., Fellows, J., Erdjument-Bromage, H., Ohba, R., Li, Y., Allis, C. D., Tempst, P. & Svejstrup, J. Q. (1999) Mol. Cell 4, 123-128] implies that the state of histone acetylation, and hence the ability of chromatin to fold, can be altered by a processive mechanism. In this article, it is proposed that tracking-mediated chromatin modification could create and͞or maintain an open configuration in a complete chromatin domain including both intra-and extragenic regions. This mechanism suggests a putative functional role for the extragenic transcription observed at the ␤-globin and other loci in vertebrate cells.T he transcriptional competence of eukaryotic chromatin requires access both for the activating gene-specific factors and for RNA polymerase II and its associated protein complexes. The reorganization of chromatin associated with activation can be mediated by large remodeling complexes and is also strongly correlated with histone hyperacetylation (1-3). The state of histone modification can modulate both the access of the transcription machinery to regulatory regions and the effectiveness of chromatin as a template for RNA synthesis. This modification is believed to stabilize the unfolded state of chromatin by antagonizing internucleosomal interactions (4) and so could facilitate the passage of the transcribing enzyme, especially in long transcription units. This article summarizes recent findings suggesting that the elongating RNA polymerase II may itself directly mediate histone acetylation and consequent chromatin unfolding by piggybacking an acetyltranferase activity. The possible implications of this observation with respect to domain opening are discussed.Local and Extensive Chromatin Modification. In Saccharomyces, several studies suggest that the extent of histone modification mediated by transcription factor-targeted histone acetyltranferases and deacetyltranferases, such as Gcn5 and Rpd3, respectively, is restricted to chromatin in the immediate vicinity of regulatory regions (5-7). In these examples, the histones of only one or, at most, two nucleosomes are modified whereas those associated with the downstream transcribed region are unaffected and could still present a barrier to efficient transcription elongation. This localized modification also contrasts with the pattern observed in certain chromatin domains of higher eukaryotes. Functionally a chromatin domain, such as that typified by the chicken ␤-globin locus, constitutes a region of positionindependent gene expression and is delimited by boundary or insulator elements that confer this property (8). Early studies of the avian globin, lysozyme, and ovalbumin loci showed that the transcriptional competence of a domain is correlated with an increased sensitivity to cle...

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“…Is the deacetylated status of Xist gene chromatin on the active X chromosome (23,30) altered by the absence of Mbd2? An antibody against the acetylated N-terminal tail of histone H4, a marker of transcriptionally active chromatin, precipitated equivalent low levels of the methylated Xist CpG island from wt and Mbd2 Ϫ/Ϫ cells in a ChIP experiment (Fig.…”

Section: Resultsmentioning

confidence: 99%

Mbd2 Contributes to DNA Methylation-Directed Repression of the Xist Gene

Barr

Hermann

Berger

et al. 2007

Molecular and Cellular Biology

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Transcription of the Xist gene triggers X chromosome inactivation in cis and is therefore silenced on the X chromosome that remains active. DNA methylation contributes to this silencing, but the mechanism is unknown. As methylated DNA binding proteins (MBPs) are potential mediators of gene silencing by DNA methylation, we asked whether MBP-deficient cell lines could maintain Xist repression. The absence of Mbd2 caused significant low-level reactivation of Xist, but silencing was restored by exogenous Mbd2. In contrast, deficiencies of Mbd1, MeCP2, and Kaiso had no detectable effect, indicating that MBPs are not functionally redundant at this locus. Xist repression in Mbd2-null cells was hypersensitive to the histone deacetylase inhibitor trichostatin A and to depletion of the DNA methyltransferase Dnmt1. These synergies implicate Mbd2 as a mediator of the DNA methylation signal at this locus. The presence of redundant mechanisms to enforce repression at Xist and other loci is compatible with the hypothesis that "stacking" of imperfect repressive tendencies may be an evolutionary strategy to ensure leakproof gene silencing.

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Chromatin structure analysis of the mouse Xist locus

Cited by 12 publications

References 46 publications

Promoter-specific hypoacetylation of X-inactivated genes

Promoter-specific hypoacetylation of X-inactivated genes

Chromatin modification by DNA tracking

Mbd2 Contributes to DNA Methylation-Directed Repression of the Xist Gene

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