Repressive but not activating epigenetic modifications are aberrant on the inactive X chromosome in live cloned cattle

X inactivation is the process of a chromosome-wide silencing of the majority of genes on the X chromosome during early mammalian development. This process may be aberrant in cloned animals. Here we show that repressive modifications, such as methylation of DNA, and the presence of methylated histones, H3K9me2 and H3K27me3, exhibit distinct aberrance on the inactive X chromosome in live clones. In contrast, H3K4me3, an active gene marker, is obviously missing from the inactive X chromosome in all cattle studied. This suggests that the disappearance of active histone modifications (H3K4me3) seems to be more important for X inactivation than deposition of marks associated with heterochromatin (DNA methylation, H3K27me3 and H3K9me2). It also implies that even apparently normal clones may have subtle abnormalities in repressive, but not activating epigenetic modifications on the inactive X when they survive to term. We also found that the histone H3 methylations were enriched and co-localized at q21-31 of the active X chromosome, which may be associated with an abundance of LINE1 repeat elements.     

Homologous recombination is reduced in female embryonic stem cells by two active X chromosomes

The reactivation of X‐linked genes is observed in some primary breast tumors. Two active X chromosomes are also observed in female embryonic stem cells (ESCs), but whether double doses of X‐linked genes affect DNA repair efficiency remains unclear. Here, we establish isogenic female/male ESCs and show that the female ESCs are more sensitive to camptothecin and have lower gene targeting efficiency than male ESCs, suggesting that homologous recombination (HR) efficiency is reduced in female ESCs. We also generate Xist‐inducible female ESCs and show that the lower HR efficiency is restored when X chromosome inactivation is induced. Finally, we assess the X‐linked genes with a role in DNA repair and find that Brcc3 is one of the genes involved in a network promoting proper HR. Our findings link the double doses of X‐linked genes with lower DNA repair activity, and this may have relevance for common diseases in female patients, such as breast cancer.     

Human Embryonic Stem Cells Do Not Change Their X Inactivation Status during Differentiation

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Xist-mediated chromatin changes that establish silencing of an entire X chromosome in mammals

X chromosome inactivation (XCI) ensures an equal gene dosage between the sexes in placental mammals. Xist, a modular multi-domain X-encoded long non-coding RNA coats the X chromosome in cis during XCI. Xist recruits chromatin remodelers and repressor complexes ensuring silencing of the inactive X (Xi). Here, we review the recent work focused on the role of Xist functional repeats and interacting RNA-binding factors in the establishment of the silent state. Xist orchestrates recruitment of remodelers and repressors that first facilitate removal of the active chromatin landscape and subsequently direct the transition into a repressive heterochromatic environment. Some of these factors affect silencing on a chromosome-wide scale, while others display gene-specific silencing defects. The temporal order of recruitment shows each silencing step is party dependent on one another. After the Xi is established, many of the factors are dispensable, and a different repertoire of proteins ensure the silenced Xi is maintained and propagated.     

RNAi in X inactivation: contrasting findings on the role of interference

X inactivation is the process that brings about the dosage equivalence of X‐linked genes in females to that of males. This complex process initiated at a very early stage of female embryonic development is orchestrated by long non‐coding RNAs transcribed in both sense and antisense orientation. Recent studies present contradicting evidence for the role of small RNAs and RNase III enzyme Dicer in the X inactivation process. In this review, I discuss these results in the overall perspective of X inactivation and gene silencing.     

Proximity-dependent recruitment of Polycomb repressive complexes by the lncRNA Airn

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An embryonic story: Analysis of the gene regulative network controlling Xist expression in mouse embryonic stem cells

In mice, dosage compensation of X‐linked gene expression is achieved through the inactivation of one of the two X‐chromosomes in XX female cells. The complex epigenetic process leading to X‐inactivation is largely controlled by Xist and Tsix, two non‐coding genes of opposing function. Xist RNA triggers X‐inactivation by coating the inactive X, while Tsix is critical for the designation of the active X‐chromosome through cis‐repression of Xist RNA accumulation. Recently, a plethora of trans‐acting factors and cis‐regulating elements have been suggested to act as key regulators of either Xist, Tsix or both; these include ubiquitous factors such as Yy1 and Ctcf, developmental proteins such as Nanog, Oct4 and Sox2, and X‐linked regulators such as Rnf12. In this paper we summarise recent advances in our knowledge of the regulation of Xist and Tsix in embryonic stem (ES) and differentiating ES cells.