Lineage-specific regulation of imprinted X inactivation in extraembryonic endoderm stem cells

Abstract

BackgroundSilencing of the paternal X chromosome (Xp), a phenomenon known as imprinted X-chromosome inactivation (I-XCI), characterises, amongst mouse extraembryonic lineages, the primitive endoderm and the extraembryonic endoderm (XEN) stem cells derived from it.ResultsUsing a combination of chromatin immunoprecipitation characterisation of histone modifications and single-cell expression studies, we show that whilst the Xp in XEN cells, like the inactive X chromosome in other cell types, globally accumulates the repressive histone mark H3K27me3, a large number of Xp genes locally lack H3K27me3 and escape from I-XCI. In most cases this escape is specific to the XEN cell lineage. Importantly, the degree of escape and the genes concerned remain unchanged upon XEN conversion into visceral endoderm, suggesting stringent control of I-XCI in XEN derivatives. Surprisingly, chemical inhibition of EZH2, a member of the Polycomb repressive complex 2 (PRC2), and subsequent loss of H3K27me3 on the Xp, do not drastically perturb the pattern of silencing of Xp genes in XEN cells.ConclusionsThe observations that we report here suggest that the maintenance of gene expression profiles of the inactive Xp in XEN cells involves a tissue-specific mechanism that acts partly independently of PRC2 catalytic activity.

Establishment of X chromosome inactivation and epigenomic features of the inactive X depend on cellular contexts

“…These studies revealed peculiar features of imprinted Xi chromatin and lower stability of iXCI. Enrichment of H3K27me3 on the Xi has been observed in all cells from the extra‐embryonic lineages , yet, to a lower extent compared to a randomly inactivated X: approximately 76–80% of X‐linked genes display female‐specific enrichment of H3K27me3 in XEN and TS cells, while this number reaches 93% in liver cells . This is in agreement with the observation that levels of H3K27me3 are globally lower in TS and XEN cells compared with post‐implantation embryonic stages .…”

“…The inactive state of the Xp is, however, passed on and maintained, at least for the period of in utero life, in cells of the extra‐embryonic lineage. The maintenance and stability of iXCI in this post‐implantation context has mainly been addressed in vitro, through the analysis of cells derived from the trophectoderm (TS, trophoblast stem cells, and differentiated derivatives such as TGC, trophoblast giant cells) and from the primitive endoderm (XEN cells) – and in rare cases, from in vivo analyses . These studies revealed peculiar features of imprinted Xi chromatin and lower stability of iXCI.…”

“…It remains to be fully understood to what extent this peculiar epigenetic Xi signature in extra‐embryonic lineages is directly linked to partial lack of XCI and/or frequent reactivation events from the Xi. Indeed, more genes escape XCI in placental progenitor cells (around 13% in TS cells) than in various adult tissues . Relaxation of XCI has been observed in up to 40% of TGCs for X‐linked genes ( Huwe1 and Atrx ), which do not escape XCI in other lineages .…”

A rapid passage through a two-active-X-chromosome state accompanies the switch of imprinted X-inactivation patterns in mouse trophoblast stem cells

“…ChIP assays were performed as previously described [ 45 ] using an H3K27me3 antibody (07-449 from EMD Millipore, Billerica, MA, USA) and analysed as described previously [ 19 , 20 ]. ChIP-chip data have been deposited under Gene Expression Omnibus accession number [GSE:68536].…”

“…Paradoxically, this latter lineage is particularly rich in gene escaping from I-XCI—i.e. genes expressed from both Xs—compared to other adult cell types [ 18 , 19 ]. In addition to this, transient and spontaneous reactivations of certain X-linked genes occur both and ex vivo [ 20 ] and, after differentiation, the relaxation of I-XCI extends to additional genes in specific subtypes of placental cells [ 21 – 25 ].…”