Abstract
Repression of maternal Xist (Xm-Xist) during preimplantation in mouse embryos is essential for establishing imprinted X chromosome inactivation. Nuclear transplantation (NT) studies using nuclei derived from non-growing (ng) and full-grown (fg) oocytes have indicated that maternal-specific repressive modifications are imposed on Xm-Xist during oogenesis, as well as on autosomal imprinted genes. Recent studies have revealed that histone H3 lysine 9 trimethylation (H3K9me3) enrichments on Xm-Xist promoter regions are involved in silencing at the preimplantation stages. However, whether H3K9me3 is imposed on Xm-Xist during oogenesis is not known. Here, we dissected the chromatin states in ng and fg oocytes and early preimplantation stage embryos. Chromatin immunoprecipitation experiments against H3K9me3 revealed that there was no significant enrichment within the Xm-Xist region during oogenesis. However, NT embryos with ng nuclei (ngNT) showed extensive Xm-Xist derepression and H3K9me3 hypomethylation of the promoter region at the 4-cell stage, which corresponds to the onset of paternal Xist expression. We also found that the chromatin state at the Xist genomic locus became markedly condensed as oocyte growth proceeded. Although the condensed Xm-Xist genomic locus relaxed during early preimplantation phases, the extent of the relaxation across Xm-Xist loci derived from normally developed oocytes was significantly smaller than those of paternal-Xist and ngNT-Xist genomic loci. Furthermore, Xm-Xist from 2-cell metaphase nuclei became derepressed following NT. We propose that chromatin condensation is associated with imprinted Xist repression and that skipping of the condensation step by NT leads to Xist activation during the early preimplantation phase.
Highlights
Expression of the large non-coding RNA X inactive specific transcript (Xist) is essential for the initiation of X chromosome inactivation (XCI) in female mice and humans (Augui et al, 2011; Sado and Sakaguchi, 2013; Lee, 2011)
H3K9me3 is comparable between ng and fg oocytes at XmXist loci We initially confirmed that Rnf12 is highly expressed during oogenesis (Fig. S1), as described elsewhere (Shin et al, 2010), indicating that the Xist repressive state is established prior to oocyte maturation
We previously demonstrated that H3K9me3 is essential for X chromosome (Xm)-Xist repression in preimplantation embryos (Fukuda et al, 2014)
Summary
Expression of the large non-coding RNA X inactive specific transcript (Xist) is essential for the initiation of X chromosome inactivation (XCI) in female mice and humans (Augui et al, 2011; Sado and Sakaguchi, 2013; Lee, 2011). Xist expression is initiated around the 4-cell stage and is restricted to the paternal allele. Using parthenogenetic embryos, which are composed of two maternal genomes, we previously demonstrated that histone 3 lysine 9 trimethylation (H3K9me3) is essential for Xm-Xist repression during early preimplantation phases (Fukuda et al, 2014)
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