Fig. 5: ERK-Deficient Pbx1-KO EpiSCs exhibit distinct chromatin and DNA methylation profiles at ExM lineage genes prior to differentiation-associated ExM defects. | Nature Communications

Fig. 5: ERK-Deficient Pbx1-KO EpiSCs exhibit distinct chromatin and DNA methylation profiles at ExM lineage genes prior to differentiation-associated ExM defects.

From: Cell-context response to germ layer differentiation signals is predetermined by the epigenome in regionalized epiblast populations

Fig. 5

a Illustration depicting of E7.5 ExM lineages: allantois, amnion, and yolk sac influenced by BMP signalling. b Confocal 3D projection of E7.5 control and Pbx1f/f;Pbx2-/-;Sox2Cre/+ embryos stained with the allantois and amnion marker HAND1 (red), DE and node marker FOXA2 (green), and DAPI (grey). N, Node; DE, definitive endoderm; ExM, extraembryonic mesoderm. Images are representative of three independent experiments. WT Scale bar: 100 μm, Pbx1/2-Sox2Cre/+ or 50 μm. c Schematics of WT and Pbx1-KO EpiSCs differentiation protocol52 to PPS and ExM cells. d Heatmap shows lineage-specific DEDG (P ≤ 0.05, fold-change ≥1.5, n = 2 biological replicates) across stages (yellow: EpiSC, grey: PPS, red: ExM). Significance was assessed by DESeq2 on the basis of two-sided Wald test with Benjamini–Hochberg adjusted P-values. e Pbx1-KO EpiSCs show a preferential bias towards primitive erythroid and yolk sac mesenchyme when subjected to ExM differentiation. f Schematic showing DMGs in WT vs Pbx1-KO EpiSCs and their transcriptional changes upon differentiation in PPS and ExM. Yellow, grey, and red circles represent DMGs in EpiSCs, DEGs in PPS, and DEGs in ExM, respectively, illustrating that methylation changes in EpiSCs precede lineage-specific gene activation. g WGBS (SeqMonk) visualization indicating differentially methylated regions across the Hand1 locus in WT and Pbx1-KO EpiSCs, showing hypermethylation in Pbx1-KO EpiSCs. The bar shows CpG methylation levels (%). h UCSC visualization of Hand1 showing increased chromatin accessibility in WT conditions, while the locus remains closed in Pbx1-KO. Accessible loci are indicated with red arrowheads, and reduced accessibility with white arrowheads. i WGBS (SeqMonk) tracks indicating CpG methylation levels across Tbx3 in WT and Pbx1-KO EpiSCs, showing hypermethylation in WT EpiSCs. CpG methylation levels (%) are depicted in the bottom bar. j ATAC-seq tracks of Tbx3 show increased chromatin accessibility in Pbx1-KO vs WT cells. Red arrowheads mark accessible loci; white indicates reduced accessibility.

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