Fig. 4: Loss of ISL1 impairs amnion signals essential for mesoderm formation.

See also Supplementary Fig. 8 and Supplementary Data 5. a–d Immunofluorescent staining of major cell types on sections of ISL1 mutant embryos at Day 14. n = 3 mutant embryos. a OCT4, VIM, and GATA3; b BRA and MIXL1; c ISL1; d GABRP. Scale bar 50 µm. Numbers represent section numbers. Arrows indicate signals of interest and dashed lines mark the amniotic cavity. e GO categories enriched among genes significantly downregulated in cells of the mesodermal clusters in mutant embryos ordered by false discovery rate (FDR). n = 4 wild-type and 4 mutant embryos collected in two batches. f STRING network of all significantly downregulated genes in cells of the mesodermal clusters in the mutant embryos. Nodes not connected to the main network have been removed. Nodes belonging to the GO category embryonic development colored in blue; nodes belonging to the STRING network cluster Wnt signaling pathway, and TGF-beta signaling pathway colored in red. g Volcano plot showing the DEGs between amnion (AM-1 and AM-2) of the wild-type and mutant embryos. Gray areas indicate the expected group mean difference in random cell subsets (99.9th percentile) and a false discovery rate cutoff of 1%. Red labeling indicates that the gene is part of the ISL1 regulon identified by SCENIC. P values were calculated using a two-tailed Welch’s t test. The x-axis reports uncorrected p values. h Violin plot of the expression levels of BMP4 and WNT6 in the different cell types identified in (Fig. 1d) at Day 14 separated between wild-type (blue) and mutant (red) embryos. Epi epiblast, AM-1 amnion 1, AM-2 amnion 2, meso-1 mesoderm 1, meso-2 mesoderm 2, wt wild type, mt mutant, D dorsal, V ventral.