Fig. 5: Integrated analysis reveals key DNMT3B regulated genes associated with neuro-ectodermal lineage commitment.

a Multiple Factor Analysis (MFA) approach on integrated RNA and WGBS sequencing data show discrimination between ESC, EpiLC and ME stages and a divergent differentiation trajectory between 3BKO and WT cells. b Trends of gene expression and DNAme dynamics in WT (grey) and 3BKO (orange) cells for the 615 DNMT3B target genes. Dark lines represent median of gene expression (top panel, as scaled RPKM) and DMRs methylation (bottom panel, as %), shaded areas represent the interquartile range. Genes are divided as early, medium and late according to when the biggest difference in transcription between 3BKO and WT occurs (indicated by the vertical line). While transcription of these genes shows distinct dynamical patterns of induction, DNAme depositions on their associated regulatory region is mostly established at the EpiLC stage. c, Integrated heatmaps of RNA-seq (left) and WGBS data (right) of differentially induced genes in medium and late stages of differentiation, displaying also differential methylation in regulatory regions (promoters/enhancers/super-enhancers). Each row represents a z-score of normalized logRPKM (for RNA-seq) and average % of DNAme (for WGBS) of the associated DMRs. The last (rightmost) 1-column heatmap indicates the regulatory region. Key neuro-ectodermal markers are indicated on the left. d Heatmap showing adjusted p-values (hypergeometric test as implemented in the ClusterProfiler package) of GO terms for enriched biological processes in early, medium and late DNMT3B target genes.