Fig. 5: H3K4me1 is dispensable for enhancer-dependent gene expression in in vitro pluripotent cells.

a dCD cells express catalytic mutant MLL3 and MLL4 proteins without histone methyltransferase activity but capable of binding to their target sites and interacting with other proteins as part of the COMPASS complex. The different Mll3/4 catalytic mutants used in this study are shown to the right. b H3K4me1 and H3K27ac levels for the Group I and II PGCLC enhancers in R1 WT and MLL3/4 catalytic mutant ESC lines as well as upon their differentiation into d2 EpiLC and EpiSC. P-values were calculated using paired two-sided Wilcoxon tests. The H3K4me1 ChIP-seq data shown for WT d2 EpiLC and EpiSC are the same ones used in Fig. 3c as a third replicate. Scales: RPGC. c H3K4me1, H3K4me2, and H3K27ac profiles in both WT and MLL3/4 catalytic mutant cells (ESC, EpiLC, and EpiSC) around representative PGCLC (Esrrb), EpiLC (Grhl2), and EpiSC (Wnt3) genes and their associated enhancers. d Percentage of CpG methylation for the Group I and II PGCLC enhancers in WT and dCD ESC as well as upon their differentiation into d2 EpiLC. P-values were calculated using paired two-sided Wilcoxon tests. The WGBS data in 2i ESC was obtained from Skvortsova et al. 2019⁶⁴. Scale: percentage of methylated CpGs. e Expression (as measured by RNA-seq) of the genes associated with the PGCLC, EpiLC, and EpiSC enhancers in WT (gray) and dCD (red) ESC cells as well as upon their differentiation into EpiLC and EpiSC. The RNA-seq data in ESC was obtained from Dorighi et al. 2017³⁸ and the experiments in d2 EpiLC and EpiSC were performed in duplicates. The horizontal line in the boxplots indicates the median, the box indicates the first and third quartiles and the whiskers indicate ± 1.5 × interquartile range. TPM transcripts per million.