KDM5B regulates H3K4 methylation during differentiation. (A) ES cells cultured in the absence of LIF for 3 to 4 days to induce differentiation. Note the loss of normal three-dimensional colony morphology of shLuc ES cells at day 3 of differentiation, while shKdm5b ES cells maintained their normal three-dimensional colony morphology in the absence of LIF. (B) Correlation between changes in histone methylation and expression level. Fold change normalized tag density ratios (shKdm5b/shLuc) of H3K4me3, H3K4me2, and H3K4me1 at all refseq genes, which were sorted into three groups based on their expression level (red line, highest expressed; green line, lowest expressed). (C) UCSC genome browser view of H3K4me3/2/1 marks at the self-renewal gene Tbx3 during differentiation. shKdm5b ES cells have altered H3K4me3/2/1 levels relative to shLuc ES cells. (D) Average profile of H3K4me3/2/1 densities at p300 enhancer regions following 4 days of shLuc and shKdm5b ES cell differentiation. RPBM, reads per base pair per million reads. (E) Promoter density of H3K4me3 in ES cells and day 3 differentiated ES cells. (F) UCSC browser view of H3K4me3 density at HoxA cluster genes during differentiation. Note the delayed decrease of H3K4me3 levels during shKdm5b ES cell differentiation relative to shLuc ES cells.