Fig. 7: Eaf3 recognized H3K36 and H3K27 methylation and is required for the enrichment of H3K27me3 occupancy.

a Pulldown assays using MBP-Eaf3 and biotinylated histone peptides carrying the indicated methylation. Two independent experiments had similar results. b Pull-down assays using truncated Eaf3 and biotinylated histone peptides carrying the indicated methylation. c MST measuring the affinity of binding between MBP-Eaf3 and peptides with H3K36, H3K27, H3K9 and H3K4 methylation. Kd, dissociation constant. NBD, no binding detected. Data points are represented as means ± SD from three independent replicates. d A Venn diagram showing significant overlap among gene sets of Ash1-H3K36me2, H3K27me3, and Eaf3. The significance of overlap was determined using two-sided Fisher’s exact test. e Relative abundance of H3K27me3 in the indicated strains. Relative abundance of H3K27me3 to H3 was calculated by ImageJ software and that of WT was set as 1. Values are means ± SD from five biological replicates. Statistical significance was assessed by a two-tailed Student’s test (**P < 0.01). The exact P-values and western blots are shown in the Source Data. f A Venn diagram showing the significant overlap between gene sets of H3K27me3 (WT/∆kmt6) and H3K27me3 (∆eaf3/∆kmt6)). P-values were calculated by two-sided Fisher’s exact test. g, h Heatmaps and metagene plots showing H3K27me3 ChIP-seq signals of the H3K27me3-marked regions in the WT and ∆eaf3 strains. i Box plots and violin plots showing average transcriptions levels of the genes with and not H3K27me3 in the indicated strains. For box-plot, the horizontal lines from top to bottom represent the maximum, first quartile, median, third quartile, and minimum of the total data respectively. Statistical significance was assessed by a two-tailed Student’s test (**P < 0.01). The exact P-values are shown in the Source Data. For g and h, metaplots and heatmaps are shown with one biological replicate of ChIP-seq or RNA-seq assays, and similar results were gained in other replicates.