Dnmt3b induces DNA methylation changes detected by RRBS. (A-B) Smoothened scatter plots of methylation values for Dnmt3b-knock-in vs wild-type control samples in Myc-Bcl2 (A) and MLL-AF9 (B) leukemias, respectively. Colors represent the density of points ranging from red (high density) to blue (low density). (C) Chromosomal distribution of DMCs in Myc-Bcl2 (left) and MLL-AF9 (right) leukemias. Every dot represents 1 DMC. Red indicates DNA hypermethylated DMCs in Dnmt3b-knock-in samples, green indicates hypomethylated DMCs. (D) Region presence of covered and DMC sites across different genomic regions. Promoter regions were defined as 1000 bp upstream of transcriptional start site (TSS) to 500 bp downstream of TSS; gene bodies were defined as 500 bp downstream of TSS to end of gene. GB, gene bodies; P, promoter. (E-F) Presence of transcription factor binding sites (E), and RNA polymerase II binding sites (F) among DMRs. The number of centers of a particular region of interest that could be expected in DMRs under the assumption of a uniform distribution in RRBS-covered regions (expected region centers) was subtracted from the number of region centers actually found in DMRs (observed region centers). The bars visualize the differences of observed and expected region centers in hypermethylated DMRs. ***P < .001. (G) Smoothed scatter plot shows CpG sites that gained DNA methylation in previously published Dnmt triple-knockout mice (TKO) with reintroduced Dnmt3b50 and the corresponding DNA methylation difference in MLL-AF9 leukemias (Dnmt3b-knock-in − wild-type). Methylation in TKO + Dnmt3b indicates CpGs that gain DNA methylation after eradication of existing DNA methylation patterns in TKO cells following reintroduction of Dnmt3b. (H) Number of CpG sites that show methylation gain in TKO murine embryonic stem cells >0, >20, or >40, and hypo- or hypermethylation in Dnmt3b-knock-in MLL-AF9 leukemias. (I) Smoothed scatter plot showing all CpGs that are differentially methylated in Myc-Bcl2 and MLL-AF9 leukemias, respectively. The majority of DMCs showed hypermethylation in both leukemias.
