Fig 1: N-Terminal domain accounting for the sequence preferences of DNMT3A. Sequence logos of the most methylated 10-mer non-CpG sequences (N = 5000) in (A) WT cells (WT), (B) DKO-zero cells, (C) DKO-zero cells expressing full-length DNMT3A (DNMT3A-FL), or (D) DKO-zero cells expressing the DNMT3A catalytic domain (DNMT3A-CD).
Fig 2: DNMT3A and -3B have different preferences for flanking sequences for CpG and non-CpG sites. Box plot of methylation levels in NCGN context after incubation with (A) DNMT3A or (B) DNMT3B for 240 min, ranked by median methylation level. Sequence logo of the 1000 most methylated 10-mer CG sequences after incubation with (C) DNMT3A or (D) DNMT3B for 30 min. Sequence logo of the 1000 most methylated 10-mer non-CpG sequences after incubation with (E) DNMT3A or (F) DNMT3B for 120 min.
Fig 3: DNMT3L stimulates DNMT3A and DNMT3B activity without altering their sequence preference. Average methylation by DNMT3A and DNMT3B in (A) CpG and (B) non-CpG sites. Sequence logo of the 1000 most methylated 10-mer CG sequences after incubation with (C) DNMT3A/3L or (D) DNMT3B/3L for 120 min. Sequence logo of the 1000 most methylated 10-mer non-CpG sequences after incubation with (E) DNMT3A/3L or (F) DNMT3B/3L for 120 min.
Fig 4: In vitro methylation assay using recombinant full-length human DNMT3A and -3B. (A) Average level of methylation introduced by DNMT3A, DNMT3B, or DNMT3L in CpG and non-CpG sites. Box plots of methylation levels at NCN sequences after incubation with (B) DNMT3A or (C) DNMT3B for 240 min, ranked by median methylation level. Sequences are written in 5′ to 3′ order.
Fig 5: Methylation signatures in the mouse and human genome depend on the presence of DNMT3A or DNMT3B. Sequence logos of the most methylated sequence contexts in chromosome 1 of mouse stem cells. (A) Top methylated 10-mer CG sequences: 100% methylation for WT (left; N = 325622), >60% methylation for TKO with Dnmt3a (middle; N = 628), and >40% for TKO with Dnmt3b (right; N = 399). (B) Top methylated 10-mer CH sequences: >40% methylation for WT (left; N = 16392), >30% for TKO with Dnmt3a (middle; N = 533), and >20% for TKO with Dnmt3b (right; N = 177). Sequence logos of the most methylated (>20% methylation) CH sequence contexts in chromosome 1 of hESCs. (C) WT hESCs (N = 163050). (D) DNMT3A-KO hESCs at early passage 7 (left; N = 28101) and late passage 22 (middle; N = 59323). (E) DNMT3B-KO hESCs at passage 6 (left; N = 24327) and passage 22 (middle; N = 20603). (F) DNMT3A/B double-knockout hESCs at passage 7 (left; N = 12736) and passage 22 (right; N = 6859). (G) Ratio between CAC and CAG methylation in mouse Dnmt-TKO cells with Dnmt3a/Dnmt3b reintroduced and hESCs with either DNMT3A or DNMT3B knocked out.
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