Fig. 6: Disease-associated PWWP-ADD interface mutations led to impaired DNMT3A autoinhibition and target selectivity.

a Mapping of the W330R, D333N and E545G to the PWWP-ADD interface, with the PWWP and ADD domains shown in ribbon representation and the MTase domain shown in surface representation. b ¹H,¹⁵N-HSQC spectral overlay of E545G-mutated DNMT3A ADD, free or in the presence of PWWP^WT. Four selected regions were shown in expanded views. c ¹H,¹⁵N-HSQC spectral overlay of the DNMT3A ADD domain, free or in the presence of PWWP^WT or mutant DNMT3A PWWP (PWWP^W330R and PWWP^D333N). Four selected regions of the NMR spectra for the W330R (left) or D333N (right) mutants were shown in expanded views. d In vitro DNA methylation assay of the DNMT3A2-DNMT3L complex, WT or disease mutant, on 36-mer (GAC)₁₂/(GTC)₁₂ DNA. Two-tailed Student’s t test was used to compare the activity of WT vs mutant. Data are mean ± s.d. (n = 3 biological repeats). e In vitro DNA methylation assay of DNMT3A2-DNMT3L, WT or disease mutant, on the nucleosome substrates with various histone modifications. The statistical analysis used two-tailed Student’s t test. Data are mean ± s.d. (n = 3 biological repeats). f Box plots showing the DNA methylation levels of 10-kb bins for non-CGI overlapping H3K36me2 positive (left) or negative (right) regions in TKO mESCs reconstituted with WT or W330R DNMT3A1 (n = 2 biological replicates). Box-and whisker plots depict 25–75% in the box, whiskers are 1.5 times the interquartile range (IQR), and median is indicated. Wilcoxon rank-sum two-sided test is used for statistical analysis. g The DNA methylation level of 10-kb bins that overlapped with quintile Q1–Q5 of non-CGI overlapping H3K36me2 in WT or W330R DNMT3A1 TKO mESCs (n = 2 biological replicates). Box-and-whisker plots depict 25–75% in the box, whiskers are 1.5 times the interquartile range (IQR), and median is indicated. Source data are provided as a Source Data file.