Fig. 2: MBD5/6 and MBD7 prefer different subsets of CG-methylated loci.

a (Left) Average methylation density (sum of % methylation across all cytosines in indicated sequence context) in 400 bp windows centered on the midpoint of the peaks shown at right. Each column was normalized separately. (Right) Heatmaps of MBD6-Flag signal and MBD7-Flag signal (log₂ IP over no-FLAG control) over the union of all MBD6 and MBD7 peaks. Each heatmap is an independent transgenic line (T2-[line number]). All heatmaps in (a) share same row order. b Browser tracks showing the location of MBD6/7 union peaks (top), as well as MBD6 and MBD7 ChIP-seq signal (log₂ IP over no-FLAG control), % DNA methylation data from inflorescences with unopened flower buds²⁵, gene annotations (black = forward strand, yellow = reverse), and transposon (TE) annotations. Genes and TEs are from the araport11 annotation⁶¹. Both Col replicates from ref. ²⁵ were pooled; only sites with 5 or more coverage shown; small negative value (−10%) corresponds to sites with coverage but no methylation, to distinguish from missing data. H3K9me2⁶³ and PolV (NRPE1)⁵⁶ ChIP-seq signal also shown. c (left) Representative image of root nucleus expressing pMBD7::MBD7-YFP and incubated with DAPI to stain chromocenters. (right) Percent of foci assayed (n = 120 for each condition) where YFP and RFP signal overlapped vs. did not overlap. Left bar shows co-expression of MBD6-RFP and MBD7-YFP, right bar shows control co-expressing MBD6-RFP with MBD6-YFP. d Percent of MBD7-dominant, mixed, and MBD6-dominant peaks from (A) overlapping annotated transcripts classified as either gene body methylated (gbM, mCG only) or TE-like methylated (teM, mC in all contexts). Annotations were from ref. ⁶. e Metaplots of DNA methylation²⁵, NRPE1⁵⁶, histone H1 and H3K9me2⁶³, histone H3 and H3K4me3⁶⁴, accessibility by ATAC-seq⁶⁵, and RNA-seq⁴, over the peaks identified in (a). f Loess-smoothed distribution of MBD7-dominant, MBD6-dominant, and mixed peaks across Chromosome 4. Pericentromere shown as a grey bar. g Metaplots of MBD6 and MBD7 ChIP-seq log₂(IP/no-FLAG control) signal over pollen mbd5/6 upregulated transcripts rescued, partially rescued, and not rescued by MBD7 loss (see Fig. 1d). Controls are an equal number of random genes matched for expression level and length. Transgenic lines were averaged (N = 2 for MBD6-Flag, N = 3 for MBD7-Flag).