Figures and data
NRPE1 accumulates in heterochromatin in an h1 mutant background:
A) Genome browser image showing increased NRPE1 enrichment in h1 over heterochromatic (long, H3K9me2 enriched) and not euchromatic (short, not H3K9me2 enriched) TEs. B) metaplot showing ChIP-seq enrichment of NRPE1 in WT vs. h1, and H1 occupancy in WT for reference (data from GSE122394), at short vs. long TEs. C) boxplot showing association between NRPE1 and TE length in WT and h1. wilcoxon rank sum test p-values indicated. D) As in B) for drm12 vs. cmt2 hypo CHH DMRs. E) Violin plot inlaid with boxplot showing enrichment of NRPE1 at TEs that overlap with H3K9me2 peaks, vs. TEs that do not, in WT vs. h1. Boxplot medians are shown in blue. Wilcoxon rank sum test p-values indicated. F) as in B) at euchromatic vs. heterochromatic TEs. G) Chromosomal plots showing NRPE1 enrichment in h1 as compared to WT, with pericentromeric regions denoted in grey.
RdDM does not reciprocally affect H1 localization:
A) Scatter plot showing H1 enrichment in WT vs. nrpe1 at euchromatic vs heterochromatic TEs. B) Metaplot showing H1 enrichment at NRPE1 peaks in WT vs. nrpe1 C) H1 and NRPE1 occupancy at ZF-DMS3 off target peaks. Upper panel: H1 occupancy in WT vs. ZF108-DMS3. Lower panel: NRPE1 occupancy in WT vs. ZF108-DMS3. Chromatin from the same sample was used for both H1 and NRPE1 ChIP-seq data, so the lower panel serves as a control showing that ZF108-DMS3 is recruited to these off-target regions in these samples, despite H1 localisation remaining unchanged. D) Same as C) at the subset of regions where CHH methylation is gained in ZF108-DMS3 (Liu et al 2018), indicating that the fully functioning RdDM pathway is recruited to these regions.
Major CG and CHG methylation gains in h1 are independent of RdDM:
A) Chromosomal view of DNA methylation levels (average of 10kb windows) in the genotypes indicated on chromosome 1. Y-axis indicates fraction methylation (0-1). B) Methylation level over heterochromatic TEs. Upper panel: methylation metaplots, Lower panel - kernel density plots. In the kernel density plots, the average methylation is calculated for each TE in both mutant and WT, then then the methylation difference is calculated. The plot shows the frequency density of TEs that gain/lose DNA methylation in the regions in the mutants indicated. C) Methylation level over euchromatic TEs. Upper panel: metaplots, Lower panel - kernel density plots (as above). Arrow highlights the gain in CHG methylation in the h1/nrpe1 double mutant, as compared to nrpe1 alone.
Overlap analysis of h1 hyper CHG DMRs with 96 methylation mutants:
A) Number of hypo vs hyper DMRs in genotype comparisons indicated. B) h1 hyper CHG DMR frequency density plot over Chromosomes 1-5. C) Similarity matrix based on pairwise overlapping scores (Co-Occurrence Statistics) of DMRs from 96 whole methylomes.. The labels on the right summarise the major functional categories of the methylation mutant genotypes in the cluster block. (individual genotypes are shown in Supplementary Figure 6). The colour scale represents the number of observed DMR overlaps between the pairwise comparison indicated over the number of overlaps expected by chance (DMRs randomly distributed throughout the genome). Red means highly enriched overlapping DMRs (similar genomic distribution), blue means highly non-overlapping (different genomic distribution) D) Zoomed in view of the cluster containing h1 hyper CHG DMRs C) (see blue box and arrow).
Patterns of NRPE1 re-localisation in h1 show corresponding methylation changes: A) Methylation metaplot in the genotypes indicated over regions of the genome that gain NRPE1 in h1. Arrow highlights the change in average CHH methylation in the h1/nrpe1 double mutant as compared to the single mutants. B) Methylation metaplot of the genotypes indicated over regions of the genome that lose NRPE1 in h1.
SUVH1 encroaches heterochromatin in an h1 mutant background:
A) Genome browser image showing SUVH1 re-localisation in h1 mirrors that of NRPE1. B-D) Boxplot inlaid violin plots showing SUVH1 enrichment in WT vs h1 at short vs long TEs (B), at euchromatic vs heterochromatic TEs (C) and at drm12 vs cmt2 hypo CHH DMRs (D).
