Supplemental Material for Klocko et al., 2018

Figure S1 shows aberrant cytosine methylation in dim-1 strains.
Figure S2 shows the complementation of a dim-1 strain with an ectopic copy of gene NCU06484.
Figure S3 shows the phenotypic characterization of a dim-1 strain, as this strain has changes in cytosine methylation, including the loss of cytosine methylation in constitutive heterochromatic regions and a gain in cytosine methylation in intergenic regions.
Figure S4 shows the cytosine (detected by bisulfite-sequencing) and histone methylation (as H3K9me3; detected by ChIP-sequencing) across all seven chromosomes of Neurospora crassa.
Figure S5 shows how the new cytosine methylation in a dim-1 strain requires the DIM-2 DNA methyltransferase.
Figure S6 shows how an ectopic copy of the dim-2 gene does not rescue the loss of cytosine methylation phenotype in constitutive heterochromatic regions.
Figure S7 shows how the new cytosine methylation peaks in dim-1 intergenic regions does not require the action of small RNAs produced by the ERI-1 helicase.
Figure S8 shows average enrichment profiles of H3K9me3 ChIP-sequencing replicate data across traditional constitutive heterochromatic regions, all regions gaining cytosine methylation in a dim-1 strain, or just the neo-heterochromatic regions that only gain cytosine methylation in a dim-1 strain.
Figure S9 shows how neo-heterochromatin requires the histone methyltransferase DIM-5
Figure S10 shows the affect of the dim-1 strain on heterochromatic silencing.
Figure S11 shows the minimal changes to facultative heterochromatin, marked by H3K27me2/3, in a dim-1 strain.
Figure S12 shows the growth rate of a dim-1 strain.
Figure S13 shows a possible correlation that genes that gain of cytosine methylation in a dim-1 strain are some of the most highly expressed in Neurospora.
Figure S14 shows the replicate analysis, using average enrichment profiles that use different reference points, of micrococcal nuclease sequencing experiments.
Figure S15 shows that an ectopic copy of the histone H3 gene does not rescue the cytosine methylation phenotype in a dim-1 strain.
Figure S16 shows the changes in nucleosome positioning in genes that are up-regulated, down-regulated, and unchanged in a dim-1 strain.
Figure S17 shows a control experiment for average enrichment profiles of nucleosome data: if random reference points are used, no periodicity is observed.
Figure S18 shows the proteins that interact with DIM-1, as determined by mass spectrometry.
Figure S19 show the proteins that interact with NCU00856, which is an interacting partner with DIM--1, and how the loss of NCU00856 does not cause a cytosine methylation phenotype.
Figure S20 shows the characterization of DIM-1 localization through DamID, including several genomic regions that were examined for DIM-1-DAM localization and how the removal of conserved domains from DIM-1 affected protein levels and localization.
File S1 shows an alignment of the DIM-1 primary structure with other orthologous proteins, and the associated E-values of those alignments.
File S2 provides the genes that are up- and down-regulated in dim-1 strains, the peak call files of H3K9me3, H3K27me2/3, and cytosine methylation, and the list of 1001 unchanged genes used as a control.
File S3 provides the python script to re-order the data within heatmaps so each heatmap is directly comparable.
Table S1 provides a list of strains used in this study.
Table S2 provides a list of oligos used in this study.