Fig. 6. Genome-wide effect of mta1 downregulation.

a Schematic diagram of the dynamic mta1 regulation experiment. From a starting point with 0 mg/L ZnSO₄ and active Mta1 expression, it was downregulated by increasing ZnSO₄ to 10 mg/L and 20 mg/L. Colonies growing in 20 mg/L ZnSO₄-supplemented media were transferred again to media without ZnSO₄. b Genomic 6mA levels measured by SMRT sequencing and HPLC-MS/MS. Data are represented as mean ± SD. Different letters indicate statistically significant differences, while the identical letters denote no significant differences, calculated using one-way ANOVA ( P < 0.001). (HPLC/MS n = 3 biological replicates for each growth condition analyzed) c 6mA frequency profile over protein-coding genes for the UM67 strain growing with 0 mg/L (blue), 10 mg/L (green), and 20 mg/L (yellow) ZnSO₄. d Proportion of symmetric 6mA sites and motif logo for UM67 growing with 0 mg/L ZnSO₄ (left) and 20 mg/L ZnSO₄ (right). e Upregulated genes (red) and downregulated genes (blue) that have lost a MAC when growing in media supplemented with 20 mg/L (top panel) and with 10 mg/L (bottom panel). Two-tailed P values were obtained from DESeq2 Wald’s test. Multiple comparison adjustments were performed with the Benjamini-Hochberg false discovery rate (FDR) correction procedure. f 6mA frequency profiles over upregulated and downregulated genes (20 mg/L vs 0 mg/L). The blue line indicates the 6mA frequency for UM67—0 mg/L, and the yellow line indicates the 6mA frequency for UM67—20 mg/L. g H3K4me3 enrichment (IP/Input ratio) over MAC lost when UM67 is grown with 20 mg/L ZnSO₄ (right panel) compared to 0 mg/L (left panel). Each MAC was extended to 2000 bp and fragmented into 200 equally sized bins. h Profile and heatmap of 6mA and H3K4me3 enrichment (IP/Input ratio) for each cluster of genes (upregulated, upregulated with MAC lost; downregulated, downregulated with MAC lost). Red and blue lines in the profiles indicated upregulated and downregulated genes in the 20 mg/L vs 0 mg/L, respectively. For both 6mA and H3K4me3 panels, the panel on the left is for UM67—0 mg/L, and the panel on the right is for UM67—20 mg/L. i Normalized occupancy of nucleosome dyads in DEGs (20 mg/L vs 0 mg/L) was plotted in reference to TSS, including 1.5 kb flanking regions. A heatmap with the average normalized occupancy signal is also displayed below. Kolmogorov-Smirnov test (KS) was performed to evaluate differences between frequency distributions (P < 0.001). j Nucleosome classification according to their fuzziness score for upregulated (top panel) and downregulated (bottom panel) genes. Nucleosomes bound to genes with MAC lost (20 mg/L vs 0 mg/L) are indicated in light pink. Kolmogorov-Smirnov test (KS) was performed to evaluate differences between frequency distributions (P = 0.0053 for upregulated and P < 0.0001 for downregulated genes). Source data are provided as a Source Data file.