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. 2017 Jul 18;6:e27406. doi: 10.7554/eLife.27406

Figure 2. H3-G34R mutants show a distinct transcriptional profile to set2Δ cells.

Schematic of genes that are (a) differentially-regulated, or (b) similarly down-regulated or up-regulated comparing either H3-G34R or set2Δ cells with H3-WT cells in RNA-seq analyses. Triplicate biological replicates were analyzed with cut-offs of 1.5 fold differences in expression, and false discovery rates of 5%. (c) RNA-seq profiles for chromosomes I, II, and III comparing Log fold change ratios for H3-G34R/H3-WT or set2Δ/H3 WT plotted against chromosome coordinates. (d) Zoomed-in regions of RHS Chr I (5.3 Mb-end, top) and LHS Chr II (first 300 Kb, bottom) as depicted in the boxed regions in (c) showing log FC data for individual biological replicates. (e) RT-PCR validation of fah1+ and grt1+ expression relative to adh1+ expression. (f) Density plots of RNA seq reads for sense (black) and antisense transcripts (blue) against log fold change ratios for set2Δ/H3 WT (left) and H3-G34R/H3 WT (right). Please see Figure 2—figure supplement 1 for analysis of heterochromatic loci, and Supplementary file 3 for full RNA-seq analysis results.

DOI: http://dx.doi.org/10.7554/eLife.27406.006

Figure 2.

Figure 2—figure supplement 1. Heterochromatin integrity is maintained in H3-G34R cells.

Figure 2—figure supplement 1.

(a) Real-time (RT) PCR analysis of pericentromeric imr, dg, and dh expression as well as the subtelomeric tlh expression in the indicated strains. Transcripts were compared relative to adh1+ expression. These domains are silenced in WT cells, so clr4Δ cells that lack heterochromatin represent a positive control. Error bars represent SEM from 2 independent experiments. (b) Chromatin immunoprecipitation (ChIP) of H3K9me2 and (c) Swi6 in the indicated cells at either the dh or tlh loci relative to adh1+. Error bars represent SEM from 5 biological replicates. (d) ChIP analysis of the Cnp1 protein to central core sequence cnt2, with ChIP enrichment depicted relative to input material. Data represents 6 biological replicates with error bars representing standard deviation.
DOI: http://dx.doi.org/10.7554/eLife.27406.007