Figure 2: YonO is a bona fide RNAP of SPβ.
From: Single-peptide DNA-dependent RNA polymerase homologous to multi-subunit RNA polymerase
(a) Western blot analysis of YonO expression upon SPβ induction with mitomycin C. DivIVA, a constitutively expressed cell division protein, was probed as a loading control. (b) Plaque formation assay with SPβ sensitive strain (CU1065) infected with lysates of mitomycin C (MitC)-induced wild type, ΔSPβ and ΔyonO B. subtilis strains. Parts of agar plates are shown. (c) A volcano plot of the gene expression fold change against P-value for the 4314 genes between WT strains with and without mitomycin C treatment. Genes were considered significant at a fold change of 2 and a P-value threshold of 0.05, following correction using the Benjamini–Hochberg false discovery rate. SPβ genes with statistically significant changes in expression are shown in green (N=157) with other significant genes in pink (N=1,072). Blue indicates all the genes (out of 4,314) without a statistically significant change in expression. (d) A volcano plot of the gene expression fold change against P-value for the 4,314 genes between wild-type and ΔyonO strains upon prophage induction. Criteria for significant change are as in panel c. Out of the 148 SPβ prophage genes induced by mitomycin C (see panel c), 37 were not expressed in the ΔyonO strain (pink dots with labels; see also Supplementary Fig. 2). Blue dots indicate all the genes (out of 4,314) without a statistically significant change in expression. (e) Confirmation of the transcription start site (TSS) of the promoter used by YonO (PYONO) to transcribe the operon of SPβ late genes, as determined by RNA-seq, using primer extension. (f) Deletion of YonO does not affect transcription from constitutive PVEG promoter of B. subtilis as confirmed by primer extension.
