Figure 1.

RNase Y allows differential expression between genes co-expressed in the saePQRS operon. (A) Schematic representation of the saePQRS operon, with its primary and mature RNA molecules (T1–T4), promoters (P1 and P3), terminator (Term), cleavage site (CS) and putative stem loops. (B) Schematic representation of sae-gfp constructs carrying different deletions (the deleted sequence is indicated in the panel with a cross). The RNAs observed in the northern blot analyses are indicated with their names and lengths below each constructs. (C) Northern blot analyses to examine sae processing in strains carrying the sae-gfp constructs. Newman saeP mutant and saeP rny double mutant strains carrying different constructs were grown to exponential phase. RNA was then harvested and hybridized with DIG-labeled DNA probes specific for gfp, saeP and rny. As a loading control, 16S rRNA detected in the ethidium bromide-stained gel was used, which is shown at the bottom of the panel. For clarity, lane numbers are indicated in the panel. (D) RT-qPCR to assess the ratio between saeR and saeP copy numbers. Newman wild-type, rny mutant and complemented strains were grown (in triplicate) to late exponential phase and RNA was extracted. After DNase I treatment, one-step RT-qPCR was performed. saeR and saeP copy numbers were calculated by reference to a standard curve. Statistically significant differences between the samples are indicated: **P = 0.001 to 0.01; ***P < 0.001.