σ^E‐dependent small RNAs of Salmonella respond to membrane stress by accelerating global omp mRNA decay

Abstract

Summary The bacterial envelope stress response (ESR) is triggered by the accumulation of misfolded outer membrane proteins (OMPs) upon envelope damage or excessive OMP synthesis, and is mediated by the alternative sigma factor, σE. Activation of the σE pathway causes a rapid downregulation of major omp mRNAs, which prevents further build‐up of unassembled OMPs and liberates the translocation and folding apparatus under conditions that require envelope remodelling. The factors that facilitate the rapid removal of the unusually stable omp mRNAs in the ESR were previously unknown. We report that in Salmonella the ESR relies upon two highly conserved, σE‐controlled small non‐coding RNAs, RybB and MicA. By using a transcriptomic approach and kinetic analyses of target mRNA decay in vivo, RybB was identified as the factor that selectively accelerates the decay of multiple major omp mRNAs upon induction of the ESR, while MicA is proposed to facilitate rapid decay of the single ompA mRNA. In unstressed bacterial cells, the two σE‐dependent small RNAs function within a surveillance loop to maintain envelope homeostasis and to achieve autoregulation of σE.

Dual RNA-seq of pathogen and host

“…2.24a,b). To confirm Dual RNA-seq-based gene expression profiling of intracellular Salmonella by unrelated methods, four of the induced sRNAs were selected for independent qRT-PCR experiments: The wellcharacterized Salmonella sRNAs RyhB (Masse and Gottesman, 2002) and RybB (Balbontin et al, 2010;Papenfort et al, 2006) were chosen as representative for a highly (>100 fold) or slightly (~2 fold) induced sRNA. In addition, STnc440 -a previously validated (Sittka et al, 2008) but poorly characterized sRNA -was included as was a computationally predicted candidate sRNA, STnc510.…”

“…Thus, in order to identify direct targets of this sRNA, a previously described pulse-expression approach was applied (Masse et al, 2005;Papenfort et al, 2006). Briefly, the sRNA of interest is transiently over-expressed for a short time period (5-10 min) from an arabinose-inducible promoter and the global abundance of mRNAs is monitored in the presence or absence of the sRNA.…”

“…MicA and MicL (Guo et al, 2014;Hobbs et al, 2010;Papenfort et al, 2006) were induced upon THP-1 invasion. At the same time numerous omp mRNAs, which are major targets of σ Eactivated sRNAs (Rhodius et al, 2006), were rapidly down-regulated.…”

A small RNA regulates multiple ABC transporter mRNAs by targeting C/A-rich elements inside and upstream of ribosome-binding sites

“…Tjaden et al (2006) suggested that functional relationship of the proteins encoded by target candidates, as it is also seen among the GcvB targets, may add confidence to predictions. Similarly, the OmrA/B and RybB sRNAs were predicted to directly target multiple mRNAs that collectively encode for outer membrane proteins Johansen et al 2006;Papenfort et al 2006). By the same token, the iron stress-responding E. coli RyhB sRNA was shown to regulate multiple mRNAs that encode proteins involved in iron metabolism (Massé and Gottesman 2002), and many of the mRNAs demonstrated to be direct targets of S. aureus RNAIII encode bacterial virulence factors (Boisset et al 2007).…”

“…Recent biocomputational and experimental approaches predicted more sRNAs to target multiple mRNAs. For example, pulse expression of several E. coli and Salmonella sRNAs (Massé et al 2005;Guillier and Gottesman 2006;Johansen et al 2006;Papenfort et al 2006;Tjaden et al 2006) followed by global transcriptome profiling showed the rapid depletion of many mRNAs. The kinetics of target decay observed in these experiments strongly argues that the regulated mRNAs are direct targets.…”