RNA-mediated regulation in pathogenic bacteria

Abstract

Pathogenic bacteria possess intricate regulatory networks that temporally control the production of virulence factors, and enable the bacteria to survive and proliferate after host infection. Regulatory RNAs are now recognized as important components of these networks, and their study may not only identify new approaches to combat infectious diseases but also reveal new general control mechanisms involved in bacterial gene expression. In this review, we illustrate the diversity of regulatory RNAs in bacterial pathogens, their mechanism of action, and how they can be integrated into the regulatory circuits that govern virulence-factor production.

Figure 1.

Prominent functions of Hfq in bacterial pathogens. (A) Summary of Hfq protein functions as inferred from the phenotypes of hfq mutants in Gram-negative bacteria (reviewed in Chao and Vogel 2010). In most bacteria examined, inactivation of the hfq gene causes a growth defect, a decrease in motility and biofilm formation, and an inability to cope with environmental stresses. Expression and/or secretion of the type III secretion system can be affected in either a negative or positive manner. (B) Gene ontology analysis of disregulated genes in a Salmonella hfq mutant strain (Sittka et al. 2008). Bars show the percentage of Hfq-dependent genes in each pathway. Orange bars denote the five major virulence regions of S. typhimurium; green bars denote metabolic functions.

Figure 2.

Dual functions of peptide-coding small RNAs. (A) Quorum sensing and RNAIII-controlled gene expression. S. aureus produces an autoinducing peptide that is sensed by a histidine kinase (AgrC). Sensing of the autoinducing peptide by AgrC leads to phosphorylation of the response regulator AgrA, which, in turn, is a transcriptional activator of the bifunctional RNAIII. RNAIII encodes the hld gene (coding for δ-hemolysin), and posttranscriptionally regulates several target mRNAs. Whereas spa, coa, rot, SA1000, and SA2353 mRNAs are repressed by RNAIII (Boisset et al. 2007), the hla mRNA is activated by this bifunctional RNA (Morfeldt et al. 1995). Red and blue arrows denote sRNA-mediated posttranscriptional activation and repression, respectively. (B) Glucose-phosphate stress and SgrS modulates virulence gene expression. The accumulation of glucose phosphate in the cytoplasm is sensed by the transcription factor SgrR, which, in turn, activates transcription of the SgrS RNA. The 5′ region of SgrS encodes a small peptide SgrT (43 amino acids), whereas the 3′ region contains a seed-pairing domain that targets several mRNAs with the help of the RNA chaperone Hfq. In addition to targeting the conserved sugar transporters ptsG and manXYZ, SgrS also represses the mRNA of the T3SS secreted virulence effector SopD in Salmonella (Papenfort et al. 2012). (C) PhrS sRNA in Pseudomonas. Binding of the PhrS sRNA induces the translation of the quorum-sensing regulator PqsR translational activation of an upstream ORF (uof), which is coupled to PqsR (Sonnleitner et al. 2011). Expression of PhrS is induced by ANR, an oxygen-responsive regulator, at high-cell density populations when oxygen becomes limiting.

Figure 3.

Several sRNAs regulate quorum sensing in Vibrio cholerae. At low cell density, the histidine kinase-receptor LuxPQ phosphorylates LuxU, which, in turn, phosphorylates LuxO (LuxO-P). LuxO-P, together with the RNAP-σ⁵⁴ holoenzyme, activates the transcription of 5 sRNAs, Qrr1-5 (Rutherford et al. 2011). These sRNAs together with Hfq repress the translation of hapR mRNA and activate the translation of ahpA mRNA through an anti-antisense mechanism. Qrr4 also represses the translation of luxO mRNA, thus creating a negative-feedback regulation. At high cell density and high concentrations of extracellular autoinducer-2, the LuxPQ receptor acts as a phosphatase and dephosphorylates LuxO. In consequence, the synthesis of Qrr sRNAs is arrested and the synthesis of HapR is derepressed, and a gradient of higher concentrations of HapR (in blue) over AphA (in pink) is established to provide a large range of responses. In addition, AphA and HapR autoregulate their own synthesis at the transcriptional level. Red and blue arrows denote sRNA-mediated posttranscriptional activation and repression, respectively.

Figure 4.

UTR-derived trans-acting small RNAs. (A) 5′ UTR-derived sRNA in L. monocytogenes. During the exponential phase of growth (i.e., under rich nutrient conditions), S-adenosylmethionine (SAM) binds to SreA, the terminator structure (in red) is formed, and transcription is prematurely halted (Loh et al. 2009). At 37°C, the prfA thermosensor (in green) melts to expose the SD for PrfA synthesis and consequently the virulence factors are produced. Under these conditions, the SreA-terminated transcript represses prfA mRNA translation by binding to the 5′ UTR 80 bp upstream of the SD and turning off its expression by an unknown mechanism. (B) 3′ UTR-derived sRNA in S. typhimurium. The synthesis of DapZ (in red) is induced by HilD, the master regulator of Salmonella invasion genes. DapZ carries a short unpaired G/U-rich region that initiates binding to the conserved C/A-rich regions in dppA and oppA mRNAs, which encode the major ABC transporters (Chao et al. 2012). The same target sites are recognized by a similar G/U-rich seed region of GcvB sRNA, the master sRNA regulator of amino acid transporters.

Figure 5.

Novel regulations by long and short antisense RNAs. (A) AmgR is a 1.2-kb cis-encoded RNA that is expressed convergently to the mgtC ORF in Salmonella (Lee and Groisman 2010). When Mg²⁺ concentrations are low, the PhoPQ two-component system activates expression of both the AmgR and mgtC transcripts. The interaction of these two RNAs results in RNase-E-dependent degradation of the RNA duplex. (B) Unusually long antisense RNAs have been predicted upstream of many operons in Listeria (Wurtzel et al. 2012). Expression of these lasRNAs may activate the downstream operons while inhibiting the expression of genes located antisense to them, which usually have opposite functions to the operon. (C) SprA1_AS is a short, ∼60-nucleotide-long antisense RNA of a toxin–antitoxin (TA) system in S. aureus. It overlaps with the 3′ end of the dual-functional SprA1 sRNA. SprA1_AS was shown to use its 5′ end to base-pair with SprA1 sRNA through imperfect pairing, and to repress the translation of the SprA1-encoded small cytolytic peptide (Sayed et al. 2011).

Figure 6.

Small RNAs directly regulate protein activity in Pseudomonas. (A) RsmA is an RNA-binding protein that modulates gene expression by antagonizing translational initiation (Lapouge et al. 2008). The RsmY/Z RNAs contain multiple RsmA binding sites and counteract RsmA activity via a titration mechanism in several different plant and human pathogens. Similar small RNAs (e.g., CsrB) target the homologous CsrA protein in enterobacteria (Romeo et al. 2012). (B) Crc is an RNA-binding protein that modulates expression by antagonizing translational initiation (Moreno et al. 2009). CrcY/Z RNAs carry up to six Crc-binding sites and counteract Crc activity via a titration mechanism.