Small RNAs in Bacterial Virulence and Communication

Genomic location and regulatory relationships between bacterial riboregulators and their mRNA targets. Riboregulators are depicted in red; target mRNAs are shown in blue. Flanking open reading frames (ORFs) are shown in black. Arrows indicate transcriptional start sites. (A) Cis-encoded antisense RNAs are transcribed from the opposite strand to their target mRNAs and can overlap with target 5′/3′ untranslated regions (UTRs) (top panel) and/or the mRNA ORF (bottom panel). (B) Trans-encoded sRNAs can be expressed from distinct regions of the chromosome from their target genes: either from stand-alone genes encoded intergenically (top panel) or from ORFs/3′UTRs via either processing or internal transcriptional start sites (bottom panel). (C) Extended UTR elements of adjacent operons can allow for coregulation of related genes at the posttranscriptional level. The long-antisense RNA (lasRNA) of the excludon paradigm arises from transcription of an extended 5′UTR that has complementarity to a divergently transcribed operon (top panel). Also, extended 3′UTR elements can potentially base-pair with transcripts expressed from convergently transcribed operons (bottom panel). (D) Cis-elements within mRNAs themselves can regulate expression of their associated transcripts. These include ligand-binding riboswitches (top panel) and temperature-responsive RNA thermosensors (bottom panel).

Mechanisms of posttranscriptional control by regulatory RNAs. (A) Gene repression (left) and activation (right) mechanisms used by base-pairing sRNAs (depicted in red) for direct regulation of target mRNAs (shown in blue) at the level of translation or stability. Base-pairing interaction sites in mRNAs and sRNAs are shown with blue- and red-lined boxes, respectively. Potential RNase cleavage sites are indicated with an orange asterisk. Also participating are ribosomes and RNases. TIR, translation initiation region including RBS and start codon. (B) Potential sRNA interaction sites in regulated target mRNAs, starting from the TSS (transcriptional start site) to the transcriptional terminator (TERM). (C) Targeting/titration of other regulatory molecules by riboregulators acting as so-called sponges to affect gene expression. RNA sponges can be stand-alone sRNAs, regions of mRNAs themselves (either intact or processed), or those derived from housekeeping RNAs such as the 3′ external transcribed spacer (3′ ETS) of tRNAs. They can target either sRNA or protein regulators and have been shown to sequester them away from their targets, trigger their degradation, and/or modulate their regulatory activity.

Numerous riboregulators participate in quorum sensing and virulence regulation of Vibrio cholerae. (A) Riboregulation of the V. cholerae ToxT virulence regulon in response to temperature. The central transcriptional regulator ToxT (blue circles, center) activates virulence and colonization factor genes, such as tcp (toxin-coregulated pilus), ctxAB (cholera toxin), and acf (accessory colonization factor). ToxT also autoregulates its own transcription. Levels of ToxT are also modulated in response to temperature by a FourU RNA thermometer, with increased translation at the 37°C host temperature. ToxT also activates the sRNAs TarB, which represses translation of the tcpF ORF of tcpA-F mRNA, and TarA, which represses ptsG mRNA (glucose uptake). The VrrA sRNA also represses tcpA.

Numerous riboregulators participate in quorum sensing and virulence regulation of Vibrio cholerae. (B) The Qrr sRNAs mediate the switch between Vibrio low and high cell-density physiologies via reciprocal posttranscriptional regulation of the master regulators AphA and HapR. Vibrio autoinducers (AI-2 and CAI-1) are made by LuxS and CqsA, respectively, and accumulate extracellularly. Phosphorelay systems headed by LuxPQ or CqsS (AI-2 and CAI-1, respectively) detect autoinducers. Left panel: Low bacterial density. Continued phosphorylation of LuxO at low autoinducer conditions leads to transcription of the Qrr sRNAs, which act along with the RNA chaperone Hfq to activate translation of aphA mRNA. In turn, AphA expression induces the ToxT virulence regulon (see panel A), as well as genes required for biofilm formation (vpsT). The Qrrs also repress the hapR mRNA, which encodes the high-density master regulator (see right panel). Right panel: High bacterial density. High autoinducer concentration reduces levels of phosphorylated LuxO and, thus, Qrr expression. The hapR mRNA is no longer destabilized, allowing translation of the HapR regulator. HapR activates genes that mediate biofilm dispersal and competence. In addition, genes activated by AphA at low density, such as vpsT, as well as aphA itself and its regulated genes, are repressed by HapR. Genes encoding the type VI secretion system are also induced. Finally, feedback regulation occurs via HapR activation of Qrr expression and Qrr repression of the luxO mRNA. The sRNA VqmR is activated by the transcriptional regulator VqmA and posttranscriptionally represses vpsT mRNA and rtx mRNA, encoding the RTX toxin, as well as six other mRNAs.

The dual-function sRNA RNAIII of Staphylococcus aureus reciprocally regulates expression of secreted virulence factors and surface proteins in response to cell density. (A) Genomic context and transcriptional regulation of the S. aureus agr quorum sensing locus, including the dual-function RNAIII. The RNAII mRNA (black) encodes proteins required for synthesis and detection of the peptide pheromone (agrBDCA, green and red open reading frames [ORFs]). Under high cell density and high autoinducer concentration, phosphorylated AgrA (red) activates transcription of the RNAIII sRNA (blue). RNAIII encodes δ-haemolysin (hld ORF) and is the major mediator of Agr regulation. (B) Overall integration of RNAIII posttranscriptional activities promotes toxin expression and represses expression of secreted proteins. Center: General structure of RNAIII with the hld coding region (light blue) and C-rich loops (red). The RNAIII molecule directly activates the mRNA encoding α-haemolysin (hla). Also, together with the double-strand-specific RNase III, the sRNA directly represses numerous genes encoding surface-associated proteins (coa, spa, SA2353, SA1000). RNAIII also represses translation of Rot, a repressor of toxin gene expression.