The Staphylococcus aureus RNome and its commitment to virulence

Abstract

Staphylococcus aureus is a major human pathogen causing a wide spectrum of nosocomial and community-associated infections with high morbidity and mortality. S. aureus generates a large number of virulence factors whose timing and expression levels are precisely tuned by regulatory proteins and RNAs. The aptitude of bacteria to use RNAs to rapidly modify gene expression, including virulence factors in response to stress or environmental changes, and to survive in a host is an evolving concept. Here, we focus on the recently inventoried S. aureus regulatory RNAs, with emphasis on those with identified functions, two of which are directly involved in pathogenicity.

Figure 1. General mechanisms given for several S. aureus regulatory RNAs.

(A) S-adenosyl methionine (SAM) riboswitch regulates several operons encoding enzymes and transporter proteins. SAM binds to an aptamer domain and stabilizes the formation of a terminator hairpin (the alternate pairings are in red) to arrest transcription . (B) Schematic representation of a T-Box involved in the regulation of aminoacyl-tRNA synthetases (aaRS). Non aminoacylated tRNA binds to the leader region at two different sites: the anticodon sequence of the tRNA base paired with a codon-like triplet present in the “specifier loop”, and the ACCA end of the tRNA binds to a complementary sequence located in the T-Box motif . This interaction stabilizes an anti-terminator structure allowing transcription of the downstream genes. *The aminoacyl-tRNA synthetases regulated by this mechanism are the ValRS, MetRS, IleRS, PheRS, GlyRS, SerRS, HisRS, and the AspRS. (C) The SprD pathogenicity island RNA (in red) binds to the ribosome binding site (The SD is green) of sbi mRNA to repress translation initiation . (D) The RsaOX (in red) (or Teg14as) cis-acting asRNA , is complementary to the coding sequence of tnp mRNA and is predicted to induce rapid degradation of tnp mRNA. Both the asRNA and the mRNA target site are highly folded, suggesting that the pairing is initiated by a loop–loop interaction.

Figure 2. Secondary structures of selected S. aureus sRNAs.

The secondary structures of RsaA and RsaE and of SprD were experimentally determined. The secondary structure of the antisense RNA RsaOX (or Teg14as) is proposed based on computer predictions , . The genomic locations and flanking genes of the sRNAs are indicated. The known regulatory domains of the sRNA, which bind to mRNA targets, are in red. The black circled nucleotides are accessible C-rich motifs that are proposed to be crucial for the initial binding with mRNA .

Figure 3. The functional RNAIII structure and its mRNA targets.

(A) Schematic view of the RNAIII-mediated antisense activation mechanism. Hairpin loops H2 and H3 of RNAIII (red) bind to the hla mRNA (black) to activate translation initiation. The ribosomal 30S subunit is schematized. The SD sequence is green. (B) RNAIII secondary structure (adapted from [52]) and its genomic location within the agr locus (bottom). RNAIII encodes the delta-hemolysin (hld, in green). The 5′UTR activates alpha-hemolysin translation and the 3′ domain represses the translation of virulence factors and of the transcriptional repressor of toxins (rot) , . The conserved C-rich sequences detected in H7, H13, and H14 is indicated. (C) Schematic views of the RNAIII-mediated antisense translation initiation repression mechanisms. RNAIII structural domains H7, H13, and H14 (in red) are involved in interactions with target mRNAs (in black). The AUG codon and SD sequence are in green. The broken black arrows are the RNase III–induced cleavages.

Figure 4. Integrating the S. aureus sRNAs into gene regulation cascades.

The “agr-RNAIII” auto-activation circuit is indicated with the two feed-forward loops involving RNAIII. When reaching optimal density, the autoinducing peptide (AIP) activates the agr autocatalytic circuit, leading to RNAIII transcription. RNAIII represses the expression of rot, which activates spa transcription and represses that of hla. In the meantime, RNAIII also activates hla mRNA translation and represses spa mRNA translation. The plain and broken lines indicate the direct or indirect gene activations, respectively. The red lines indicate the down-regulations mediated by the various RNAs. The black question marks above the “see-sawing” triangles point to the unknown triggering factors. The transcriptional regulatory proteins are in blue. The complexity of this scheme will certainly increase as we learn more on the sRNA functions.