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Rsp promotes the transcription of virulence factors in an agr-independent manner in Staphylococcus aureus.

ABSTRACT: Staphylococcus aureus is a major human pathogen that causes a great diversity of community- and hospital-acquired infections. Rsp, a member of AraC/XylS family of transcriptional regulators (AFTRs), has been reported to play an important role in the regulation of virulence determinants in S. aureus via an agr-dependent pathway. Here we demonstrated that Rsp could bind to the rsp promoter to positively regulate its own expression. We then constructed an isogenic rsp deletion strain and compared the haemolysis in the wild-type and rsp mutant strains. Our results indicated that the rsp mutant strain displayed decreased haemolytic activity, which was correlated with a dramatic decrease in the expression of hla and psm. Furthermore, we analysed the regulatory effects of Rsp in the agr mutant strain and found that they are agr-independent. Electrophoretic mobility shift assay indicated that Rsp can directly bind to the promoter regions of hla and psm. The mouse model of subcutaneous abscess showed that the rsp mutant strain displayed a significant defect in virulence compared to the wild-type strain. These findings reveal that Rsp positively regulates the virulence of S. aureus by promoting the expression of hla and psm through direct binding to their promoter regions.

SUBMITTER: Liu B

PROVIDER: S-EPMC7241556 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:The expression of most Staphylococcus aureus virulence factors is controlled by the agr locus, which encodes a two-component signaling pathway whose activating ligand is an agr-encoded autoinducing peptide (AIP). A polymorphism in the amino acid sequence of the AIP and of its corresponding receptor divides S. aureus strains into four major groups. Within a given group, each strain produces a peptide that can activate the agr response in the other member strains, whereas the AIPs belonging to different groups are usually mutually inhibitory. We investigated a possible relationship between agr groups and human S. aureus disease by studying 198 S. aureus strains isolated from 14 asymptomatic carriers, 66 patients with suppurative infection, and 114 patients with acute toxemia. The agr group and the distribution of 24 toxin genes were analyzed by PCR, and the genetic background was determined by means of amplified fragment length polymorphism (AFLP) analysis. The isolates were relatively evenly distributed among the four agrgroups, with 61 strains belonging to agr group I, 49 belonging to group II, 43 belonging to group III, and 45 belonging to group IV. Principal coordinate analysis performed on the AFLP distance matrix divided the 198 strains into three main phylogenetic groups, AF1 corresponding to strains of agr group IV, AF2 corresponding to strains of agr groups I and II, and AF3 corresponding to strains of agr group III. This indicated that the agr type was linked to the genetic background. A relationship between genetic background, agr group, and disease type was observed for several toxin-mediated diseases: for instance, agr group IV strains were associated with generalized exfoliative syndromes, and phylogenetic group AF1 strains with bullous impetigo. Among the suppurative infections, endocarditis strains mainly belonged to phylogenetic group AF2 and agr groups I and II. While these results do not show a direct role of the agr type in the type of human disease caused by S. aureus, the agr group may reflect an ancient evolutionary division of S. aureus in terms of this species' fundamental biology.

Project description:BACKGROUND:The Cnt system is crucial for the optimal import of essential metals in metal-limiting conditions and contributes to virulence in S. aureus. In a screen for regulators of efflux pumps in a phage-based ultra-high-density transposon library, we identified Rsp as a candidate regulator of the cntE gene. RESULTS:A two-fold decrease in expression of all genes of the cnt operon was observed by RT-qPCR in the rsp mutant compared to the parental strain, indicating that Rsp acts as an activator of the cnt operon. To determine whether the Rsp activation depends on iron, we compared mutant and parent cnt expression under varying metal conditions. A 2-fold reduction in cnt gene expression was detected in the rsp mutant in TSB, and a slightly smaller decrease (1.9, 1.7, and 1.5-fold changes for cntK, cmtA, and cntE respectively) was observed after addition of dipyridyl. The greatest decrease was seen with addition of FeSO4 (4.1, 5.3 and 6.3-fold changes for cntK, cmtA and cntE respectively). These findings suggest that Rsp activates the cnt operon in low and high iron conditions. To study the relationship between Rsp and the cnt repressors Fur and Zur, we created single and double mutants. Both fur and zur single mutants had significant increases in cnt gene expression compared to the parental strain, as did the fur rsp double mutant. The zur rsp double mutant also had a significant increase in cntK expression and a trend in increases in cntA and cntE expression just below statistical significance. Thus, the ability of Fur and Zur to repress cnt gene expression are not eliminated by the presence of Rsp. However, there were significantly smaller increases in cnt gene expression in the double mutants compared to single mutants, suggesting that Rsp activation can still occur in the absence of these repressors. To determine if Rsp directly modulates expression of cnt genes, incubation of purified Rsp caused a DNA-specific band shift for the cntK and cntA promoters. CONCLUSIONS:Rsp activation may act to maintain basal cellular levels of staphylopine to scavenge free metals when needed, in addition to metal dependent regulation by Fur and Zur.

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Rsp promotes the transcription of virulence factors in an agr-independent manner in Staphylococcus aureus.

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