The sae locus of Staphylococcus aureus encodes a two-component regulatory system (original) (raw)
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Regulation of exoprotein expression in Staphylococcus aureus by a locus (sar) distinct from agr
Proceedings of the National Academy of Sciences, 1992
A single insertion of transposon Tn917LTV1 into the chromosome ofa Staphylococcus aureus Cdinical isolate, strain DB, resulted in a pleiotropic effect on the expression of a number of extraceilular and cell-wall-as ted proteins. Detailed comparison ofphenotype associated with the mutant, 11D2, and the parent, DB, indicted that the chromosomal locus inactivated as a result of transposon mutagenesis differs from the S. aureus accessory gene regulator locus (agr). In particular, the expression of a-hemolysin, which is not detectable in Agr mutants, was enhanced in mutant 11D2, while it remained at a low level in strain DB. Likewise, protease activity was significantly enhanced in 11D2 compared with DB. In addition, most of the cell-bound proteins were expressed at lower levels in the mutant than the parent strain. This pattern is contrary to that found in switching from Agr+ to Agrphenotypes. Southern blot hybridization with an agr probe indicated that the inactivated chromosomal locus is distinct from agr. Transduction experiments demonstrated that the phenotypes associated with mutant 11D2 could be transferred to the parental strain DB as well as to RN450, an S. aureus strain with a genetic background similar to strain 8325-4. This locus on the S. aureus chromosome, possibly regulatory in nature, has been designated sar for staphylococcal accessory regulator.
Studies on the Expression of Regulatory Locus sae in Staphylococcus aureus
Current Microbiology, 2003
Global regulatory locus sae consists of a two-component signal transduction system coded by saeR and saeS genes that upregulates the transcription of several exoproteins. Northern analysis carried out in this study reveals the synthesis at late and post-exponential phases of a cotranscript of saeR and saeS structural genes of about 2.4 kb. This transcript is diminished in the isogenic agr::tetM mutant. Likewise, transcriptional fusion experiments show that sae expression is downregulated in the agr null mutant. Complementation analyses with plasmids carrying fragments of about 1.2 or 0.2 kbp upstream of saeR-saeS genes, which restore fully or only partially, respectively, the wild-type phenotype to the sae mutant, are in agreement with two initiation start points of transcription revealed by primer extension experiments. This work, as well as previous studies, reveals a complex hierarchical regulatory network involving several loci that control the expression of virulence determinants in S. aureus. Staphylococcus aureus is a major pathogen of man and animals; it synthesizes a large number of extracellular and cell wall-associated proteins that contribute to its virulence. Several global regulatory loci, such as agr, sar and sae, have been found to regulate the production of these virulence factors [2, 7, 13]. The agr locus consists of two divergent transcripts, RNAIII transcribed from the P3 promoter, which encodes ␦-hemolysin and acts in the regulation of secretory and cell wall-associated proteins, and RNAII, transcribed from the P2 promoter, which encodes the products of agrB, D, C, and A. Genes agrD and agrB encode and process an autoinducing peptide that acts as a signal that activates the expression of RNAII and RNAIII through a two-component signal system coded by agrC and agrA. Activation of RNAIII leads to increased production of ␣and -hemolysins, serine protease, DNase, and other exotoxins and to repression of the production of protein A, coagulase, and other cell wall-associated proteins [12, 14, 18]. The main product of the sar locus is a DNA-binding protein, SarA, which is transcribed from three different promoters. SarA acts on the expression of virulent factors indirectly by upregulating RNAIII transcription and also stimulating or repressing the transcription of several virulence genes such as hla, fnbA, cna, spa, and ssp (which code for ␣-hemolysin, fibronectin, and collagenbinding proteins, proteinA and serin-protease, respectively) in an agr-independent way [1, 4, 15, 25]. The regulatory locus designated sae (for S. aureus exoprotein expression) encodes a two-component regulatory system involving saeR, a response regulator, and saeS, a histidine protein kinase, of 687 and 1062 bp, respectively, and upregulates the production of ␣and -hemolysins, DNase, and coagulase at the transcriptional level [9, 10]. More recently, other virulence regulators have been described. The sarH1 or sarS locus, which belongs to a family of sar homologs, represses the transcription of ␣-hemolysin and activates spa transcription while its own transcription is repressed by sarA and agr [3, 23]. Other characterized genes of this family are sarT, which like sarH1 is negatively controlled by sarA and agr and † Died October 11, 2000.
Journal of Bacteriology, 2003
We characterized the sae operon, a global regulator for virulence gene expression in Staphylococcus aureus. A Tn917 sae mutant was obtained by screening a Tn917 library of the agr mutant ISP479Mu for clones with altered hemolytic activity. Sequence analysis of the sae operon revealed two additional open reading frames (ORFs) (ORF3 and ORF4) upstream of the two-component regulatory genes saeR and saeS. Four overlapping sae-specific transcripts (T1 to T4) were detected by Northern blot analysis, and the transcriptional initiation points were mapped by primer extension analysis. The T1, T2, and T3 mRNAs are probably terminated at the same stem-loop sequence downstream of saeS. The T1 message (3.1 kb) initiates upstream of ORF4, T2 (2.4 kb) initiates upstream of ORF3, and T3 (2.0 kb) initiates in front of saeR. T4 (0.7 kb) represents a monocistronic mRNA encompassing ORF4 only. sae-specific transcripts were detectable in all of the 40 different clinical S. aureus isolates investigated. Transcript levels were at maximum during the post-exponential growth phase. The sae mutant showed a significantly reduced rate of invasion of human endothelial cells, consistent with diminished transcription and expression of fnbA. The expression of type 5 capsular polysaccharide is activated in the sae mutant of strain Newman, as shown by immunofluorescence and promoter-reporter fusion experiments. In summary, the sae operon constitutes a four-component regulator system which acts on virulence gene expression in S. aureus.
Regulatory organization of the staphylococcal sae locus
Microbiology, 2008
This paper describes an investigation of the complex internal regulatory circuitry of the staphylococcal sae locus and the impact of modifying this circuitry on the expression of external genes in the sae regulon. The sae locus contains four genes, the saeR and S two ...
Journal of Bacteriology, 2006
The two-component system SaeRS consisting of the histidin kinase SaeS and the response regulator SaeR is known to act on virulence gene expression in Staphylococcus aureus. In order to get a more comprehensive picture on SaeR-regulated genes, we studied the contribution of the two-component system on global gene expression by using both the proteomic and transcriptomic approach. Altogether, a loss of SaeRS resulted in a decreased amount of at least 17 extracellular proteins and two cell surface-associated proteins, among them several important virulence factors such as HlgA, HlgB, HlgC, LukF, and LukM. SaeRS activates the expression of these genes at the transcriptional level. The amount of the five proteins Aur, SspA, SsaA, Plc, and GlpQ was negatively influenced by SaeRS. However, the transcription of the corresponding genes was not affected by the two-component system. SaeRS had also no measurable influence on the transcription of the regulatory genes agr, sarA, arlRS, and sigB that contribute to the regulation of SaeRS-dependent virulence factors identified in this investigation. Our results clearly show that SaeRS is strongly involved in the tight temporal control of virulence factor expression in S. aureus. Its precise role within the regulatory network remains to be determined.
In vivo expression of exoprotein synthesis with a Sae mutant of Staphylococcus aureus
Canadian journal of veterinary research = Revue canadienne de recherche vétérinaire, 1996
The expression of exoprotein synthesis of Staphylococcus aureus Sae mutant RC121 and its parental strain was studied under in vivo growth conditions. Cultures of both strains were inoculated into dialysis sacs implanted in sheep peritoneum. Results indicated that similar to in vitro grown mutant cells, Sae mutant RC121 shows diminished synthesis of alpha- and beta-hemolysin, coagulase, DNase and protein A. However, in vitro and in vivo grown mutant cultures showed different exoprotein profiles in SDS-PAGE; some bands from in vivo mutant cultures were diminished or missing and others appeared as more concentrated, when compared with the pattern of the in vivo grown parental strain, while all the exoprotein bands from the in vitro cultures of the mutant were diminished or missing as compared to the in vitro grown parental strain. The virulence of the Sae mutant, assayed by intraperitoneal injection in mice, was lower than that of the parental strain after both in vivo and in vitro gro...
The SaeRS Two-Component System of Staphylococcus aureus
In the Gram-positive pathogenic bacterium Staphylococcus aureus, the SaeRS two-component system (TCS) plays a major role in controlling the production of over 20 virulence factors including hemolysins, leukocidins, superantigens, surface proteins, and proteases. The SaeRS TCS is composed of the sensor histidine kinase SaeS, response regulator SaeR, and two auxiliary proteins SaeP and SaeQ. Since its discovery in 1994, the sae locus has been studied extensively, and its contributions to staphylococcal virulence and pathogenesis have been well documented and understood; however, the molecular mechanism by which the SaeRS TCS receives and processes cognate signals is not. In this article, therefore, we review the literature focusing on the signaling mechanism and its interaction with other global regulators.
Journal of Bacteriology, 2010
Staphylococcus aureus uses the SaeRS two-component system to control the expression of many virulence factors such as alpha-hemolysin and coagulase; however, the molecular mechanism of this signaling has not yet been elucidated. Here, using the P1 promoter of the sae operon as a model target DNA, we demonstrated that the unphosphorylated response regulator SaeR does not bind to the P1 promoter DNA, while its C-terminal DNA binding domain alone does. The DNA binding activity of full-length SaeR could be restored by sensor kinase SaeS-induced phosphorylation. Phosphorylated SaeR is more resistant to digestion by trypsin, suggesting conformational changes. DNase I footprinting assays revealed that the SaeR protection region in the P1 promoter contains a direct repeat sequence (GTTAAN 6 GTTAA [where N is any nucleotide]). This sequence is critical to the binding of phosphorylated SaeR. Mutational changes in the repeat sequence greatly reduced both the in vitro binding of SaeR and the in vivo function of the P1 promoter. From these results, we concluded that SaeR recognizes the direct repeat sequence as a binding site and that binding requires phosphorylation by SaeS.
FEMS Microbiology Letters, 1998
The presence of sequences related to the agr of Staphylococcus aureus was demonstrated in Staphylococcus epidermidis by agr-specific PCR, and Southern blot. The agr-like locus of S. epidermidis A086 was cloned and sequenced. An overall homology of 68% was found between the agr locus from S. epidermidis and S. aureus. The agr locus from S. epidermidis was organized similar to those from S. aureus and S. lugdunensis. The putative RNAII molecule contains four open reading frames, agrA, B, C and D. AgrA was a response regulator. AgrB showed homology with transducer and translocase molecules. AgrC is expected to act as a histidine protein kinase in which a leucine zipper is present. AgrD is presumably processed into an autoinducer peptide. The putative RNAIII molecule contained an open reading frame encoding a putative 26 amino acid (aa) polypeptide, which differed in 3 aa from the RNAIII encoded N-toxin of S. aureus. Kinetic studies showed that the production of this RNAIII was elevated during the post-exponential phase. N-Toxin activity was demonstrated for 21 of 23 tested S. epidermidis strains. Kinetic studies of the production of N-toxin showed that the toxin was produced during the postexponential phase. Sequencing of S. epidermidis A097, which showed a delayed agr-response, revealed a truncated AgrC lacking the histidine kinase domain. These data indicate that an agr-like locus is active in S. epidermidis during the postexponential phase. z
Differential Target Gene Activation by the Staphylococcus aureus Two-Component System saeRS
Journal of Bacteriology, 2010
The saePQRS system of Staphylococcus aureus controls the expression of major virulence factors and encodes a histidine kinase (SaeS), a response regulator (SaeR), a membrane protein (SaeQ), and a lipoprotein (SaeP). The widely used strain Newman is characterized by a single amino acid change in the sensory domain of SaeS (Pro18 in strain Newman [SaeS P ], compared with Leu18 in other strains [SaeS L ]). SaeS P determines activation of the class I sae target genes (coa, fnbA, eap, sib, efb, fib, sae), which are highly expressed in strain Newman. In contrast, class II target genes (hla, hlb, cap) are not sensitive to the SaeS polymorphism. The SaeS L allele (saeS L ) is dominant over the SaeS P allele, as shown by single-copy integration of saePQRS L in strain Newman, which results in severe repression of class I target genes. The differential effect on target gene expression is explained by different requirements for SaeR phosphorylation. From an analysis of saeS deletion strains and strains with mutated SaeR phosphorylation sites, we concluded that a high level of SaeR phosphorylation is required for activation of class I target genes. However, a low level of SaeR phosphorylation, which can occur independent of SaeS, is sufficient to activate class II target genes. Using inducible saeRS constructs, we showed that the expression of both types of target genes is independent of the saeRS dosage and that the typical growth phase-dependent gene expression pattern is not driven by SaeRS.