σ ^B and the σ ^B -Dependent arlRS and yabJ-spoVG Loci Affect Capsule Formation in Staphylococcus aureus

Abstract

The alternative transcription factor B of Staphylococcus aureus affects the transcription of the cap gene cluster, required for the synthesis of capsular polysaccharide (CP), although this operon is lacking an apparent B -dependent promoter. Regulation of cap expression and CP production in S. aureus strain Newman was shown here to be influenced by B , the two-component signal transduction regulatory system ArlRS, and the yabJ-spoVG locus to different extents. Inactivation of arlR or deletion of the sigB operon strongly suppressed capA (CP synthesis enzyme A) transcription. Deletion of spoVG had a polar effect on yabJ-spoVG transcription and resulted in a two-to threefold decrease in capA transcription. Interestingly, immunofluorescence showed that CP production was strongly impaired in all three mutants, signaling that the yabJ-spoVG inactivation, despite its only partial effect on capA transcription, abolished capsule formation. trans-Complementation of the ⌬spoVG mutant with yabJ-spoVG under the control of its native promoter restored CP-5 production and capA expression to levels seen in the wild type. Northern analyses revealed a strong impact of B on arlRS and yabJ-spoVG transcription. We hypothesize that ArlR and products of the yabJ-spoVG locus may serve as effectors that modulate B control over B -dependent genes lacking an apparent B promoter.Staphylococcus aureus is a major nosocomial pathogen with the ability to cause a variety of diseases, including life-threatening infections. Like most microorganisms that are able to cause invasive diseases, S. aureus produces extracellular capsular polysaccharides (CPs), which are thought to be of importance in pathogenesis (reviewed in reference 35). Although 11 serologically distinct CPs were identified in S. aureus, the majority of clinical isolates produce CPs of serotype 5 (CP-5) or serotype 8 (CP-8). CPs protect S. aureus against opsonophagocytic killing by polymorphonuclear leukocytes (16,17,25,53,56) and enhance virulence in a number of animal models of staphylococcal infection (34,40,53,54,57). Expression of CPs is known to be influenced by various environmental signals in vitro and in vivo (reviewed in references 35 and 56), and transcription of the cap operon was shown to be modulated by regulatory elements, such as arlRS, agr, ccpA, mgr, sae, and sarA (7,8,23,24,26,27,39,48,52,55). Recent microarray analyses added the alternative factor B to the regulatory network controlling cap operon expression (3, 38) and indicated B to control capA transcription in a growth phasedependent manner (3). However, the lack of an apparent B consensus sequence in the promoter of capA suggested that B regulates cap transcription indirectly. Candidates for such downstream-acting regulators might be ArlRS and SarA, which are positively controlled by B in S. aureus (2, 3), although SarA was previously shown to have only a minor effect on cap expression and CP production in S. aureus (24). RNAIII of the agr locus, known to positively affect capA expression (7, 24, 55), could ...

The σ B -Dependent yabJ-spoVG Operon Is Involved in the Regulation of Extracellular Nuclease, Lipase, and Protease Expression in Staphylococcus aureus

“…The molecular mechanisms of YabJ, which belongs to the YjgF protein family of unknown biochemical function (49), and of the stage V sporulation protein G SpoVG (30), originally identified in Bacillus subtilis as being involved in the formation of the spore cortex, are yet unknown for the nonsporulating S. aureus. However, the finding that inactivation of the staphylococcal yabJ-spoVG operon, like inactivation of B , reduces the transcription of the capA gene, which is not preceded by a B promoter (36), and reduces the resistance level of methicillin-resistant S. aureus (MRSA) and glycopeptide intermediate-resistant S. aureus (GISA) (46) support the hypothesis that the yabJ-spoVG operon may encode a B downstream regulatory element. Early work on B activity in S. aureus showed that extracellular nuclease and lipase activities are negatively regulated by the alternative sigma factor B via a yet unknown indirect mechanism (31).…”

“…For this purpose we complemented SM148 and the B mutant IK184 with either gene separately or both genes together, expressed from the B -independent constitutive promoter. In analogy to the complementation of capA transcription in a ⌬yabJ-spoVG background, where SpoVG was shown to be the effector of the yabJ-spoVG operon (36,46), the plasmids pspoVG and pyabJspoVG did largely restore nuc expression (27). Sequences matching the GNNTNN-12-15-NGGTAN consensus, with no more than three mismatches of the degenerated bases from the GTTTAA-12-15-GGGTAT sequence and lying within a range of 500 bp upstream of the ORF were accepted.…”

“…The B -controlled yabJ-spoVG operon has pleiotropic effects in S. aureus, stimulating cap operon transcription and capsule formation, and supporting resistance to cell wall antibiotics (36,46). To explore the extent of the yabJ-spoVG regulon, we used full-genome DNA microarrays and compared the transcriptional patterns of S. aureus Newman and its isogenic ⌬yabJ-spoVG mutant, SM148, in the early stationary growth phase, when B activity and spoVG transcription are known to be maximal (7,36,47). Forty-three genes belonging to 27 predicted transcriptional units were found to be differently regulated.…”

“…The bicistronic yabJ-spoVG operon, whose transcription depends heavily on B activity, was hypothesized to play a role in the indirect B -dependent regulation of genes lacking an obvious B promoter (7,36,46). The molecular mechanisms of YabJ, which belongs to the YjgF protein family of unknown biochemical function (49), and of the stage V sporulation protein G SpoVG (30), originally identified in Bacillus subtilis as being involved in the formation of the spore cortex, are yet unknown for the nonsporulating S. aureus.…”

MsaB activates capsule production at the transcription level in Staphylococcus aureus

“…The deletion of the msaABCR operon alters the expression of several global regulators, including sarA, agr and sigB, all of which have been shown to be indirectly involved in capsule production (Bischoff et al, 2004;Pané-Farré et al, 2006;Meier et al, 2007;Dassy et al, 1993;Sahukhal & Elasri, 2014). The regulatory mechanism for msaABCR has not yet been defined.…”