Deletion of hypothetical wall teichoic acid ligases in Staphylococcus aureus activates the cell wall stress response

Abstract

The Staphylococcus aureus cell wall stress stimulon (CWSS) is activated by cell envelope-targeting antibiotics or depletion of essential cell wall biosynthesis enzymes. The functionally uncharacterized S. aureus LytR-CpsA-Psr (LCP) proteins, MsrR, SA0908 and SA2103, all belong to the CWSS. Although not essential, deletion of all three LCP proteins severely impairs cell division. We show here that VraSR-dependent CWSS expression was up to 250-fold higher in single, double and triple LCP mutants than in wild type S. aureus in the absence of external stress. The LCP triple mutant was virtually depleted of wall teichoic acids (WTA), which could be restored to different degrees by any of the single LCP proteins. Subinhibitory concentrations of tunicamycin, which inhibits the first WTA synthesis enzyme TarO (TagO), could partially complement the severe growth defect of the LCP triple mutant. Both of the latter findings support a role for S. aureus LCP proteins in late WTA synthesis, as in Bacillus subtilis where LCP proteins were recently proposed to transfer WTA from lipid carriers to the cell wall peptidoglycan. Intrinsic activation of the CWSS upon LCP deletion and the fact that LCP proteins were essential for WTA-loading of the cell wall, highlight their important role(s) in S. aureus cell envelope biogenesis.

LytR-CpsA-Psr Enzymes as Determinants of Bacillus anthracis Secondary Cell Wall Polysaccharide Assembly

“…In agreement with this model, the crystal structure of recombinant Cps2A and TagT revealed polyprenyl lipids bound to the active site as well as intrinsic pyrophosphatase activity for both proteins (29,30). The genome of Staphylococcus aureus contains three lcp genes, designated lcpA, lcpB, and lcpC (31,32). Staphylococcal mutants lacking all three lcp genes (⌬lcp), although viable, display growth defects, are unable to attach either WTA or capsular polysaccharide to pepti-doglycan, and release small amounts of these polymers into the extracellular medium (32)(33)(34).…”

“…Mutant alleles were verified by PCR using chromosomal DNA as a template and primers flanking the cloning sites. S. aureus MSSA1112 and its isogenic variant lacking lcpA, lcpB, and lcpC (⌬lcp) have been described previously (31). For complementation studies, B. anthracis lcp genes were cloned into pWWW412 and expressed via the constitutive hprK promoter of S. aureus (45).…”

SagB Glucosaminidase Is a Determinant of Staphylococcus aureus Glycan Chain Length, Antibiotic Susceptibility, and Protein Secretion

“…6). The genetic requirements for staphylococcal replication are known to vary among different strains, which was previously observed for LytR-CpsA-Psr (LCP) enzymes immobilizing secondary cell polymers via murein linkage units to bacterial peptidoglycan (41,74). Thus, N-acetylglucosaminidase enzymes may represent yet another example for genetic heterogeneity in S. aureus.…”

Pathways and roles of wall teichoic acid glycosylation in Staphylococcus aureus

“…(tagGH), and d-alanylation (dltABCD) of S. aureus WTA were identified and studied in genetic and/or in vitro reconstitution approaches (Swoboda et al, 2010;Xia et al, 2010a). Recently the ligation of WTA from lipid-linked WTA precursors to peptidoglycan was proposed to be accomplished by a new family of enzymes named LCP proteins (Chan et al, 2013;Dengler et al, 2012;Kawai et al, 2011) ( Fig. 1A and B).…”