Fig. 6: Accumulation of peptidoglycan is associated with a GraRS-dependent reduction in autolysis.

Triton X-100-triggered lysis of TSB-grown and serum-adapted cultures of S. aureus as measured by following OD₆₀₀ values during a 6 h exposure to 0.05% Triton X-100 (a). Cell wall extracts of TSB-grown and serum-adapted S. aureus were separated by SDS-PAGE using gels containing heat-killed S. aureus cells. Hydrolases were allowed to refold and zones of peptidoglycan degradation were visualised using methylene blue (b). Triton X-100-triggered lysis of serum-adapted cultures of S. aureus WT, the graS::Tn mutant, and the graS::Tn mutant complemented with empty pCN34 or PgraXRS as measured by OD₆₀₀ values after a 6 h exposure to 0.05% Triton X-100 (c). Cell wall extracts of serum-adapted cultures of S. aureus WT, the graS::Tn mutant, and the graS::Tn mutant complemented with empty pCN34 or PgraXRS were analysed by zymography (d). Graphs in (a) and (c) represent the mean ± standard deviation of three or four independent experiments, respectively. Three independent replicates of zymography were carried out and one representative image is shown. Data in (a) were analysed by two-way ANOVA with Sidak’s post-hoc test; *, P = 0.0267 (120 min), 0.0031 (135 min), 0.0011 (150 min), 0.0005 (165 min) and <0.0001 at all time-points from 180 min onwards. Data in (c) were analysed by one-way ANOVA with Dunnett’s post-hoc test; WT vs mutants (* P = 0.0002 (graS::Tn), <0.0001 (pCN34)). TSB, tryptic soy broth; OD₆₀₀, optical density at 600 nm. Source data are provided as a Source Data file.