Opposing roles of the Staphylococcus aureus virulence regulators, Agr and Sar, in Triton X-100- and penicillin-induced autolysis - PubMed (original) (raw)

Opposing roles of the Staphylococcus aureus virulence regulators, Agr and Sar, in Triton X-100- and penicillin-induced autolysis

D F Fujimoto et al. J Bacteriol. 1998 Jul.

Abstract

The regulation of murein hydrolases is a critical aspect of peptidoglycan growth and metabolism. In the present study, we demonstrate that mutations within the Staphylococcus aureus virulence factor regulatory genes, agr and sar, affect autolysis, resulting in decreased and increased autolysis rates, respectively. Zymographic analyses of these mutant strains suggest that agr and sar exert their effects on autolysis, in part, by modulating murein hydrolase expression and/or activity.

PubMed Disclaimer

Figures

FIG. 1

FIG. 1

Effect of agr and sar on Triton X-100-induced autolysis. S. aureus RN6390 (wild type) (diamonds), RN6911 (agr mutant) (squares), ALC488 (sar mutant) (triangles), and ALC135 (agr sar mutant) (circles) were grown to an optical density at 580 nm of 0.6 to 0.8. Triton X-100-induced autolysis assays were performed as described by Mani et al. (8) except that the cells were grown in NZY broth (3% NZ amine, 1% yeast extract). Triton X-100-induced autolysis was measured as the decline of optical density versus time and is expressed as the percent of the initial optical density. The data presented are representative of three independent experiments.

FIG. 2

FIG. 2

Effect of agr and sar on penicillin-induced autolysis. Penicillin-induced autolysis of S. aureus RN6390 (wild type) (diamonds), RN6911 (agr mutant) (squares), ALC488 (sar mutant) (triangles), and ALC135 (agr sar mutant) (circles) was measured with a Klett-Summerson colorimeter (filter no. 60; 1 Klett unit = 5 × 106 CFU/ml). Penicillin G (final concentration of 0.4 μg/ml) was added to early-exponential-phase cells (20 Klett units) in tryptic soy broth at 37°C, and the changes in culture turbidity were monitored over an 8-h period. Autolysis was measured as the decline in culture turbidity versus time and is expressed as the percent of the initial Klett reading when penicillin was added. The data presented are representative of three independent experiments.

FIG. 3

FIG. 3

Effect of agr and sar on penicillin-induced killing. Penicillin-induced killing of S. aureus RN6390 (wild type) (diamonds), RN6911 (agr mutant) (squares), ALC488 (sar mutant) (triangles), and ALC135 (agr sar mutant) (circles) was examined by performing viable cell counts every 2 h on the penicillin-treated cultures for which data are presented in Fig. 2. The viability of the cultures was assessed immediately prior to and every 2 h (for 8 h) after the addition of penicillin G by performing serial dilutions and spreading the cells on tryptic soy broth agar medium. The data presented are representative of three independent experiments.

FIG. 4

FIG. 4

Zymographic analysis of S. aureus murein hydrolases. Cell wall-associated proteins (1.5 μg) were extracted from S. aureus cells grown to early exponential phase (optical density at 580 nm of 0.6 to 0.8) in NZY broth and analyzed as described by Qoronfleh and Wilkinson (14). Murein hydrolase activities were visualized as zones of hydrolysis by staining the gel with methylene blue. Lane 2, RN6390 (wild type); lane 3, RN6911 (agr mutant); lane 4, ALC488 (sar mutant); lane 5, ALC135 (agr sar mutant). The sizes of the prestained broad-range MW standards (Bio-Rad Laboratories, Hercules, Calif.) are given in kilodaltons (lane 1).

Similar articles

Cited by

References

    1. Archibald A R, Hancock I C, Harwood C R. Cell wall structure, synthesis, and turnover. In: Sonenshein A L, Hoch J A, Losick R, editors. Bacillus subtilis and other gram-positive bacteria. Washington, D.C: American Society for Microbiology; 1993. pp. 381–410.
    1. Brunskill E W, Bayles K W. Identification and molecular characterization of a putative regulatory locus that affects autolysis in Staphylococcus aureus. J Bacteriol. 1996;178:611–618. - PMC - PubMed
    1. Brunskill E W, Bayles K W. Identification of LytSR-regulated genes from Staphylococcus aureus. J Bacteriol. 1996;178:5810–5812. - PMC - PubMed
    1. Foster S J. Molecular characterization and functional analysis of the major autolysin of Staphylococcus aureus 8325/4. J Bacteriol. 1995;177:5723–5725. - PMC - PubMed
    1. Foster S J. Analysis of the autolysins of Bacillus subtilis 168 during vegetative growth and differentiation by using renaturing polyacrylamide gel electrophoresis. J Bacteriol. 1992;174:464–470. - PMC - PubMed

Publication types

MeSH terms

Substances

LinkOut - more resources