SpoVG Regulates Cell Wall Metabolism and Oxacillin Resistance in Methicillin-Resistant Staphylococcus aureus Strain N315

Abstract

bIncreasing cases of infections caused by methicillin-resistant Staphylococcus aureus (MRSA) strains in healthy individuals have raised concerns worldwide. MRSA strains are resistant to almost the entire family of ␤-lactam antibiotics due to the acquisition of an extra penicillin-binding protein, PBP2a. Studies have shown that spoVG is involved in oxacillin resistance, while the regulatory mechanism remains elusive. In this study, we have found that SpoVG plays a positive role in oxacillin resistance through promoting cell wall synthesis and inhibiting cell wall degradation in MRSA strain N315. Deletion of spoVG in strain N315 led to a significant decrease in oxacillin resistance and a dramatic increase in Triton X-100-induced autolytic activity simultaneously. Real-time quantitative reverse transcription-PCR revealed that the expression of 8 genes related to cell wall metabolism or oxacillin resistance was altered in the spoVG mutant. Electrophoretic mobility shift assay indicated that SpoVG can directly bind to the putative promoter regions of lytN (murein hydrolase), femA, and lytSR (the two-component system). These findings suggest a molecular mechanism in which SpoVG modulates oxacillin resistance by regulating cell wall metabolism in MRSA. Staphylococcus aureus is a versatile and dangerous pathogen in humans, especially while the frequency of staphylococcal infections caused by methicillin-resistant Staphylococcus aureus (MRSA) increases steadily (1). S. aureus has four penicillin-binding proteins (PBPs) which are involved in the last stages of peptidoglycan synthesis (2). MRSA strains have acquired an extra PBP (PBP2a), encoded by the mecA gene on the staphylococcal cassette chromosome mec (SCCmec), which has a remarkably lower affinity for ␤-lactams than does PBP2 and is responsible for resisting these drugs (3). Besides the expression of PBP2a, which is responsible for a higher level of ␤-lactam resistance, S. aureus has another primary ␤-lactam resistance mechanism: the expression of ␤-lactamase enzymes, encoded by the blaZ gene, which hydrolyze ␤-lactams such as penicillin. In addition, several additional native genes, such as vraSR, pbp4, and fem (factor essential for methicillin resistance), have been identified as being essential to the full expression of oxacillin resistance (4-6). Inactivation of femA and pbp4 genes in the presence of an intact mecA gene usually results in strains with a low resistance to oxacillin. The VraS/VraR twocomponent regulatory system regulates the genes which are associated with cell wall biosynthesis, such as pbp2, sgtB, and murZ (7).Most MRSA and glycopeptide-intermediately resistant Staphylococcus aureus (GISA) isolates are resistant to Triton X-100-induced autolysis, while instances of reduced resistance have been observed with concomitant increases of the autolysis rate (8-11). The expression of autolytic enzymes, including Atl, LytM, LytN, and Sle1 (12-15), is controlled by pleiotropic regulators, such as MgrA, SarV, SarA, LytSR, and ArlSR (autolysis-related locus...

Strain-Specific Metabolic Requirements Revealed by a Defined Minimal Medium for Systems Analyses of Staphylococcus aureus

“…The SigB transcription factor is related to the expression of high-level methicillin resistance and regulated in response to growth phase and environmental stress (38,39). Moreover, spoVG, a SigB-controlled protein, is a critical factor that impacts cell wall synthesis, and it is likewise involved in antibiotic resistance and virulence (40,41).…”

Role of the msaABCR Operon in Cell Wall Biosynthesis, Autolysis, Integrity, and Antibiotic Resistance in Staphylococcus aureus

“…WTAs play a crucial role in septal localization of autolysins in S. aureus cells and interact with them, thereby regulating autolysis (4,19,20,43,55,56). WTAs are also known to have an important role in peptidoglycan cross-linking, as well as in ␤-lactam and vancomycin resistance, determining septal localization of PBP4 and FmtA (9-11, 16, 17).…”

“…Overall, these findings suggest that the extra murein hydrolase bands in zymogram analysis produced by the msaABCR mutants in both VISA and MRSA strains were due to increased AtlA production as well as increased processing of AtlA and/or other murein hydrolases by the increased activity of proteases. Therefore, increased protease- Previous studies have shown that WTAs are cell-surface glycopolymers that are covalently attached to peptidoglycan and modulate AtlA murein hydrolase activity as well as its localization (19,43). In addition, WTAs are also known to play a role in PBP4 localization, lysozyme resistance, and cell wall cross-linking (10,11,31).…”

“…Several recent studies have uncovered the role of other factors/genes involved in the biosynthesis of cell envelope components (such as peptidoglycan and teichoic acids), in modulating cell wall hydrolase activities of ␤-lactams, and in vancomycin resistance. These reports have rekindled interest in targeting genes involved in the maintenance of cell wall integrity and ultimately resensitizing resistant strains (4,7,(13)(14)(15)(16)(17)(18)(19)(20). Previously, we showed that the msaABCR operon plays a role in biofilm development, virulence, persister cell formation, and antibiotic resistance in S. aureus (21)(22)(23)(24)(25)(26).…”

Role of substrate temperature at graphene synthesis in an arc discharge

“…It has been shown to successfully synthesize graphene flakes with and without the enhancement of a magnetic field. 25,26,27,28,29,30,31 Graphene was synthesized using anodic arc ablation in a helium environment at atmospheric pressure. Samples were collected from the chamber wall or the surface of the magnet, and were characterized with SEM, TEM, AFM and RAMAN spectroscope.…”