Extensive and Genome-Wide Changes in the Transcription Profile ofStaphylococcus aureusInduced by Modulating the Transcription of the Cell Wall Synthesis GenemurF

Abstract

A murF conditional mutant was used to evaluate the effect of suboptimal transcription of this gene on the transcriptome of the methicillin-resistant Staphylococcus aureus strain COL. The mutant was grown in the presence of optimal and suboptimal concentrations of the inducer, and the relative levels of transcription of genes were evaluated genome wide with an Affymetrix DNA microarray that included all open reading frames (ORFs) as well as intergenic sequences derived from four sequenced S. aureus strains. Using a sensitivity threshold value of 1.5, suboptimal expression of murF altered the transcription of a surprisingly large number of genes, i.e., 668 out of the 2,740 ORFs (close to one-fourth of all ORFs), of the genome of S. aureus strain COL. The genes with altered transcription were distributed evenly around the S. aureus chromosome, and groups of genes involved with distinct metabolic functions responded in unique and operon-specific manners to modulation in murF transcription. For instance, all genes belonging to the isd operon and all but 2 of the 35 genes of prophage L54a were down-regulated, whereas all but one of the 21 members of the vraSR regulon and most of the 79 virulence-related genes (those for fibronectin binding proteins A and B, clumping factor A, gamma hemolysin, enterotoxin B, etc.) were up-regulated in cells with suboptimal expression of murF. Most importantly, the majority of these altered gene expression profiles were reversible by resupplying the optimal concentration of IPTG (isopropyl-␤-D-thiogalactopyranoside) to the culture. The observations suggest the coordinate regulation of a large sector of the S. aureus transcriptome in response to a disturbance in cell wall synthesis.Transcriptomic approaches are powerful tools for the identification of groups of genes with similar expression patterns and for the establishment of putative regulatory relations between them. Using microarray technology, recent reports have described extensive changes in gene expression profiles when Staphylococcus aureus was challenged by different cell wall inhibitors such as oxacillin, bacitracin, D-cycloserine, and vancomycin (12, 19, 20, 34). The group of genes whose transcription was altered during treatment with antibiotics that target different steps in cell wall synthesis was proposed to represent members of a cell wall stimulon responding to a specific form of stress in a coordinate manner.The large number and diversity of genes that respond to cell wall inhibitors by altered transcription suggest that many functions of the bacterial cell are connected through regulatory loops to one or more steps in cell wall synthesis which may also serve as control points of metabolism. Such a control should also be operational in the absence of antibiotic treatment, and we searched for a system in which the existence of such a regulatory system could be experimentally demonstrated.The method of choice became available with the isolation of an S. aureus murF conditional mutant in which the rate of transcription o...

The Pneumococcal Cell Envelope Stress-Sensing System LiaFSR Is Activated by Murein Hydrolases and Lipid II-Interacting Antibiotics

“…For S. aureus VraSR, the antibiotics D-cycloserine and tunicamycin, which inhibit the enzyme (MraY) catalyzing the synthesis of lipid I, and even ␤-lactams and reduction in the level of pbpB transcription have been found to induce the system (9, 23). Furthermore, suboptimal transcription of murF, an enzyme that adds the D-alanyl-D-alanine dipeptide to the UDP-linked MurNAc-tripeptide, results in increased transcription of the VraSR system in S. aureus (40).…”

cwrA, a gene that specifically responds to cell wall damage in Staphylococcus aureus

“…Understanding the molecular and cellular response that occurs subsequent to peptidoglycan biosynthesis inhibition is of critical importance in understanding how pathogens adapt to, resist and evade killing by what are typically bactericidal agents. Herein, we studied cwrA, a gene that has appeared as the highest-fold upregulated in a variety of transcriptomic studies examining cell wall inhibition (Balibar et al, 2009;McAleese et al, 2006;Sobral et al, 2007).…”

Reduced Vancomycin Susceptibility inStaphylococcus aureus, Including Vancomycin-Intermediate and Heterogeneous Vancomycin-Intermediate Strains: Resistance Mechanisms, Laboratory Detection, and Clinical Implications

“…Recently, a core and accessory set of cell wall-associated genes (the "cell wall stimulon") has been described based on the results of microarray transcriptional analysis experiments after the exposure of S. aureus to cell wall-active agents (92,168,329,361). These genes are predominately under the control of the two-component regulatory (2CR) system vraSR.…”