RsbU-Dependent Regulation of Staphylococcus epidermidis Biofilm Formation Is Mediated via the Alternative Sigma Factor σB by Repression of the Negative Regulator Gene icaR

摘要

Transposon mutagenesis of rsbU leads to a biofilm-negative phenotype in Staphylococcus epidermidis. However, the pathway of this regulatory mechanism was unknown. To investigate the role of RsbU in the regulation of the alternative sigma factor σ^B and biofilm formation, we generated different mutants of the σ^B operon in S. epidermidis strains 1457 and 8400. The genes rsbU, rsbV, rsbW, and sigB, as well as the regulatory cascade rsbUVW and the entire σ^B operon, were deleted. Transcriptional analysis of sarA and the σ^B-dependent gene asp23 revealed the functions of RsbU and RsbV as positive regulators and of RsbW as a negative regulator of σ^B activity, indicating regulation of σ^B activity similar to that characterized for Bacillus subtilis and Staphylococcus aureus. Phenotypic characterization of the mutants revealed that the dramatic decrease of biofilm formation in rsbU mutants is mediated via σ^B, indicating a crucial role for σ^B in S. epidermidis pathogenesis. However, biofilm formation in mutants defective in σ^B or its function could be restored in the presence of subinhibitory ethanol concentrations. Transcriptional analysis revealed that icaR is up-regulated in mutants lacking σ^B function but that icaA transcription is down-regulated in these mutants, indicating a σ^B-dependent regulatory intermediate negatively regulating IcaR. Supplementation of growth media with ethanol decreased icaR transcription, leading to increased icaA transcription and a biofilm-positive phenotype, indicating that the ethanol-dependent induction of biofilm formation is mediated by IcaR. This icaR-dependent regulation under ethanol induction is mediated in a σ^B-independent manner, suggesting at least one additional regulatory intermediate in the biofilm formation of S. epidermidis.