The Extracytoplasmic Function Sigma Factor σ ^S Protects against both Intracellular and Extracytoplasmic Stresses in Staphylococcus aureus

Abstract

Previously we identified a novel component of the Staphylococcus aureus regulatory network, an extracytoplasmic function -factor, S , involved in stress response and disease causation. Here we present additional characterization of S , demonstrating a role for it in protection against DNA damage, cell wall disruption, and interaction with components of the innate immune system. Promoter mapping reveals the existence of three unique sigS start sites, one of which appears to be subject to autoregulation. Transcriptional profiling revealed that sigS expression remains low in a number of S. aureus wild types but is upregulated in the highly mutated strain RN4220. Further analysis demonstrates that sigS expression is inducible upon exposure to a variety of chemical stressors that elicit DNA damage, including methyl methanesulfonate and ciprofloxacin, as well as those that disrupt cell wall stability, such as ampicillin and oxacillin. Significantly, expression of sigS is highly induced during growth in serum and upon phagocytosis by RAW 264.7 murine macrophage-like cells. Phenotypically, S mutants display sensitivity to a broad range of DNA-damaging agents and cell wall-targeting antibiotics. Furthermore, the survivability of S mutants is strongly impacted during challenge by components of the innate immune system. Collectively, our data suggest that S likely serves dual functions within the S. aureus cell, protecting against both cytoplasmic and extracytoplasmic stresses. This further argues for its important, and perhaps novel, role in the S. aureus stress and virulence responses. Staphylococcus aureus is an exceedingly virulent and successful pathogen, capable of causing a wide range of infections, from relatively benign skin lesions to life-threatening septicemia. With an overwhelming ability to adapt to its environment, S. aureus has become the most common cause of both hospital-and community-acquired infections and is believed to be the leading cause of death by a single infectious agent in the United States (20, 34). The threat posed by this organism to human health is further heightened by the rapid and continued emergence of multidrug-resistant isolates (1,20,34,43).Many components govern the adaptive nature of S. aureus, including complex regulatory networks, which allow it to respond to constantly changing environments via rapid shifts in gene expression. There are a number of different elements that mediate this fine-tuning, including DNA-binding proteins, two-component systems, regulatory RNAs, and alternative factors (10,11,18,21,22,32,44,50,51). The last class acts by binding to core RNA polymerase and redirecting promoter recognition to coordinate gene expression, bringing about expedient and wide-reaching alterations within the cell.From a classification perspective, factors are divided into five discrete subfamilies, with the essential housekeeping factors ( A or 70 ), which are responsible for the majority of transcription, constituting group 1. The remaining families (groups 2 to 5) contain alterna...

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The δ Subunit of RNA Polymerase Guides Promoter Selectivity and Virulence in Staphylococcus aureus

“…Quantitative real-time PCR (qPCR) analysis was conducted as described previously (20) using the rpoE gene-specific primers OL1976 and OL1978 and OL2981 and OL2982 and 16S rRNA primers OL1184 and OL1185. Three independent replicates were used to calculate final values.…”

Regulation of antimicrobial resistance by extracytoplasmic function (ECF) sigma factors

“…In these pathogens, other regulatory mechanisms, such as two-component systems may fulfill similar functions to compensate for the lack of ECF sigma factors [136–141]. Other pathogens, such as Haemophilus influenza, Neisseria gonnorhea, and Listeria monocytogenes have ECF sigma factors that are clearly involved in membrane stress responses, but their specific roles in antibiotic resistance have not been determined [142–145]. …”