RsbT and RsbV Contribute to σ B -Dependent Survival under Environmental, Energy, and Intracellular Stress Conditions in Listeria monocytogenes

Abstract

ABSTRACT Sigma B (σ B ) is a stress-responsive alternative sigma factor that has been identified in various gram-positive bacteria. Seven different regulators of sigma B (Rsbs) are located in the sigB operons of both Bacillus subtilis and Listeria monocytogenes . In B. subtilis , these proteins contribute to regulation of σ B activity by conveying environmental and energy stress signals through two well-established branches of a signal transduction pathway. RsbT contributes to regulation of σ B activity in response to environmental stresses, while RsbV contributes to σ B activation under both environmental and energy stresses in B. subtilis . To probe L. monocytogenes Rsb roles in σ B -mediated responses to various stresses, in-frame deletions were created in rsbT and rsbV . Phenotypic characterization of the L. monocytogenes rsbT and rsbV null mutants revealed that both mutants were similar to the Δ sigB strain in their abilities to survive under environmental stress conditions (exposure to synthetic gastric fluid, pH 2.5, acidified brain heart infusion broth [BHI], or oxidative stress [13 mM cumene hydroperoxide]). Under energy stress conditions (carbon starvation in defined media, entry into stationary phase, or reduced intracellular ATP), both Δ rsbT and Δ rsbV showed survival reductions similar to that of the Δ sigB strain. These observations suggest that the pathways for Rsb-dependent regulation of σ B activity differ between L. monocytogenes and B. subtilis . As σ B also activates transcription of the L. monocytogenes prfA P2 promoter, we evaluated virulence-associated characteristics of Δ prfA P1 rsbT and Δ prfA P1 rsbV double mutants in hemolysis and tissue culture assays. Both double mutants showed identical phenotypes to Δ prfA P1P2 and Δ prfA P1 sigB double mutants, i.e., reduced hemolysis activity and reduced plaque size in mouse fibroblast cells. These findings indicate that RsbT and RsbV both contribute to σ B activation in L. monocytogenes during exposure to environmental and energy stresses as well as during tissue culture infection.