Role of σBin Regulating the Compatible Solute Uptake Systems ofListeria monocytogenes: Osmotic Induction ofopuCIs σBDependent

Abstract

The regulation of the compatible solute transport systems inListeria monocytogenesby the stress-inducible sigma factor σ^Bwas investigated. Using wild-type strain 10403S and an otherwise isogenic strain carrying an in-frame deletion insigB, we have examined the role of σ^Bin regulating the ability of cells to utilize betaine and carnitine during growth under conditions of hyperosmotic stress. Cells lacking σ^Bwere defective for the utilization of carnitine but retained the ability to utilize betaine as an osmoprotectant. When compatible solute transport studies were performed, the initial rates of uptake of both betaine and carnitine were found to be reduced in thesigBmutant; carnitine transport was almost abolished, whereas betaine transport was reduced to approximately 50% of that of the parent strain. Analysis of the cytoplasmic pools of compatible solutes during balanced growth revealed that both carnitine and betaine steady-state pools were reduced in thesigBmutant. Transcriptional reporter fusions to theopuC(which encodes an ABC carnitine transporter) andbetL(which encodes an a secondary betaine transporter) operons were generated by using a promoterless copy of thegusgene fromEscherichia coli. Measurement of β-glucuronidase activities directed byopuC-gusandbetL-gusrevealed that transcription ofopuCis largely σ^Bdependent, consistent with the existence of a potential σ^Bconsensus promoter motif upstream fromopuCA. The transcription ofbetLwas found to besigBindependent. Reverse transcriptase PCR experiments confirmed these data and indicated that the transcription of all three known compatible solute uptake systems (opuC,betL, andgbu), as well as a gene that is predicted to encode a compatible solute transporter subunit (lmo1421) is induced in response to elevated osmolarity. The osmotic induction ofopuCAandlmo1421 was found to be strongly σ^Bdependent. Together these observations suggest that σ^Bplays a major role in the regulation of carnitine utilization byL. monocytogenesbut is not essential for betaine utilization by this pathogen.