Vibrio cholerae LeuO Links the ToxR Regulon to Expression of Lipid A Remodeling Genes

Abstract

Vibrio cholerae is an intestinal pathogen that causes the diarrheal disease cholera. Colonization of the intestine depends upon the expression of genes that allow V. cholerae to overcome host barriers, including low pH, bile acids, and the innate immune system. ToxR is a major contributor to this process. ToxR is a membrane-spanning transcription factor that coordinates gene expression in response to environmental cues. In previous work we showed that ToxR upregulated leuO expression in response to bile salts. LeuO is a LysR family transcription factor that contributes to acid tolerance, bile resistance, and biofilm formation in V. cholerae. Here, we investigated the function of ToxR and LeuO in cationic antimicrobial peptide (CAMP) resistance. We report that ToxR and LeuO contribute to CAMP resistance by regulating carRS transcription. CarRS is a two-component regulatory system that positively regulates almEFG expression. AlmEFG confers CAMP resistance by glycinylation of lipid A. We found that the expression of carRS and almEFG and the polymyxin B MIC increased in mutants lacking toxRS or leuO. Conversely, leuO overexpression decreased the polymyxin B MIC. Furthermore, we found that LeuO directly bound to the carRS promoter and that ToxR-dependent activation of leuO transcription regulated carRS transcription in response to bile salts. Our results suggest that LeuO functions downstream of ToxR to modulate carRS expression in response to environmental cues. This study extends the functional role of ToxR and LeuO in environmental adaptation to include cell surface remodeling and CAMP resistance. Vibrio cholerae is a Gram-negative facultative human pathogen and the causative agent of the diarrheal disease cholera. V. cholerae is native to aquatic ecosystems, where it often associates with chitinous aquatic organisms (reviewed in reference 1). People acquire V. cholerae by ingestion of contaminated food or water. Following ingestion, V. cholerae colonizes the small intestine, where it produces virulence factors that result in a severe secretory diarrhea that is the hallmark of the disease cholera. The secretory diarrhea then contributes to V. cholerae transmission to new hosts and dissemination into the aquatic ecosystem.The ability of V. cholerae to rapidly transition between the aquatic ecosystem and the host is essential for its success as a pathogen. Colonization of the human gastrointestinal tract is mediated by transcriptional responses that facilitate adaptation to dynamic environments in the gastrointestinal tract. Following ingestion, V. cholerae activates the expression of virulence factors that are essential for both colonization and disease development. This includes the production of the enterotoxin cholera toxin (CT) and an adhesin called the toxin coregulated pilus (TCP) (2, 3). The expression of many V. cholerae virulence genes is coordinately regulated by the ToxR regulon (4). The ToxR regulon is divided into two branches: the ToxT-dependent branch and the ToxT-independent branch. In the ToxT-depe...

Acidic pH promotes lipopolysaccharide modification and alters colonization in a bacteria–animal mutualism

“…Unlike V. fischeri, V. cholerae encodes an additional LPS-modifying system that confers PMB resistance: the aminoacyl lipid modifying enzymes AlmEFG. In contrast to the acid-induced modification described here, transcription of almEFG is repressed by exposure to acid via the LeuO-ToxRS-CarRS/VprAB cascade of transcription factors (Herrera et al, 2014;Ante et al, 2015;Bilecen et al, 2015;Bina et al, 2016). The V. cholerae PMB resistance phenotype is also regulated by the envelope stress response through the alternative sigma factor RpoE and the outer membrane protein OmpU (Mathur et al, 2007;Shaw, 2012).…”

Novel coordination of lipopolysaccharide modifications in Vibrio cholerae promotes CAMP resistance

“…). Recently, Bina and co‐workers reported that ToxR controls VprA in a LeuO‐dependent manner (Bina et al ., ). Both LeuO and ToxR are broad regulators of Vibrio pathogenicity (Shimada et al ., ; Kazi et al ., ) and deletion of either increases polymyxin resistance 3‐fold in mid‐log‐phase cells.…”

Control of virulence gene transcription by indirect readout in Vibrio cholerae and Salmonella enterica serovar Typhimurium

“…Among the genes that are depressed by ToxR in V. cholerae is leuO , encoding the LysR‐like wHTH DNA binding protein LeuO (Bina et al ., ). LysR‐type transcription factors (LTTRs) have very weak consensus DNA binding site sequences: T‐N 11 ‐A, where N is any base.…”