The Vibrio cholerae VprA-VprB Two-Component System Controls Virulence through Endotoxin Modification

Abstract

The bacterial cell surface is the first structure the host immune system targets to prevent infection. Cationic antimicrobial peptides of the innate immune system bind to the membrane of Gram-negative pathogens via conserved, surface-exposed lipopolysaccharide (LPS) molecules. We recently reported that modern strains of the global intestinal pathogen Vibrio cholerae modify the anionic lipid A domain of LPS with a novel moiety, amino acids. Remarkably, glycine or diglycine addition to lipid A alters the surface charge of the bacteria to help evade the cationic antimicrobial peptide polymyxin. However, the regulatory mechanisms of lipid A modification in V. cholerae are unknown. Here, we identify a novel two-component system that regulates lipid A glycine modification by responding to important biological cues associated with pathogenesis, including bile, mildly acidic pH, and cationic antimicrobial peptides. The histidine kinase Vc1319 (VprB) and the response regulator Vc1320 (VprA) respond to these signals and are required for the expression of the almEFG operon that encodes the genes essential for glycine modification of lipid A. Importantly, both the newly identified two-component system and the lipid A modification machinery are required for colonization of the mammalian host. This study demonstrates how V. cholerae uses a previously unknown regulatory network, independent of well-studied V. cholerae virulence factors and regulators, to respond to the host environment and cause infection.

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

“…Once across the outer membrane, CAMPs can disrupt the cytoplasmic membrane and/or inhibit critical cytoplasmic processes, resulting in cell death. V. cholerae resistance to CAMPs is mediated by active efflux via the VexAB-TolC RND-efflux system (22), expression of the extracytoplasmic stress response (17), and covalent modification of LPS (24)(25)(26)(27). LPS remodeling has been shown to be critical for high-level CAMP resistance.…”

“…In V. cholerae the production of hexa-acylated lipid A via the MsbB acyltransferase confers resistance to polymyxin B (PXB), a CAMP antibiotic (28). Hexacylated lipid A can be further modified with glycine and diglycine residues by AlmEFG to effect high-level PXB resistance (25)(26)(27). Glycinylation of lipid A results in a net increase in the positive charge of lipid A which reduces the electrostatic interactions between CAMPs and lipid A, leading to CAMP resistance.…”

“…cholerae LPS glycinylation is regulated by CarRS (also known as VprAB), a two-component regulatory system (24,25). CarRS was first identified as a calcium-responsive negative regulator of biofilm production (29).…”

“…CarRS was first identified as a calcium-responsive negative regulator of biofilm production (29). Two subsequent studies showed that CarRS also regulated CAMP resistance by positively regulating almEFG expression (24,25). Mutations in carR or almEFG resulted in an ϳ100-fold decrease in V. cholerae PXB resistance and attenuated intestinal colonization (24,25), highlighting the importance of lipid A modification in V. cholerae.…”

“…The molecular mechanisms controlling carRS expression are unknown. Previous studies showed that carRS expression was influenced by environmental cues, with carRS expression being activated by PXB and repressed by bile salts and calcium (25,29).…”