Abstract
BackgroundLuxS may function as a metabolic enzyme or as the synthase of a quorum sensing signalling molecule, auto-inducer-2 (AI-2); hence, the mechanism underlying phenotypic changes upon luxS inactivation is not always clear. In Helicobacter pylori, we have recently shown that, rather than functioning in recycling methionine as in most bacteria, LuxS (along with newly-characterised MccA and MccB), synthesises cysteine via reverse transsulphuration. In this study, we investigated whether and how LuxS controls motility of H. pylori, specifically if it has its effects via luxS-required cysteine metabolism or via AI-2 synthesis only.ResultsWe report that disruption of luxS renders H. pylori non-motile in soft agar and by microscopy, whereas disruption of mccAHp or mccBHp (other genes in the cysteine provision pathway) does not, implying that the lost phenotype is not due to disrupted cysteine provision. The motility defect of the ΔluxSHp mutant was complemented genetically by luxSHp and also by addition of in vitro synthesised AI-2 or 4, 5-dihydroxy-2, 3-pentanedione (DPD, the precursor of AI-2). In contrast, exogenously added cysteine could not restore motility to the ΔluxSHp mutant, confirming that AI-2 synthesis, but not the metabolic effect of LuxS was important. Microscopy showed reduced number and length of flagella in the ΔluxSHp mutant. Immunoblotting identified decreased levels of FlaA and FlgE but not FlaB in the ΔluxSHp mutant, and RT-PCR showed that the expression of flaA, flgE, motA, motB, flhA and fliI but not flaB was reduced. Addition of DPD but not cysteine to the ΔluxSHp mutant restored flagellar gene transcription, and the number and length of flagella.ConclusionsOur data show that as well as being a metabolic enzyme, H. pylori LuxS has an alternative role in regulation of motility by modulating flagellar transcripts and flagellar biosynthesis through production of the signalling molecule AI-2.
Highlights
LuxS may function as a metabolic enzyme or as the synthase of a quorum sensing signalling molecule, auto-inducer-2 (AI-2); the mechanism underlying phenotypic changes upon luxS inactivation is not always clear
The H. pylori ΔluxS mutant lost the ability to produce AI-2 while the wild-type, ΔmccAHp and ΔmccBHp mutants did not Our previous study has demonstrated that luxSHp, mccAHp and mccBHp genes comprise a reverse transulphuration pathway in H. pylori, which is the sole cysteine biosynthesis pathway [15]
We wanted to determine whether these mutants in a motile strain of H. pylori, J99, would be useful in differentiating whether H. pylori motility was affected by luxS associated AI-2 production or by cysteine provision
Summary
LuxS may function as a metabolic enzyme or as the synthase of a quorum sensing signalling molecule, auto-inducer-2 (AI-2); the mechanism underlying phenotypic changes upon luxS inactivation is not always clear. It is generally assumed that auto-inducers are employed to regulate aspects of bacterial behaviour in response to cell population density (so-called quorum sensing). This includes changes in the expression of genes crucial for bacterial survival or virulence [1,2]. Many Gram-positive and Gram-negative bacterial species possess LuxS, and in some it has been shown to catalyse AI-2 production and to control quorum sensing (QS). Good examples include Vibrio harveyi and Vibrio cholera, where AI-2 has been shown to regulate densitydependent bioluminescence and virulence factor production, respectively [4,5]. luxS inactivation has been shown to cause phenotypic alterations such as biofilm formation, changes in motility, toxin production, and reduced colonisation in various experimental infection models [3,6]
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