In Helicobacter pylori auto-inducer-2, but not LuxS/MccAB catalysed reverse transsulphuration, regulates motility through modulation of flagellar gene transcription.
BACKGROUND
LuxS 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.


RESULTS
We 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 DeltaluxSHp 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 DeltaluxSHp 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 DeltaluxSHp mutant. Immunoblotting identified decreased levels of FlaA and FlgE but not FlaB in the DeltaluxSHp 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 DeltaluxSHp mutant restored flagellar gene transcription, and the number and length of flagella.


CONCLUSIONS
Our 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.
Shen
Feifei
F
Centre for Biomolecular Sciences, University of Nottingham, University Park, Nottingham NG7 2RD, UK. fayeshen316@msn.com
Hobley
Laura
L
Doherty
Neil
N
Loh
John T
JT
Cover
Timothy L
TL
Sockett
R Elizabeth
RE
Hardie
Kim R
KR
Atherton
John C
JC
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D013437:Carbon-Sulfur Lyases / Q000235:genetics / Q000378:metabolism*; D003545:Cysteine / Q000378:metabolism*; D005407:Flagella / Q000235:genetics* / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D016480:Helicobacter pylori / Q000235:genetics / Q000502:physiology*; D006714:Homoserine / Q000031:analogs & derivatives* / Q000378:metabolism; D007783:Lactones / Q000378:metabolism*; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D007783:Lactones; C091088:N-octanoylhomoserine lactone; D006714:Homoserine; D013437:Carbon-Sulfur Lyases; C117753:LuxS protein, Bacteria; D003545:Cysteine