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The human pathogenic species of Yersinia all synthesise AHLs and orthologues of LuxI (the AHL synthase) and LuxR (the response regulator) of Vibrio fischeri have been cloned and sequenced from Y. pseudotuberculosis (ypsR/I, ytbR/I), Y. enterocolitica (yenR/I) and Y. pestis (ypeR/I, yepR/I) . Previously we have shown that Y. pseudotuberculosis synthesises three short chain AHLs, N-3-(oxo-hexanoyl)homoserine lactone (3-oxo-C6-HSL), N-hexanoylhomoserine lactone (C6-HSL) and N-octanoylhomoserine lactone (C8-HSL) . Using liquid chromatography (LC) coupled to hybrid quadruple-linear ion trap mass spectrometry we have recently revealed that Y. pseudotuberculosis actually produces at least twenty four different AHLs, eight of which have been detected at levels which are considered to be biologically significant. Both YpsI and YtbI are capable of synthesising C6-HSL, 3-oxo-C6-HSL and N-3-(oxo-septanoyl)homoserine lactone (3-oxo-C7-HSL) while YtbI is solely responsible for the synthesis of N-3-(hydroxy-octanonoyl)homoserine lactone (3-OH-C8-HSL), N-3-(oxo-octanoyl)homoserine lactone (3-oxo-C8-HSL), C8-HSL and N-3-(oxo-decanoyl)homoserine lactone (3-oxo-C10-HSL). Both the YpsI and YtbI synthases are required in combination for the synthesis of N-septanoylhomoserine lactone (C7-HSL) . During growth in LB medium at 28oC or 37oC, Y. pseudotuberculosis ypsR mutant cells aggregate, a phenomenon which does not occur at 22oC or in either the parent or YpIII ypsI mutant at any temperature. This phenotype implied the involvement of surface components which were likely to be regulated via AHL-dependent QS. SDS-PAGE of cell surface extracts prepared from parent, ypsI and ypsR mutants (grown at 28oC) revealed that the expression of numerous proteins was affected including an upregulated 42 kDa protein with significant homology to the major flagellin structural proteins FleA, FleB and FleC of Y. enterocolitica . These data implicated ypsR in the control of flagellar-mediated motility. On soft-agar motility plate assays, both ypsR and ypsI mutants, but not the parent Y. pseudotuberculosis strain, were motile after 24h when grown at 22oC. None of the strains were motile after 24 h or 48 h at 37oC (Fig 1). By following the onset of motility over the growth curve in batch culture, it was clear that at 22oC, motility is induced after 6 h in both ypsI and ypsR mutants but delayed until 26 h in the parent. Thus cell population density appears to be one important environmental cue for activating swimming motility in Y. pseudotuberculosis. Furthermore, although the ypsRI locus is involved in the repression of motility, mutation of this locus does not overcome the temperature dependence of swimming motility.
