Swarming-coupled expression of the Proteus mirabilis hpmBA haemolysin operon.
The HpmA haemolysin toxin of Proteus mirabilis is encoded by the hpmBA locus and its production is upregulated co-ordinately with the synthesis and assembly of flagella during differentiation into hyperflagellated swarm cells. Primer extension identified a sigma(70) promoter upstream of hpmB that was upregulated during swarming. Northern blotting indicated that this promoter region was also required for concomitant transcription of the immediately distal hpmA gene, and that the unstable hpmBA transcript generated a stable hpmA mRNA and an unstable hpmB mRNA. Transcriptional luxAB fusions to the DNA regions 5' of the hpmB and hpmA genes confirmed that hpmB sigma(70) promoter activity increased in swarm cells, and that there was no independent hpmA promoter. Increased transcription of the hpmBA operon in swarm cells was dependent upon a 125 bp sequence 5' of the sigma(70) promoter -35 hexamer. This sequence spans multiple putative binding sites for the leucine-responsive regulatory protein (Lrp), and band-shift assays with purified Lrp confirmed the presence of at least two such sites. The influence on hpmBA expression of the key swarming positive regulators FlhD(2)C(2) (encoded by the flagellar master operon), Lrp, and the membrane-located upregulator of the master operon, UmoB, was examined. Overexpression of each of these regulators moderately increased hpmBA transcription in wild-type P. mirabilis, and the hpmBA operon was not expressed in any of the flhDC, lrp or umoB mutants. Expression in the mutants was not recovered by cross-complementation, i.e. by overexpression of FlhD(2)C(2), Lrp or UmoB. Expression of the zapA protease virulence gene, which like hpmBA is also upregulated in swarm cells, did not require Lrp, but like flhDC it was upregulated by UmoB. The results indicate intersecting pathways of control linking virulence gene expression and swarm cell differentiation.
Fraser
Gillian M
GM
Cambridge University Department of Pathology, Tennis Court Road, Cambridge CB2 1QP, UK1.
Claret
Laurent
L
Cambridge University Department of Pathology, Tennis Court Road, Cambridge CB2 1QP, UK1.
Furness
Richard
R
Cambridge University Department of Pathology, Tennis Court Road, Cambridge CB2 1QP, UK1.
Gupta
Srishti
S
Cambridge University Department of Pathology, Tennis Court Road, Cambridge CB2 1QP, UK1.
Hughes
Colin
C
Cambridge University Department of Pathology, Tennis Court Road, Cambridge CB2 1QP, UK1.
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D012321:DNA-Directed RNA Polymerases; D005407:Flagella / Q000502:physiology; D015964:Gene Expression Regulation, Bacterial; D006460:Hemolysin Proteins / Q000235:genetics / Q000378:metabolism*; D007930:Leucine; D008565:Membrane Proteins; D008969:Molecular Sequence Data; D009876:Operon; D011401:Promoter Regions, Genetic; D011513:Proteus mirabilis / Q000235:genetics / Q000472:pathogenicity / Q000502:physiology*; D017422:Sequence Analysis, DNA; D012808:Sigma Factor; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D006460:Hemolysin Proteins; C064194:HpmA hemolysin; C104106:HpmB protein, Proteus mirabilis; D008565:Membrane Proteins; D012808:Sigma Factor; C081775:RNA polymerase sigma 70; D012321:DNA-Directed RNA Polymerases; D007930:Leucine