Quorum sensing in Vibrio fischeri: elements of the luxl promoter.
Although cell density-dependent regulation of the luminescence genes in Vibrio fischeri is a model for quorum sensing in Gram-negative bacteria, relatively little is known about the promoter of the luminescence operon. The luminescence operon is activated by the LuxR protein, which requires a diffusible acylhomoserine lactone signal. The lux box, a 20 bp inverted repeat, is located in the luxl promoter region and is required for LuxR-dependent induction of the luminescence genes. Using primer extension, we mapped the LuxR-dependent transcriptional start site of the lux operon to 19 bp upstream of the luxl start codon. This indicates that the lux box is centred at -42.5 bp from the start of transcription. To gain evidence about the location of the -10 sequence, we placed a consensus -35 hexamer at different locations relative to the luxl transcriptional start site and measured constitutive levels of luminescence in recombinant Escherichia coli. The strongest constitutive promoter contained a TATAGT hexamer 17 bp from the -35 consensus sequence and 6 bp from the transcriptional start site. We propose that this is the -10 hexamer. Also in recombinant E. coli, both half-sites of the lux box were required for LuxR-dependent gene activation and for activation by an autoinducer-independent, monomeric LuxR deletion protein. LuxR-dependent activation of luminescence was eliminated when the lux box was centred at -47.5, -52.5 and -62.5 with respect to the luxl transcriptional start site. Our evidence, taken together with other information, points to a model in which a LuxR dimer overlaps the -35 region of the luxl promoter and functions as an ambidextrous activator with each LuxR subunit interacting with a different region of RNA polymerase.
Egland
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KA
Department of Microbiology, University of Iowa, Iowa City 52242, USA.
Greenberg
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EP
D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism*; D001483:Base Sequence; D017931:DNA Primers; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D008163:Luminescent Measurements; D008969:Molecular Sequence Data; D009876:Operon; D010957:Plasmids; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000032:analysis / Q000378:metabolism; D011995:Recombination, Genetic; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014733:Vibrio / Q000235:genetics / Q000254:growth & development / Q000502:physiology*
D001426:Bacterial Proteins; D017931:DNA Primers; D012333:RNA, Messenger; C084289:luxL protein, Vibrio harveyi; D012321:DNA-Directed RNA Polymerases