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Proximal and distal sites bind LuxR independently and activate expression of the Vibrio harveyi lux operon
Author(s) -
Miyamoto Carol M.,
Smith Eric E.,
Swartzman Eiana,
Cao JieGang,
Graham Angus F.,
Meighen Edward A.
Publication year - 1994
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1111/j.1365-2958.1994.tb01286.x
Subject(s) - vibrio harveyi , operon , biology , autoinducer , binding site , promoter , transcription (linguistics) , repressor , gene , microbiology and biotechnology , lac operon , genetics , transcription factor , plasmid , gene expression , escherichia coli , quorum sensing , vibrio , virulence , bacteria , linguistics , philosophy
Summary The LuxR regulatory protein of Vibrio harveyi as well as the autoinducer molecule, N‐(3‐hydroxybutanoyl) homoserine lactone, are known to be required for expression of luminescence. Although LuxR has been implicated in the activation of the promoter of the lux operon of V. harveyi , and can bind to two distinct sites upstream of the transcription initiation start site, its mode of action is unknown, in the present experiments, mobility shift assays were used to demonstrate that LuxR bound to the distal and proximal sites in an independent rather than co‐operative interaction with a much tighter binding to the distal site. Deletion and mutation analyses of DNA upstream of the lux promoter followed by transconjugation Into V. harveyi in trans using the chloramphenicol acetyl‐transferase ( cat ) gene as a reporter demonstrated, however, that the proximal site for LuxR was absolutely critical for promoter activation while the distal LuxR site was only necessary for maximum activation. This result was confirmed by mutation of the proximal site which blocked activation of the lux promoter and binding of LuxR to this site, but did not prevent LuxR binding to the distal site.