Journal of Bacteriology, November 2000, p. 6401-6411, Vol. 182, No. 22
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
School of Pharmaceutical Sciences1 and Institute of Infections and Immunity,2 University of Nottingham, University Park, Nottingham NG7 2RD, United Kingdom, and Faculty of Life Sciences, Bar-Ilan University, Ramat-Gan 52 900, Israel3
Received 15 May 2000/Accepted 5 September 2000
In Pseudomonas aeruginosa, many exoproduct virulence
determinants are regulated via a hierarchical quorum-sensing cascade involving the transcriptional regulators LasR and RhlR and their cognate activators,
N-(3-oxododecanoyl)-L-homoserine lactone
(3O-C12-HSL) and N-butanoyl-L-homoserine
lactone (C4-HSL). In this paper, we demonstrate that the cytotoxic
lectins PA-IL and PA-IIL are regulated via quorum sensing. Using
immunoblot analysis, the production of both lectins was found to be
directly dependent on the rhl locus while, in a
lasR mutant, the onset of lectin synthesis was delayed but
not abolished. The PA-IL structural gene, lecA, was cloned
and sequenced. Transcript analysis indicated a monocistronic organization with a transcriptional start site 70 bp upstream of the
lecA translational start codon. A lux box-type
element together with RpoS (
S) consensus sequences was
identified upstream of the putative promoter region. In
Escherichia coli, expression of a
lecA::lux reporter fusion was
activated by RhlR/C4-HSL, but not by LasR/3O-C12-HSL, confirming direct
regulation by RhlR/C4-HSL. Similarly, in P. aeruginosa
PAO1, the expression of a chromosomal
lecA::lux fusion was enhanced but not
advanced by the addition of exogenous C4-HSL but not 3O-C12-HSL.
Furthermore, mutation of rpoS abolished lectin synthesis in
P. aeruginosa, demonstrating that both RpoS and
RhlR/C4-HSL are required. Although the C4-HSL-dependent expression
of the lecA::lux reporter in E. coli could be inhibited by the presence of 3O-C12-HSL, this did
not occur in P. aeruginosa. This suggests that, in the
homologous genetic background, 3O-C12-HSL does not function as a
posttranslational regulator of the RhlR/C4-HSL-dependent activation of
lecA expression.
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