The Pseudomonas aeruginosa lectins PA-IL and PA-IIL are controlled by quorum sensing and by RpoS.
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 (sigma(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.
Winzer
K
K
School of Pharmaceutical Sciences, University of Nottingham, University Park, Nottingham NG7 2RD, United Kingdom.
Falconer
C
C
Garber
N C
NC
Diggle
S P
SP
Camara
M
M
Williams
P
P
D018829:Adhesins, Bacterial / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D018398:Homeodomain Proteins / Q000235:genetics; D007783:Lactones / Q000494:pharmacology; D037102:Lectins / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D011550:Pseudomonas aeruginosa / Q000235:genetics* / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics*; D012808:Sigma Factor / Q000235:genetics*; D015534:Trans-Activators / Q000235:genetics; D014158:Transcription, Genetic
D018829:Adhesins, Bacterial; D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; D007783:Lactones; C080365:LasR protein, Pseudomonas aeruginosa; C102929:LecA protein, bacteria; D037102:Lectins; D012097:Repressor Proteins; D012808:Sigma Factor; D015534:Trans-Activators; C069567:sigma factor KatF protein, Bacteria