Phenotypic and physiological alterations by heterologous acylhomoserine lactone synthase expression in Pseudomonas putida

Abstract

Many bacteria harbour an incomplete quorum-sensing (QS) system, whereby they possess LuxR homologues without the QS acylhomoserine lactone (AHL) synthase, which is encoded by a luxI homologue. An artificial AHL-producing plasmid was constructed using a cviI gene encoding the C6-AHL [N-hexanoyl homoserine lactone (HHL)] synthase from Chromobacterium violaceum, and was introduced successfully into both the wild-type and a ppoR (luxR homologue) mutant of Pseudomonas putida. Our data provide evidence to suggest that the PpoR-HHL complex, but neither PpoR nor HHL alone, could attenuate growth, antibiotic resistance and biofilm formation ability. In contrast, swimming motility, siderophore production and indole degradation were enhanced by PpoR-HHL. The addition of exogenous indole increased biofilm formation and reduced swimming motility. Interestingly, indole proved ineffective in the presence of PpoR-HHL, thereby suggesting that the PpoR-HHL complex masks the effects of indole. Our data were supported by transcriptome analyses, which showed that the presence of the plasmid-encoded AHL synthase altered the expression of many genes on the chromosome in strain KT2440. Our results showed that heterologous luxI expression that occurs via horizontal gene transfer can regulate a broad range of specific target genes, resulting in alterations of the phenotype and physiology of host cells.

Indole toxicity involves the inhibition of adenosine triphosphate production and protein folding inPseudomonas putida

“…In our previous study, the addition of indole promoted biofilm formation in KT2440 cells (Lee et al ., ). Furthermore, others have shown that indole increases biofilm formation in other pseudomonads including P. aeruginosa and P. fluorescens (Lee et al ., ).…”

“…Furthermore, others have shown that indole increases biofilm formation in other pseudomonads including P. aeruginosa and P. fluorescens (Lee et al ., ). In contrast, biofilm formation and cell adhesion decreased in E. coli upon treatment with indole (Domka et al ., ; Bansal et al ., ; Lee & Lee, ; Lee et al ., ), suggesting that indole acts differently in different bacterial species. Biofilm formation in KT2440 cells increased in the presence of indole in both rich and minimal media (Fig.…”

“…The E. coli LuxR homolog, SdiA, is considered to be related to this indole signaling (Lee et al ., ). In our previous study, the addition of exogenous indole increased the expression of the ppoR gene, an sdiA homolog, in Pseudomonas putida (Lee et al ., ). Furthermore, increased biofilm formation and reduced swimming motility were also observed upon treatment with indole, but these effects were inhibited by the expression of the cviI gene (encoding N ‐hexanoyl homoserine lactone synthase, HHL) in P. putida .…”

Role of Glyoxylate Shunt in Oxidative Stress Response

“…Total RNA was isolated using the RNeasy Mini kit (Qiagen). Specific steps for further RNA isolation were conducted as previously described (20). cDNA probes for cDNA microarray analysis were prepared by the reverse transcription of total RNA (50 g) in the presence of aminoallyl-dUTP and 6 g of random primers (Invitrogen) for 3 h. The cDNA probes were cleaned up using a Microcon YM-30 column (Millipore) followed by coupling to the Cy3 dye (for the reference) or Cy5 dye (for the test samples) (Amersham Biosciences).…”

Indole-Induced Activities of β-Lactamase and Efflux Pump Confer Ampicillin Resistance in Pseudomonas putida KT2440

“…P. putida KT2440 and P. aeruginosa strains were grown at 30°C and 37°C in Luria-Bertani (LB) and modified M9 media [Na 2 HPO 4 7H 2 O (6.8 g/l), KH 2 PO 4 (3 g/l), NaCl (0.5 g/l), NH 4 Cl (1 g/l), MgSO 4 (2 mM), and CaCl 2 (0.1 mM)] (Sambrook et al, 1989) containing 10 mM glucose and 10 mM succinate (Lee Y. et al, 2010) with aeration and shaking. Reagents in media were purchased from Sigma (USA).…”