Transcriptional kinetics of the cross-talk between the ortho-cleavage and TOL pathways of toluene biodegradation in Pseudomonas putida mt-2

Abstract

The TOL plasmid promoters are activated by toluene leading to gene expression responsible for the degradation of the environmental signal. Benzoate is formed as an intermediate, activating the BenR protein of the chromosomal ortho-cleavage pathway that up-regulates the chromosomal PbenA promoter and the TOL Pm promoter resulting in cross-talk between the two networks. Herein, the transcriptional kinetics of the PbenR and PbenA promoters in conjunction with TOL promoters was monitored by real-time PCR during toluene biodegradation of different concentrations in batch cultures. The cross-talk between the two pathways was indicated by the simultaneous maximal expression of the Pm and PbenR promoters, as well as the transcriptional activation from PbenA occurring prior to PbenR, which indicates the potential up-regulation of PbenA by the TOL XylS protein. The repressory effect of toluene on Pr was evident for concentrations higher than 0.3 mM suggesting a threshold value for restoring the promoter's activity, while all the other promoters followed a specific expression pattern, regardless of the initial inducer concentration. Induction of the system with higher toluene concentrations revealed an oscillatory behaviour of Pm, the expression of which remained at high levels until the late exponential phase, demonstrating a novel function of this network.