The logic layout of the TOL network of Pseudomonas putida pWW0 plasmid stems from a metabolic amplifier motif (MAM) that optimizes biodegradation of m-xylene.

BACKGROUND

The genetic network of the TOL plasmid pWW0 of the soil bacterium Pseudomonas putida mt-2 for catabolism of m-xylene is an archetypal model for environmental biodegradation of aromatic pollutants. Although nearly every metabolic and transcriptional component of this regulatory system is known to an extraordinary molecular detail, the complexity of its architecture is still perplexing. To gain an insight into the inner layout of this network a logic model of the TOL system was implemented, simulated and experimentally validated. This analysis made sense of the specific regulatory topology out on the basis of an unprecedented network motif around which the entire genetic circuit for m-xylene catabolism gravitates.

RESULTS

The most salient feature of the whole TOL regulatory network is the control exerted by two distinct but still intertwined regulators (XylR and XylS) on expression of two separated catabolic operons (upper and lower) for catabolism of m-xylene. Following model reduction, a minimal modular circuit composed by five basic variables appeared to suffice for fully describing the operation of the entire system. In silico simulation of the effect of various perturbations were compared with experimental data in which specific portions of the network were activated with selected inducers: m-xylene, o-xylene, 3-methylbenzylalcohol and 3-methylbenzoate. The results accredited the ability of the model to faithfully describe network dynamics. This analysis revealed that the entire regulatory structure of the TOL system enables the action an unprecedented metabolic amplifier motif (MAM). This motif synchronizes expression of the upper and lower portions of a very long metabolic system when cells face the head pathway substrate, m-xylene.

CONCLUSION

Logic modeling of the TOL circuit accounted for the intricate regulatory topology of this otherwise simple metabolic device. The found MAM appears to ensure a simultaneous expression of the upper and lower segments of the m-xylene catabolic route that would be difficult to bring about with a standard substrate-responsive single promoter. Furthermore, it is plausible that the MAM helps to avoid biochemical conflicts between competing plasmid-encoded and chromosomally-encoded pathways in this bacterium.