BenR, a XylS homologue, regulates three different pathways of aromatic acid degradation in Pseudomonas putida

Pseudomonas putida converts benzoate to catechol using two enzymes that are encoded on the chromosome and whose expression is induced by benzoate. Ben zoate also binds to the regulator XylS to induce expression of the TOL (tol uene degradation) plasmid-encoded meta pathway operon for benzoate and meth ylbenzoate degradation. Finally, benzoate represses the ability of P. putid a to transport 4-hydroxybenzoate (4-HBA) by preventing transcription of pca K, the gene encoding the 4-HBA permease. Here we identified a gene, benR, a s a regulator of benzoate, methylbenzoate, and 4-MBA degradation genes. A b enR mutant isolated by random transposon mutagenesis was unable to grow on benzoate. The deduced amino acid sequence of BenR showed high similarity (6 2% identity) to the sequence of XylS, a member of the AraC family of regula tors. An additional seven genes located adjacent to berta were inferred to be involved in benzoate degradation based on their deduced amino acid seque nces. The benABC genes likely encode benzoate dioxygenase, and BenD likely encodes 2-hydro-1,2-dihydroxybenzoate dehydrogenase. benK and benF were ass igned functions as a benzoate permease and porin, respectively. The possibl e function of a final gene, benE, is not known. benR activated expression o f a benA-lacZ reporter fusion in response to benzoate. It also activated ex pression of a meta cleavage operon promoter-lacZ fusion inserted in an E. c oli chromosome. Third, benR was required for benzoate-mediated repression o f pcaK-lacZ fusion expression. The benA promoter region contains a direct r epeat sequence that matches the XylS binding site previously defined for th e meta cleavage operon promoter. It is likely that BenR binds to the promot er region of chromosomal benzoate degradation genes and plasmid-encoded met hylbenzoate degradation genes to activate gene expression in response to be nzoate. The action of BenR in repressing 4-MBA uptake is probably indirect.