Function and regulation of pob genes for 4-hydroxybenzoate catabolism in Agrobacterium tumefaciens

Agrobacterium tumefaciens is a pathogen that causes tumors in plants. Phenolic acids present in the soil and rhizosphere may affect the interaction between A. tumefaciens and plants. An important pathway for microorganisms to degrade phenolic acids is the β-ketoadipate pathway, which has been annotated in the genome of A. tumefaciens. The ability of the PobA (atu4544) enzyme to catalyze the conversion of 4-hydroxybenzoate to protocatechuate was essential for cell growth using 4-hydroxybenzoate as the sole carbon source. The pobA gene is located upstream of atu4545, encoding an AraC transcription factor (PobR). Strains with deleted or supplemented atu4545 exhibited similar growth characteristics on common and phenolic acid-containing carbon sources as strains with deleted or supplemented atu4544. Strains with a pobA::lacZ reporter fusion showed that PobR induced pobA expression. In addition, the use of a pobR::lacZ reporter fusion showed that PobR represses its expression. Electromobility shift assay revealed that the PobR regulator can bind specifically to DNA. The binding site was identified as CGTGCGATGGTGGATT. Deletions of atu4544 (pobA) and atu4545 (pobR) decreased A. tumefaciens pathogenicity by infecting carrot roots and kalanchoe leaves, with no effect on virB genes, and decreased bacterial biomass when phenolic acids were present. The collective findings demonstrate how transcriptional regulation by A. tumefaciens controls the metabolism of 4-hydroxybenzoate and imply that PobA and PobR aid in bacterial survival during host plant infection.IMPORTANCEAgrobacterium tumefaciens is a widely distributed environmental bacterium and a recognized phytopathogen. Phenolic acids influence the relationship between A. tumefaciens and plants. One of the most important phenolic acids found in soil is 4-hydroxybenzoate, which is generated by plants. Mutants defective in the atu4544 and atu4545 genes inhibit A. tumefaciens tumor development. The atu4544-encoded enzyme, PobA, can metabolize 4-hydroxybenzoate, and the expression of its gene is positively regulated by the transcription factor encoded by atu4545. The atu4545 gene is subject to negative autoregulation. The binding site of atu4545 is CGTGCGATGGTCGGATT. Dual regulation of regulators for phenolic acid catabolism may aid in the maintenance of appropriate quantities of phenolic compounds. These results clarify the pathogenic mechanisms of A. tumefaciens and broaden the understanding of the metabolic control mechanisms of phenolic chemicals.