The toluene-3-monooxygenase from Burkholderia pickettii PKO1 and the toluene/benzene-2-monooxygenase from Burkholderia (Pseudomonas) sp. strain JS150 are distinct enzymes which differ not only in catalytic specificity and substrate range but also in the arrangement and sequence of the genes within the operons that encode the enzymes, tbuA1UBVA2C and tbmABCDEF, respectively. In the present study, we examined the transcriptional activation of the PtbuA1 and PtbmA promoters by their cognate regulators, TbuT and TbmR. TbmR and TbuT each exhibited activation of both PtbmA and PtbuA1, with toluene, benzene, and chlorobenzene serving as strong effectors. These results strongly suggest that TbmR is an NtrC-like regulator which is functionally homologous to TbuT, and they provide evidence for the evolutionary "recruitment" of the same or a similar type of regulator for both monooxygenase pathways.