Analysis of transcription of the bph locus of Burkholderia sp strain LB400 and evidence that the ORF0 gene product acts as a regulator of the bphA1 promoter

Although gene clusters for the degradation of biphenyls and polychlorobiphenyls have been extensively characterized, comparatively little is known about the regulation of their expression. In the present work, different aspects of transcription of the bph locus of the potent polychlorobiphenyl degrader Burkholderia sp. strain LB400 were investigated. An RNA blot analysis of the entire gene cluster revealed that the transcription of all genes encoding biphenyl catabolic enzymes responded similarly to the presence of biphenyl, succinate or a mixture of the two. One region of the locus, encompassing ORF0, was separately transcribed and differently regulated. A single start position was mapped for this monocistronic transcript. Synthesis of the adjacent RNA, encoding subunits of biphenyl dioxygenase, was strongly biphenyl-inducible. In this case, four major 5 ' -ends were mapped between 25 and 70 bp upstream of the start codon of gene bphA1. Sequence elements between approximately positions 710 and 1080 upstream were required in cis for full functioning of the respective promoter(s) (P-bphA1). ORF0(-) mutants of strain LB400 retained the ability to grow on biphenyl, but showed decreased concentrations of bphA1A2 RNA and decreased lacZ expression in strains harbouring a reporter system with a bphA1-lacZ transcriptional fusion. This effect was compensated by the introduction of an intact ORF0 in trans, indicating that the ORF0 gene product mediates activation of P-bphA1.