Study of factors which negatively affect expression of the phenol degradation operon pheBA in Pseudomonas putida.
Transcription of the plasmid-borne phenol catabolic operon pheBA in Pseudomonas putida is activated by the LysR-family regulator CatR in the presence of the effector molecule cis,cis-muconate (CCM), which is an intermediate of the phenol degradation pathway. In addition to the positive control of the operon, several factors negatively affect transcription initiation from the pheBA promoter. First, the activation of the pheBA operon depends on the extracellular concentration of phenol. The pheBA promoter is rapidly activated in the presence of micromolar concentrations of phenol in minimal growth medium, but the initiation of transcription from this promoter is severely delayed after sudden exposure of bacteria to 2.5 mM phenol. Second, the transcriptional activation from this promoter is impeded when the growth medium of bacteria contains amino acids. The negative effects of amino acids can be suppressed either by overproducing CatR or by increasing, the intracellular amount of CCM. However, the intracellular amount of CCM is a major limiting factor for the transcriptional activation of the pheBA operon, as accumulation of CCM in a P. putida catB-defective strain, unable to metabolize CCM (but expressing CatR at a natural level), almost completely relieves the negative effects of amino acids. The intracellular amount of CCM is negatively affected by the catabolite repression control protein via downregulating at the post-transcriptional level the expression of the pheBA-encoded catechol 1,2-dioxygenase and the phenol monooxygenase, the enzymes needed for CCM production.
Putrinš
Marta
M
Department of Genetics, Institute of Molecular and Cell Biology, Tartu University and Estonian Biocentre, 51010 Tartu, Estonia.
Tover
Andres
A
Department of Genetics, Institute of Molecular and Cell Biology, Tartu University and Estonian Biocentre, 51010 Tartu, Estonia.
Tegova
Radi
R
Department of Genetics, Institute of Molecular and Cell Biology, Tartu University and Estonian Biocentre, 51010 Tartu, Estonia.
Saks
Ülle
Ü
Department of Genetics, Institute of Molecular and Cell Biology, Tartu University and Estonian Biocentre, 51010 Tartu, Estonia.
Kivisaar
Maia
M
Department of Genetics, Institute of Molecular and Cell Biology, Tartu University and Estonian Biocentre, 51010 Tartu, Estonia.
D000596:Amino Acids / Q000494:pharmacology; D019677:Artificial Gene Fusion; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001565:Benzoates / Q000494:pharmacology; D015153:Blotting, Western; D050548:Catechol 1,2-Dioxygenase / Q000096:biosynthesis / Q000235:genetics*; D003470:Culture Media; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D017930:Genes, Reporter; D005998:Glycine / Q000378:metabolism; D008156:Luciferases / Q000032:analysis / Q000235:genetics; D014442:Monophenol Monooxygenase / Q000096:biosynthesis / Q000235:genetics*; D009876:Operon; D019800:Phenol / Q000378:metabolism* / Q000494:pharmacology; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic; D016958:Pseudomonas putida / Q000235:genetics / Q000378:metabolism*; D013011:Sorbic Acid / Q000031:analogs & derivatives / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic
D000596:Amino Acids; D001426:Bacterial Proteins; D001565:Benzoates; D003470:Culture Media; D004268:DNA-Binding Proteins; D014157:Transcription Factors; C086843:CatR protein, bacteria; D019800:Phenol; C005939:muconic acid; D050548:Catechol 1,2-Dioxygenase; D008156:Luciferases; D014442:Monophenol Monooxygenase; D005998:Glycine; D013011:Sorbic Acid