The role of FIS protein in the physiological control of the expression of the Escherichia coli meta-hpa operon.
Expression from the Escherichia coli W meta-hpa operon promoter (Pg) is under a strict catabolic repression control mediated by the cAMP-catabolite repression protein (CRP) complex in a glucose-containing medium. The Pg promoter is also activated by the integration host factor (IHF) and repressed by the specific transcriptional regulator HpaR when 4-hydroxyphenylacetate (4HPA) is not present in the medium. Expression from the hpa promoter is also repressed in undefined rich medium such as LB, but the molecular basis of this mechanism is not understood. We present in vitro and in vivo studies to demonstrate the involvement of FIS protein in this catabolic repression. DNase I footprinting experiments show that FIS binds to multiple sites within the Pg promoter. FIS-site I overlaps the CRP-binding site. By using an electromobility shift assay, we demonstrated that FIS efficiently competes with CRP for binding to the Pg promoter, suggesting an antagonist/competitive mechanism. RT-PCR showed that the Pg repression effect is relieved in a FIS deleted strain. The repression role of FIS at Pg was further demonstrated by in vitro transcription assays. These results suggest that FIS contributes to silencing the Pg promoter in the exponential phase of growth in an undefined rich medium when FIS is predominantly expressed.
Galán
Beatriz
B
Department of Molecular Microbiology, Centro de Investigaciones Biológicas, CSIC, Madrid, Spain.
Manso
Isabel
I
Department of Molecular Microbiology, Centro de Investigaciones Biológicas, CSIC, Madrid, Spain.
Kolb
Annie
A
Unité de Génétique Moléculaire-URA 2172, Institut Pasteur, Paris, France.
García
José Luis
JL
Department of Molecular Microbiology, Centro de Investigaciones Biológicas, CSIC, Madrid, Spain.
Prieto
María A
MA
Department of Molecular Microbiology, Centro de Investigaciones Biológicas, CSIC, Madrid, Spain.
D001483:Base Sequence; D001665:Binding Sites; D003470:Culture Media / Q000737:chemistry; D004926:Escherichia coli / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D035021:Factor For Inversion Stimulation Protein / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D015964:Gene Expression Regulation, Bacterial; D034982:Integration Host Factors / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D009876:Operon; D011401:Promoter Regions, Genetic; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic
D003470:Culture Media; D029968:Escherichia coli Proteins; D035021:Factor For Inversion Stimulation Protein; C501034:Fis protein, E coli; D034982:Integration Host Factors; D012097:Repressor Proteins; C440502:integration host factor, E coli