title: Tandem DNA-bound cAMP-CRP complexes are required for transcriptional repression of the deoP2 promoter by the CytR repressor in Escherichia coli.
abstract: We have studied the deoP2 promoter in Escherichia coli to define features important for its interaction with the CytR repressor. As is characteristic for CytR-regulated promoters, deoP2 encodes tandem binding sites for the activating complex cAMP-CRP. One of these sites, CRP-1, overlaps the -35 region, and is sufficient for activation; the second site, CRP-2, centred around -93, is indispensable for repression. Here we demonstrate, by means of in vivo titration, that CytR interaction with deoP2 depends not only on CRP-2, but also on CRP-1 and the length and possibly the sequence separating these two sites. Also, point mutations in either CRP site reduce or abolish CytR titration; however, no co-operativity is observed in the interaction of CytR with the two CRP binding sites. Furthermore, the reduction in CytR titration parallels the reduction in binding of cAMP-CRP to the mutated CRP sites in vitro. These observations are not easily explained by current models for the action of prokaryotic repressors; instead we favour a model in which the interaction of CytR with deoP2 depends on the presence of tandem DNA-bound cAMP-CRP complexes.
authors.affiliation: Department of Molecular Biology, Odense University, Denmark.
authors.forename: L
authors.initials: L
authors.lastname: Søgaard-Andersen
authors.forename: N E
authors.initials: NE
authors.lastname: Møllegaard
authors.forename: S R
authors.initials: SR
authors.lastname: Douthwaite
authors.forename: P
authors.initials: P
authors.lastname: Valentin-Hansen
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D000242:Cyclic AMP / Q000378:metabolism; D002373:Cyclic AMP Receptor Protein / Q000235:genetics / Q000378:metabolism*; D004269:DNA, Bacterial / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D029968:Escherichia coli Proteins; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D009154:Mutation; D011401:Promoter Regions, Genetic; D012097:Repressor Proteins / Q000378:metabolism*; D014158:Transcription, Genetic
chemical_list: D001426:Bacterial Proteins; D002373:Cyclic AMP Receptor Protein; C048425:CytR protein, E coli; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D012097:Repressor Proteins; D000242:Cyclic AMP