The cAMP-CRP/CytR nucleoprotein complex in Escherichia coli: two pairs of closely linked binding sites for the cAMP-CRP activator complex are involved in combinatorial regulation of the cdd promoter.
Transcription initiation at CytR regulated promoters in Escherichia coli is controlled by a combinatorial regulatory system in which the cAMP receptor protein (CRP) functions as both an activator and a co-repressor. By combining genetic studies and footprinting analyses, we demonstrate that regulated expression of the CytR controlled cdd promoter requires three CRP-binding sites: a high affinity site (CRP-1) and two overlapping low affinity sites (CRP-2 and CRP-3) centred at positions -41, -91 and -93, respectively. In the absence of CytR, cAMP-CRP interacts at one set of sites (CRP-1 and CRP-2) and both of these binding sites are required for full promoter activation. In the presence of CytR, however, the two regulators bind cooperatively to cddP forming a nucleoprotein complex in which cAMP-CRP binds to CRP-1 and CRP-3 and CytR occupies the sequence between these sites. Thus, association of the two regulators involves a repositioning of the cAMP-CRP complex. Moreover, mutant cdd promoters in which CRP-2 and CRP-3 have been deleted are partially regulated by CytR, and cAMP-CRP and CytR still bind cooperatively to these promoters. These findings provide clues to an understanding of how cAMP-CRP and CytR interact at a structurally diverse set of promoters.
Holst
B
B
Department of Molecular Biology, Odense University, Denmark.
Søgaard-Andersen
L
L
Pedersen
H
H
Valentin-Hansen
P
P
D001426:Bacterial Proteins / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites; D002352:Carrier Proteins / Q000378:metabolism*; D000242:Cyclic AMP / Q000378:metabolism*; D002373:Cyclic AMP Receptor Protein; D004269:DNA, Bacterial / Q000235:genetics; D004268:DNA-Binding Proteins / Q000378:metabolism*; D003850:Deoxyribonuclease I; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D029968:Escherichia coli Proteins; D017353:Gene Deletion; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011953:Receptors, Cyclic AMP / Q000378:metabolism*; D012097:Repressor Proteins / Q000378:metabolism*; D015183:Restriction Mapping
D001426:Bacterial Proteins; D002352:Carrier Proteins; D002373:Cyclic AMP Receptor Protein; C048425:CytR protein, E coli; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D011953:Receptors, Cyclic AMP; D012097:Repressor Proteins; D000242:Cyclic AMP; D003850:Deoxyribonuclease I