Transcription activation by the Escherichia coli cyclic AMP receptor protein. Receptors bound in tandem at promoters can interact synergistically.
Starting with a semi-synthetic Escherichia coli promoter with a binding site for the cyclic AMP receptor protein (CRP) centred between base-pairs 41 and 42 upstream from the transcription start site, a second upstream CRP-binding site, centred between base-pairs 90 and 91, was introduced. CRP binding to this second upstream site results in a several-fold greater stimulation of CRP-dependent transcription initiation, compared to activation at the starting promoter with just one CRP-binding site. Activation of transcription by the upstream CRP molecule is blocked by the HL159 substitution, suggesting that the upstream-bound CRP makes a direct contact with RNA polymerase. Footprinting experiments suggest that RNA polymerase contacts the promoter DNA between the two CRP-binding sites, most likely due to interactions involving the C-terminal part of the alpha subunit. Synergy between tandem bound CRP molecules in transcription activation requires that the two CRP-binding sites be separated by around 40 or 50 base-pairs, but is not found at intermediate spacings. An experiment in which the upstream CRP-binding site is replaced by a site for the related transcription factor, FNR, shows that heterologous synergistic interactions between FNR and CRP are possible.
Busby
S
S
School of Biochemistry, University of Birmingham, U.K.
West
D
D
Lawes
M
M
Webster
C
C
Ishihama
A
A
Kolb
A
A
D001426:Bacterial Proteins / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D002373:Cyclic AMP Receptor Protein / Q000378:metabolism* / Q000502:physiology; D004269:DNA, Bacterial / Q000378:metabolism*; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins; D007506:Iron-Sulfur Proteins; D008954:Models, Biological; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012329:RNA, Bacterial / Q000096:biosynthesis; D012333:RNA, Messenger / Q000096:biosynthesis; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic; D015533:Transcriptional Activation / Q000502:physiology*
D001426:Bacterial Proteins; D002373:Cyclic AMP Receptor Protein; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; C060704:FNR protein, E coli; D007506:Iron-Sulfur Proteins; D012329:RNA, Bacterial; D012333:RNA, Messenger; D014157:Transcription Factors; D012321:DNA-Directed RNA Polymerases