lacP1 promoter with an extended -10 motif. Pleiotropic effects of cyclic AMP protein at different steps of transcription initiation.
The cyclic AMP receptor protein (CRP), which activates transcription from the wild-type lacP1 promoter and most of its mutants, represses productive RNA synthesis from a lacP1 promoter variant that contains an extended -10 element, although CRP enhances RNA polymerase binding as well as open complex formation in both promoters. Moreover, abortive RNA synthesis, which is already higher in the extended -10 variant compared with the parent promoter, was further enhanced by CRP. These results, together with the observed decrease in productive RNA synthesis, indicate that CRP, while facilitating the earlier steps of initiation, inhibits transcription from the extended -10 lacP1 by hindering promoter clearance. We propose that CRP decreases energetic barriers to RNA polymerase binding, isomerization, and abortive RNA synthesis but stabilizes the abortive RNA initiating complex, which results in increasing the activation energy of the transition state before the elongation complex. The results demonstrate for the first time that a DNA-binding regulatory protein acts as an activator or a repressor in different steps of the transcription initiation pathway because of the energetic differences of the intermediate complex in the same promoter.
Liu
Mofang
M
Laboratory of Molecular Biology, Center for Cancer Research, NCI, National Institutes of Health, Bethesda, Maryland 20892-4264, USA.
Garges
Susan
S
Adhya
Sankar
S
D001483:Base Sequence; D002373:Cyclic AMP Receptor Protein / Q000235:genetics / Q000502:physiology*; D018983:DNA Footprinting; D004269:DNA, Bacterial / Q000737:chemistry / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000235:genetics / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D004926:Escherichia coli / Q000235:genetics; D007763:Lac Operon / Q000235:genetics*; D009154:Mutation; D016133:Polymerase Chain Reaction; D011196:Potassium Permanganate; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012329:RNA, Bacterial / Q000096:biosynthesis; D012097:Repressor Proteins / Q000235:genetics / Q000502:physiology; D013816:Thermodynamics; D014158:Transcription, Genetic / Q000502:physiology*
D002373:Cyclic AMP Receptor Protein; D004269:DNA, Bacterial; D012329:RNA, Bacterial; D012097:Repressor Proteins; D011196:Potassium Permanganate; D012321:DNA-Directed RNA Polymerases