Activation of RNA polymerase II transcription by the specific DNA-binding protein LSF. Increased rate of binding of the basal promoter factor TFIIB.
While the components of the initiation complex at an RNA polymerase II basal promoter have been well characterized, few mechanistic studies have focused on how upstream DNA-binding, transcriptional activators influence protein assembly at the initiation site. Our analysis of basal transcription on both the simian virus 40 and adenovirus major late promoters demonstrates that two slow steps in initiation of transcription are the assembly of the general transcription factors TFIID and TFIIB onto the template DNA. On the simian virus 40 major late promoter, the rate of initiation complex formation is dramatically increased in the presence of the cellular transcriptional activator LSF. Direct analysis by band mobility shift assays demonstrates that LSF has no effect on the rate of binding, or the stability of TFIID on the promoter, predicting that LSF would not affect the template commitment step. Rather, kinetic analyses demonstrate that LSF reduces the lag in the rate of initiation complex formation attributable to the slow addition of TFIIB and suggest that LSF increases the rate of association of TFIIB with the committed template. In addition, LSF increases the total number of transcription complexes in long term assays, which is also consistent with LSF increasing the rate of association of TFIIB, where TFIIB is not saturating. These results indicate a mechanism for the activation of the initiation of RNA polymerase II transcription by one upstream activating protein, LSF. This mechanism may also be applicable to other activators that function in cases where limiting concentrations of TFIIB in the cell dictate slow binding of TFIIB.
Sundseth
R
R
Laboratory of Eukaryotic Transcription, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts 02115.
Hansen
U
U
D001483:Base Sequence; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D012319:RNA Polymerase II / Q000378:metabolism*; D013539:Simian virus 40 / Q000235:genetics; D035581:Transcription Factor TFIIB; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic
D004268:DNA-Binding Proteins; D035581:Transcription Factor TFIIB; D014157:Transcription Factors; D004247:DNA; D012319:RNA Polymerase II