Mechanism of action of an acidic transcriptional activator in vitro.
Transcription of a eukaryotic structural gene by RNA polymerase II requires the ordered assembly of general transcription factors on the promoter to form a pre-initiation complex. Here we analyze affinity-purified complexes at various stages of assembly to determine the mechanism of action of an acidic transcriptional activator. We show that the activator can function in the absence of ATP and stimulates transcription by increasing the number of functional preinitiation complexes. The activator effects this increase by recruiting the general transcription factor TFIIB to the promoter. Using protein affinity chromatography we demonstrate a specific interaction between an acidic activating region and TFIIB. Based on these combined results, we propose that TFIIB is a direct target of an acidic activator.
Lin
Y S
YS
Department of Biochemistry and Molecular Biology, Harvard University, Cambridge, Massachusetts 02138.
Green
M R
MR
D002467:Cell Nucleus / Q000378:metabolism*; D004268:DNA-Binding Proteins; D004926:Escherichia coli / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D005796:Genes; D006367:HeLa Cells / Q000378:metabolism; D006801:Humans; D008957:Models, Genetic; D010957:Plasmids; D011401:Promoter Regions, Genetic; D012319:RNA Polymerase II / Q000302:isolation & purification / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D013698:Templates, Genetic; D035581:Transcription Factor TFIIB; D014157:Transcription Factors / Q000302:isolation & purification / Q000378:metabolism*; D014158:Transcription, Genetic
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D035581:Transcription Factor TFIIB; D014157:Transcription Factors; D012319:RNA Polymerase II