title: A severely defective TATA-binding protein-TFIIB interaction does not preclude transcriptional activation in vivo.
abstract: In yeast cells, mutations in the TATA-binding protein (TBP) that disrupt the interaction with the TATA element or with TFIIA can selectively impair the response to acidic activator proteins. We analyzed the transcriptional properties of TBP derivatives in which residues that directly interact with TFIIB were replaced by alanines. Surprisingly, a derivative with a 50-fold defect in TBP-TFIIB-TATA complex formation in vitro (E188A) supports viability and responds efficiently to activators in vivo. The E186A derivative, which displays a 100-fold defect in TBP-TFIIB-TATA complex formation, does not support viability, yet it does respond to activators. Conversely, the L189A mutation, which has the mildest effect on the interaction with TFIIB (10-fold), can abolish transcriptional activation and cell viability when combined with mutations on the DNA-binding surface. This "synthetic lethal" effect is not observed with E188A, suggesting that the previously described role of L189 in transcriptional activation may be related to its location on the DNA-binding surface and not to its interaction with TFIIB. Finally, when using TBP mutants defective on multiple interaction surfaces, we observed synthetic lethal effects between mutations on the TFIIA and TFIIB interfaces but found that mutations implicated in association with polymerase II and TFIIF did not have significant effects in vivo. Taken together, these results argue that, unlike the TBP-TATA and TBP-TFIIA interactions, the TBP-TFIIB interaction is not generally limiting for transcriptional activation in vivo.
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115, USA.
authors.forename: M
authors.initials: M
authors.lastname: Lee
authors.forename: K
authors.initials: K
authors.lastname: Struhl
mesh_terms: D000409:Alanine; D000595:Amino Acid Sequence; D017124:Conserved Sequence; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D009154:Mutation; D010641:Phenotype; D011485:Protein Binding; D012319:RNA Polymerase II / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D035181:TATA-Box Binding Protein; D035561:Transcription Factor TFIIA; D035581:Transcription Factor TFIIB; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation / Q000502:physiology*; D015003:Yeasts / Q000235:genetics / Q000254:growth & development
chemical_list: D004268:DNA-Binding Proteins; D035181:TATA-Box Binding Protein; D035561:Transcription Factor TFIIA; D035581:Transcription Factor TFIIB; D014157:Transcription Factors; D012319:RNA Polymerase II; D000409:Alanine