Differential requirement of SAGA components for recruitment of TATA-box-binding protein to promoters in vivo.
The multisubunit Saccharomyces cerevisiae SAGA (Spt-Ada-Gcn5-acetyltransferase) complex is required to activate transcription of a subset of RNA polymerase II-dependent genes. However, the contribution of each SAGA component to transcription activation is relatively unknown. Here, using a formaldehyde-based in vivo cross-linking and chromatin immunoprecipitation assay, we have systematically analyzed the role of SAGA components in the recruitment of TATA-box binding protein (TBP) to SAGA-dependent promoters. We show that recruitment of TBP is diminished at a number of SAGA-dependent promoters in ada1delta, spt7delta, and spt20delta null mutants, consistent with previous biochemical data suggesting that these components maintain the integrity of the SAGA complex. We also find that Spt3p is generally required for TBP binding to SAGA-dependent promoters, consistent with biochemical and genetic experiments, suggesting that Spt3p interacts with and recruits TBP to the core promoter. By contrast, Spt8p, which has been proposed to be required for the interaction between Spt3p and TBP, is required for TBP binding at only a subset of SAGA-dependent promoters. Ada2p and Ada3p are both required for TBP recruitment to Gcn5p-dependent promoters, supporting previous biochemical data that Ada2p and Ada3p are required for the histone acetyltransferase activity of Gcn5p. Finally, our results suggest that TBP-associated-factor components of SAGA are differentially required for TBP binding to SAGA-dependent promoters. In summary, we show that SAGA-dependent promoters require different combinations of SAGA components for TBP recruitment, revealing a complex combinatorial network for transcription activation in vivo.
Bhaumik
Sukesh R
SR
Howard Hughes Medical Institute, Programs in Gene Expression and Function and Molecular Medicine, University of Massachusetts Medical School, Worcester, Massachusetts 01605, USA.
Green
Michael R
MR
D000123:Acetyltransferases / Q000378:metabolism*; D048868:Adaptor Proteins, Signal Transducing; D002843:Chromatin / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D004305:Dose-Response Relationship, Drug; D005557:Formaldehyde / Q000494:pharmacology; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism* / Q000502:physiology; D015966:Gene Expression Regulation, Fungal; D005800:Genes, Fungal; D051548:Histone Acetyltransferases; D009154:Mutation; D016133:Polymerase Chain Reaction; D011233:Precipitin Tests; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011494:Protein Kinases; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000502:physiology; D035181:TATA-Box Binding Protein; D013696:Temperature; D013997:Time Factors; D015534:Trans-Activators / Q000502:physiology; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D000099224:p300-CBP-Associated Factor
D000123:Acetyltransferases; D048868:Adaptor Proteins, Signal Transducing; D002843:Chromatin; D004268:DNA-Binding Proteins; D005557:Formaldehyde; D005656:Fungal Proteins; D051548:Histone Acetyltransferases; D011494:Protein Kinases; D029701:Saccharomyces cerevisiae Proteins; D035181:TATA-Box Binding Protein; D015534:Trans-Activators; D014157:Transcription Factors; C090575:ADA2 protein, S cerevisiae; C106162:HFI1 protein, S cerevisiae; C083323:NGG1 protein, S cerevisiae; C099690:SPT20 protein, S cerevisiae; C075582:SPT3 protein, S cerevisiae; C093107:SPT7 protein, S cerevisiae; C089445:SPT8 protein, S cerevisiae; C501392:GCN5 protein, S cerevisiae; D000099224:p300-CBP-Associated Factor