Program in Gene Function and Expression; Program in Molecular Medicine
Binding Sites; Biological Assay; DNA, Fungal; DNA-Binding Proteins; Fluorescence; Galactokinase; Galactose; Gene Expression Regulation, Fungal; Histone Acetyltransferases; Mutation; Promoter Regions, Genetic; Protein Binding; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Trans-Activators; Transcription Factors; Transcriptional Activation
Genetics and Genomics
Promoter-specific transcriptional activators (activators) stimulate transcription through direct interactions with one or more components of the transcription machinery, termed the "target." The identification of direct in vivo targets of activators has been a major challenge. Previous studies have provided evidence that the Tra1 subunit of the yeast SAGA (Spt-Ada-Gcn5-acetyltransferase) complex is the target of the yeast activator Gal4. However, several other general transcription factors, in particular the mediator complex, have also been implicated as Gal4 targets. Here we perform a large-scale genetic screen to derive and characterize tra1 alleles that are selectively defective for interaction with Gal4 in vivo [Gal4 interaction defective (GID) mutants]. In contrast to WT Tra1, Tra1 GID mutants are not recruited by Gal4 to the promoter and cannot support Gal4-directed transcription, demonstrating the essentiality of the Gal4-Tra1 interaction. In yeast strains expressing a Tra1 GID mutant, binding of Gal4 to the promoter is unexpectedly also diminished, indicating that Gal4 and Tra1 bind cooperatively. Consistent with cooperative binding, we demonstrate that the Gal4-Tra1 interaction occurs predominantly on the promoter and not off DNA. Finally, we show that although Tra1 is targeted by other activators, these interactions are unaffected by GID mutations, revealing an unanticipated specificity of the Gal4-Tra1 interaction.
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DOI of Published Version
Ling Lin, Lynn Chamberlain, Lihua J. Zhu, and Michael R. Green. Analysis of Gal4-directed transcription activation using Tra1 mutants selectively defective for interaction with Gal4. Proc Natl Acad Sci U S A. 2012 Feb 7;109(6):1997-2002. Epub 2012 Jan 23. Link to article on publisher's site
Proceedings of the National Academy of Sciences of the United States of America
Lin, Ling; Chamberlain, Lynn; Zhu, Lihua Julie; and Green, Michael R., "Analysis of Gal4-directed transcription activation using Tra1 mutants selectively defective for interaction with Gal4" (2012). Program in Gene Function and Expression Publications and Presentations. 180.