Analysis of Gal4-directed transcription activation using Tra1 mutants selectively defective for interaction with Gal4
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.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2012 |
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| Erschienen: |
2012 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:109 |
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| Enthalten in: |
Proceedings of the National Academy of Sciences of the United States of America - 109(2012), 6 vom: 07. Feb., Seite 1997-2002 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Lin, Ling [VerfasserIn] |
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| Links: |
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| Anmerkungen: |
Date Completed 30.03.2012 Date Revised 04.04.2024 published: Print-Electronic GEO: GSE31391 Citation Status MEDLINE |
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| doi: |
10.1073/pnas.1116340109 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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| PPN (Katalog-ID): |
NLM215167856 |
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| 245 | 1 | 0 | |a Analysis of Gal4-directed transcription activation using Tra1 mutants selectively defective for interaction with Gal4 |
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| 500 | |a published: Print-Electronic | ||
| 500 | |a GEO: GSE31391 | ||
| 500 | |a Citation Status MEDLINE | ||
| 520 | |a 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 | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a Research Support, N.I.H., Extramural | |
| 650 | 4 | |a Research Support, Non-U.S. Gov't | |
| 650 | 7 | |a DNA, Fungal |2 NLM | |
| 650 | 7 | |a DNA-Binding Proteins |2 NLM | |
| 650 | 7 | |a GAL4 protein, S cerevisiae |2 NLM | |
| 650 | 7 | |a Saccharomyces cerevisiae Proteins |2 NLM | |
| 650 | 7 | |a TRA1 protein, S cerevisiae |2 NLM | |
| 650 | 7 | |a Trans-Activators |2 NLM | |
| 650 | 7 | |a Transcription Factors |2 NLM | |
| 650 | 7 | |a Histone Acetyltransferases |2 NLM | |
| 650 | 7 | |a EC 2.3.1.48 |2 NLM | |
| 650 | 7 | |a GAL1 protein, S cerevisiae |2 NLM | |
| 650 | 7 | |a EC 2.7.1.6 |2 NLM | |
| 650 | 7 | |a Galactokinase |2 NLM | |
| 650 | 7 | |a EC 2.7.1.6 |2 NLM | |
| 650 | 7 | |a Galactose |2 NLM | |
| 650 | 7 | |a X2RN3Q8DNE |2 NLM | |
| 700 | 1 | |a Chamberlain, Lynn |e verfasserin |4 aut | |
| 700 | 1 | |a Zhu, Lihua J |e verfasserin |4 aut | |
| 700 | 1 | |a Green, Michael R |e verfasserin |4 aut | |
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| 856 | 4 | 0 | |u http://dx.doi.org/10.1073/pnas.1116340109 |3 Volltext |
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