Analysis of Gal4-directed transcription activation using Tra1 mutants selectively defective for interaction with Gal4
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PROVIDER: PRJNA145875 | ENA |
REPOSITORIES: ENA
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Project description:This SuperSeries is composed of the following subset Series: GSE31389: Gene expression profile of Tra1 GID (Gal4 interaction defective) mutants GSE31390: Gene expression profile of Tra1 dependent genes Refer to individual Series
2011-12-30 | E-GEOD-31391 | biostudies-arrayexpress
Project description:Promoter-specific transcriptional activators (activators) stimulate transcription through direct interactions with one or more components of the transcription machinery, termed the “target”. 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. To investigate the essentiality of Tra1 as a target of Gal4, here we derive Tra1 mutants that are selectively defective for interaction with Gal4 in vivo (Gal4 Interaction Defective (GID) mutants). In contrast to wild-type Tra1, Tra1 GID mutants are not recruited by Gal4 to the promoter and cannot support Gal4-directed transcription activation, demonstrating that the Gal4–Tra1 interaction is required for Gal4 function. In yeast strains expressing a Tra1 GID mutant, Gal4 promoter binding is unexpectedly also diminished indicating that Gal4 and Tra1 bind cooperatively. Consistent with cooperative binding, we demonstrate that the interaction between Gal4 and Tra1 occurs predominantly on the promoter and not off DNA. Finally, we show that although Tra1 is also targeted by other activators, these interaction are unaffected by GID mutations, revealing an unanticipated specificity of the Gal4-Tra1 interaction.
2011-12-30 | GSE31389 | GEO
Project description:Promoter-specific transcriptional activators (activators) stimulate transcription through direct interactions with one or more components of the transcription machinery, termed the “target”. 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. To investigate the essentiality of Tra1 as a target of Gal4, here we derive Tra1 mutants that are selectively defective for interaction with Gal4 in vivo (Gal4 Interaction Defective (GID) mutants). In contrast to wild-type Tra1, Tra1 GID mutants are not recruited by Gal4 to the promoter and cannot support Gal4-directed transcription activation, demonstrating that the Gal4–Tra1 interaction is required for Gal4 function. In yeast strains expressing a Tra1 GID mutant, Gal4 promoter binding is unexpectedly also diminished indicating that Gal4 and Tra1 bind cooperatively. Consistent with cooperative binding, we demonstrate that the interaction between Gal4 and Tra1 occurs predominantly on the promoter and not off DNA. Finally, we show that although Tra1 is also targeted by other activators, these interaction are unaffected by GID mutations, revealing an unanticipated specificity of the Gal4-Tra1 interaction. 3 samples were analyzed in duplicate with completely randomized design
2011-12-30 | E-GEOD-31389 | biostudies-arrayexpress
Project description:Promoter-specific transcriptional activators (activators) stimulate transcription through direct interactions with one or more components of the transcription machinery, termed the “target”. 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. To investigate the essentiality of Tra1 as a target of Gal4, here we derive Tra1 mutants that are selectively defective for interaction with Gal4 in vivo (Gal4 Interaction Defective (GID) mutants). In contrast to wild-type Tra1, Tra1 GID mutants are not recruited by Gal4 to the promoter and cannot support Gal4-directed transcription activation, demonstrating that the Gal4–Tra1 interaction is required for Gal4 function. In yeast strains expressing a Tra1 GID mutant, Gal4 promoter binding is unexpectedly also diminished indicating that Gal4 and Tra1 bind cooperatively. Consistent with cooperative binding, we demonstrate that the interaction between Gal4 and Tra1 occurs predominantly on the promoter and not off DNA. Finally, we show that although Tra1 is also targeted by other activators, these interaction are unaffected by GID mutations, revealing an unanticipated specificity of the Gal4-Tra1 interaction.
2011-12-30 | GSE31390 | GEO
Project description:Promoter-specific transcriptional activators (activators) stimulate transcription through direct interactions with one or more components of the transcription machinery, termed the “target”. 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. To investigate the essentiality of Tra1 as a target of Gal4, here we derive Tra1 mutants that are selectively defective for interaction with Gal4 in vivo (Gal4 Interaction Defective (GID) mutants). In contrast to wild-type Tra1, Tra1 GID mutants are not recruited by Gal4 to the promoter and cannot support Gal4-directed transcription activation, demonstrating that the Gal4–Tra1 interaction is required for Gal4 function. In yeast strains expressing a Tra1 GID mutant, Gal4 promoter binding is unexpectedly also diminished indicating that Gal4 and Tra1 bind cooperatively. Consistent with cooperative binding, we demonstrate that the interaction between Gal4 and Tra1 occurs predominantly on the promoter and not off DNA. Finally, we show that although Tra1 is also targeted by other activators, these interaction are unaffected by GID mutations, revealing an unanticipated specificity of the Gal4-Tra1 interaction. Strain MDC1(TRA1 WT) and MDC3 (Tra1 ts) were grown at 30˚C and then shifted to 37˚C for 60, 90 and 120 mins.
2011-12-30 | E-GEOD-31390 | biostudies-arrayexpress
Project description: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.
| S-EPMC3277556 | biostudies-literature
Project description:This SuperSeries is composed of the SubSeries listed below.
2011-12-30 | GSE31391 | GEO
Project description:Tra1 is an essential component of the S. cerevisiae SAGA and NuA4 complexes. Using targeted mutagenesis, we identified residues within its C-terminal phosphatidylinositol-3 kinase (PI3K) domain that are required for function. The phenotype of tra1-P3408A, S3463A and SRR3413-3415AAA included temperature sensitivity and reduced growth in media containing 6% ethanol or aminotriazole. These alleles resulted in ≥2-fold change in expression of ~7% of yeast genes in rich media and reduced activation of PHO5 and ADH2 promoters. Tra1-SRR3413 associated with components of both the NuA4 and SAGA complexes and with the Gal4 transcriptional activation domain similar to wild-type protein. Tra1-SRR3413 resulted in a decreased ratio of acetylated histone H3 and H4 to total histone H3 at the PHO5 promoter that appeared largely due to increased unacetylated histone. Slow growth of a tra1-SRR3413 strain in aminotriazole was suppressed by deletion of Vps1 and the Vps1 interacting protein, Rad16. In addition, ethanol sensitivity was suppressed by loss of Vam3, suggesting that Tra1 is involved with protein trafficking and membrane sorting. Furthermore, tra1-SRR3413 results in generation dependent telomere shortening. While the tra1 alleles have some phenotypic similarities with deletions of SAGA and/or NuA4 components, the pattern of gene expression, genetic interactions and telomere shortening suggest possible novel functions for the PI3K-domain of Tra1. Keywords: yeast, Tra1, PI3-kinase domain, gene expression, genetic modification
2007-10-10 | GSE6847 | GEO
Project description:Gene expression profile of Tra1 GID (Gal4 interaction defective) mutants
| PRJNA154141 | ENA
Project description:Tra1 is a component of the Saccharomyces cerevisiae SAGA and NuA4 complexes and a member of the phosphatidylinositol 3-kinase kinase family that contain a C-terminal PI3K domain followed by a 35-residue FATC domain. We analyzed alleles with single residue changes in the FATC domain. Of these tra1-L3733A had the most pronounced effects, resulting in phenotypes similar but not identical to those resulting from mutations in the PI3K domain or SAGA or NuA4 components. To analyze the transcriptional effects of the tra1-L3733A allele, we performed gene profiling of wild-type and tra1-L3733A strains. Expression of 11 genes was elevated ≥ 2-fold, 79 genes were decreased ≥ 2-fold. Of the 11 genes with elevated expression, four (HSP26, PIR3, DDR2 and GRE1) have roles in cellular stress response. The only gene ontology process significantly enriched in the genes with decreased expression greater than 2-fold was lysine biosynthesis (GO: 0009085). Broader analysis of gene ontology processes revealed expression of genes involved in cellular respiration (GO:0045333) and the overlapping process generation of precursor metabolites and energy (GO: 0006091) showed an average upward shift of ~10% relative to the total collection of genes. A similar trend of less magnitude was seen for vitamin metabolic processes (GO:0006766). A slight downward shift was seen for genes involved in translation (GO:0006412) and membrane organization (GO:0016044).
2010-09-20 | GSE18591 | GEO