Two roles for the yeast transcription coactivator SAGA and a set of genes redundantly regulated by TFIID and SAGA
- Fred Hutchinson Cancer Research Center, United States
Figures
Figure 1 with 4 supplements
Two distinct classes of yeast genes based on SAGA and TFIID dependence.
(A) Scatter plots comparing normalized signal per gene in corresponding samples. Log10 scale is used for X and Y axes. Mean values from replicate experiments are plotted. All presented experiments …
Figure 1—figure supplement 1
Biological replicates for 4-thioU RNA-seq experiments show low variability.
(A) Average coefficients of variation (CV) for selected experiments vs the percentage of genes analyzed (after first filtering out 639 genes which did not have RNA-seq signal in at least one …
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Figure 1—figure supplement 1—source data 1
Coefficient of variation values for different experiments used to plot Figure 1—figure supplement 1.
- https://cdn.elifesciences.org/articles/50109/elife-50109-fig1-figsupp1-data1-v2.xlsx
Figure 1—figure supplement 2
Auxin-degron system allows for efficient degradation of target TFIID and SAGA subunits.
(A) Degron depletion in single or double degron strains. Western blot showing degron efficiency in cells grown in synthetic complete media. Experiment was done in biological triplicates (A, B, and C …
Figure 1—figure supplement 3
Validation of mutant strain fitness and enrichment of nascent transcripts in 4thioU RNA-seq experiments.
(A) Box plot of gene expression level in selected experiments following DMSO treatment. Mean value for replicate samples was used. Log10 scale is used for Y axis. (B) Genome browser images showing …
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Figure 1—figure supplement 3—source data 1
Mean values of log10 expression per gene for selected DMSO treated samples used to plot Figure 1—figure supplement 3A.
- https://cdn.elifesciences.org/articles/50109/elife-50109-fig1-figsupp3-data1-v2.xlsx
Figure 1—figure supplement 4
Mutant genotype and IAA treatment for 30 minutes do not compromise viability of tested strains.
(A) Correlation matrix of Pearson correlation coefficient values between log2 changes in transcription in SAGA and TFIID depletion experiments and the slow growth signature defined in O'Duibhir et …
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Figure 1—figure supplement 4—source data 1
Average log2 changes in transcription in the degron and deletion strains same as in Supplementary file 3 with additional column (SGS) containing the values of slow growth signature defined in O'Duibhir et al. (2014).
Table was used to plot the heatmap in Figure 1—figure supplement 4A.
- https://cdn.elifesciences.org/articles/50109/elife-50109-fig1-figsupp4-data1-v2.xlsx
Figure 2 with 1 supplement
Simultaneous depletion of SAGA and TFIID severely affects transcription from almost all genes.
(A) Box plot showing the log2 change in transcription per gene upon perturbing cellular levels of SAGA or TFIID. Genes are divided into two classes according to results of the k-means clustering …
Figure 2—figure supplement 1
Coactivator-redundant genes show extensive loss transcription upon depletion of both TFIID and SAGA.
(A) Scatter plots comparing results of Taf13-deg and Taf13/Spt3-deg experiments for coactivator-redundant (CR) genes and TFIID genes. (B) Scatter plot of gene expression rank vs the difference …
Figure 3 with 2 supplements
Properties associated with TFIID-dependent and coactivator-redundant genes.
(A) Bar plot comparing the number of genes in each gene class as a function of expression level (left panel) and among the top 10% highly transcribed genes (right panel). Quintile five contains the …
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Figure 3—source data 1
Table containing gene classification defined in this work and the TATA-containing/TATA-less definitions from Rhee and Pugh (2012).
Data was used to plot Figure 3D.
- https://cdn.elifesciences.org/articles/50109/elife-50109-fig3-data1-v2.xlsx
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Figure 3—source data 2
Tables containing gene classification defined in this work and the classification of genes into categories based on the presence of a TATA-box or Msn2/4 binding site based on promoter search performed in this work.
Data were used to plot Figure 3E.
- https://cdn.elifesciences.org/articles/50109/elife-50109-fig3-data2-v2.xlsx
Figure 3—figure supplement 1
Properties of the TFIID and coactivator-redundant gene classes.
(A) Barplot comparing previously defined categories ‘TFIID-dominated’ and ‘SAGA-dominated’ genes (Huisinga and Pugh, 2004) with categories defined in this study. (B) Boxplot representation of …
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Figure 3—figure supplement 1—source data 1
Table containing gene classification defined in this work and the gene classes defined in Huisinga and Pugh (2004).
Data was used to plot Figure 3—figure supplement 1A.
- https://cdn.elifesciences.org/articles/50109/elife-50109-fig3-figsupp1-data1-v2.xlsx
Figure 3—figure supplement 2
The number of ribosomal protein (RP) genes within each expression quintile.
Figure 4
Changes in transcription (from 4thioU RNA-seq) observed in the SAGA deletion strains and in the Spt3/7 degron strain after rapid depletion.
Mean values from replicate experiments are plotted and the genes are divided into TFIID-dependent and coactivator-redundant (CR) categories.
Figure 5 with 2 supplements
Histone modifications in SPT deletion and degron strains.
(A, B) Quantitation of total H3K18-Ac and H2B-Ub in the SPT deletion and degron strains, normalized to the TFIIH subunit Tfg2 as a loading control. Data are from Figure 5—figure supplement 1. In …
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Figure 5—source data 1
Quantification of western blot results presented in Figure 5—figure supplement 1.
Data was used to make graphs in Figure 5AB.
- https://cdn.elifesciences.org/articles/50109/elife-50109-fig5-data1-v2.xlsx
Figure 5—figure supplement 1
Histone modifications in SPT deletion and degron strains.
(A, B) Quantitation of total H3K18-Ac and H2B-Ub in the SPT deletion and degron strains as indicated. Depletion of the degron-tagged subunits is also shown. Data used for quantitation in Figure 5. …
Figure 5—figure supplement 2
ChIP-seq analysis of H3K18-Ac signal in the SAGA deletion mutants and Spt3/7 degron strains.
The degron strain was treated for 30 min with IAA or DMSO before formaldehyde crosslinking. (A) Genome browser images showing comparison of normalized signal at representative genomic locations. …
Figure 6 with 1 supplement
TFIID and SAGA show widespread binding to both gene classes.
(A) Average plots of Taf1, Taf7, Taf13, Spt3 and Spt7 ChEC DNA cleavage versus free MNase cleavage. Signals were averaged for bound promoters only. (B) Venn diagram showing the overlap of bound …
Figure 6—figure supplement 1
Different TFIID and SAGA subunits show extensive overlap of binding sites.
(A) Venn diagrams showing the overlap of promoters bound by different factors. (B) Genome browser images showing comparison of normalized signal at representative genomic locations. Identified …
Figure 7 with 1 supplement
Rapid SAGA depletion does not prevent transcription activation by Gcn4.
(A) RT-qPCR analysis of 4-thio Uracil labeled RNA purified from indicated SPT deletion strains in the presence or absence of SM or (B) from SPT-degron strains induced with SM either before or after …
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Figure 7—source data 1
Data from RT-qPCR analysis used for the plots in Figure 7.
- https://cdn.elifesciences.org/articles/50109/elife-50109-fig7-data1-v2.xlsx
Figure 7—figure supplement 1
Degron efficiency in synthetic complete media with or without SM induction and IAA addition.
(A) Western analysis of the Spt/7-degron strain without SM and with or without IAA. (B) Western analysis of the Spt3/7-degron strain treated with SM either before or after IAA addition. Blot was …
Figure 8
Model for roles of TFIID and SAGA and two different gene classes.
Of the ~83% of yeast protein-coding genes analyzed in this study (4900 genes), the number of genes in each class is shown. Expression of genes in the TFIID set is strongly TFIID-dependent with …
Additional files
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Supplementary file 1
Spike-in normalized signal for all genes (5158) which had detectable transcription in 48 RNA-seq samples collected in this study (samples for simultaneous depletion of SAGA and TFIID, GCN5 and UBP8 deletion experiments and WT control auxin experiment were not used for this analysis).
Expression column is the average signal for all DMSO and WT samples. This value was further normalized by the gene length to give normalized expression which was used to sort the genes from the highest to the lowest expression.
- https://cdn.elifesciences.org/articles/50109/elife-50109-supp1-v2.xlsx
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Supplementary file 2
Average spike-in normalized signal for replicate experiments for the final set of 4900 genes analyzed in this study and average expression per gene (based on DMSO and WT experiments).
Rows are sorted by the average expression.
- https://cdn.elifesciences.org/articles/50109/elife-50109-supp2-v2.xlsx
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Supplementary file 3
Average log2 changes in transcription in the degron and deletion strains, results of k-means clustering and average expression per gene (based on DMSO and WT experiments).
Rows are sorted by the average expression.
- https://cdn.elifesciences.org/articles/50109/elife-50109-supp3-v2.xlsx
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Supplementary file 4
Average log2 changes in transcription from degron experiments simultaneously depleting SAGA and TFIID components (Spt3/Taf13 and Spt7/Taf13).
Data for other strains, results of k-means clustering and average expression per gene are the same as shown in Supplementary file 3. Rows are sorted by the average expression.
- https://cdn.elifesciences.org/articles/50109/elife-50109-supp4-v2.xlsx
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Supplementary file 5
Motif enrichments found in the TFIID-dependent and coactivator-redundant gene classes.
- https://cdn.elifesciences.org/articles/50109/elife-50109-supp5-v2.xlsx
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Supplementary file 6
Average log2 changes in transcription for GCN5 and UBP8 deletion experiments.
Data for other strains, results of k-means clustering and average expression per gene are the same as shown in Supplementary file 3. Rows are sorted by the average expression.
- https://cdn.elifesciences.org/articles/50109/elife-50109-supp6-v2.xlsx
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Supplementary file 7
Average log2 changes in H3K18-Ac signal in the SAGA deletion mutants and Spt3/7 degron strain.
- https://cdn.elifesciences.org/articles/50109/elife-50109-supp7-v2.xlsx
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Supplementary file 8
Average spike-in normalized ChEC signals at bound promoters for the following MNase-fusions: Taf1, Taf7, Taf13, Spt3 and Spt7.
Genes in each table are sorted by signal intensity.
- https://cdn.elifesciences.org/articles/50109/elife-50109-supp8-v2.xlsx
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Supplementary file 9
S. cerevisiae and S. pombe strains used in this study.
Strains were validated using a combination of genetic assays, phenotypic analysis, Western analysis, PCR analysis and nucleic acid sequencing.
- https://cdn.elifesciences.org/articles/50109/elife-50109-supp9-v2.docx
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Transparent reporting form
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