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Adr1 and Cat8 mediate coactivator recruitment and chromatin remodeling at glucose-regulated genes.

Biddick RK, Law GL, Young ET - PLoS ONE (2008)

Bottom Line: We determined the individual contributions of Cat8 and Adr1 on the expression of a cohort of glucose-repressed genes and found three broad categories: genes that need both activators for full derepression, genes that rely mostly on Cat8 and genes that require only Adr1.These promoter-specific contributions are also apparent in the chromatin remodeling that accompanies derepression: ADH2 requires both Adr1 and Cat8, whereas, at FBP1, significant remodeling occurs with Cat8 alone.Thus, at many of the glucose-repressed genes, Cat8 and Adr1 appear to have interchangeable roles and promoter architecture may dictate the roles of these activators.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, University of Washington, Seattle, Washington, United States of America.

ABSTRACT

Background: Adr1 and Cat8 co-regulate numerous glucose-repressed genes in S. cerevisiae, presenting a unique opportunity to explore their individual roles in coactivator recruitment, chromatin remodeling, and transcription.

Methodology/principal findings: We determined the individual contributions of Cat8 and Adr1 on the expression of a cohort of glucose-repressed genes and found three broad categories: genes that need both activators for full derepression, genes that rely mostly on Cat8 and genes that require only Adr1. Through combined expression and recruitment data, along with analysis of chromatin remodeling at two of these genes, ADH2 and FBP1, we clarified how these activators achieve this wide range of co-regulation. We find that Adr1 and Cat8 are not intrinsically different in their abilities to recruit coactivators but rather, promoter context appears to dictate which activator is responsible for recruitment to specific genes. These promoter-specific contributions are also apparent in the chromatin remodeling that accompanies derepression: ADH2 requires both Adr1 and Cat8, whereas, at FBP1, significant remodeling occurs with Cat8 alone. Although over-expression of Adr1 can compensate for loss of Cat8 at many genes in terms of both activation and chromatin remodeling, this over-expression cannot complement all of the cat8Delta phenotypes.

Conclusions/significance: Thus, at many of the glucose-repressed genes, Cat8 and Adr1 appear to have interchangeable roles and promoter architecture may dictate the roles of these activators.

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Recruitment profiles under derepressed conditions.A–G: ChIP analysis for coactivators/general transcription machinery at the indicated promoter regions in a wildtype strain (black), adr1Δ (pink), cat8Δ (blue) and adr1Δcat8Δ (green). Binding is expressed as the percent of the wildtype derepressed value (set to 100%) after normalization to the TEL negative control locus. Error bars represent the standard deviation of biological replicates (two or more). Data is shown at 4 hours of derepression. H: ChIP analysis for Taf1 in repressing (grey bars) and derepressing (four hours, black bars) conditions. Error bars represent technical replicates. Inset: ChIP analysis by PCR at for Taf1 at ACT1. The protein(s) assayed for in each case is listed in Table 2.
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pone-0001436-g001: Recruitment profiles under derepressed conditions.A–G: ChIP analysis for coactivators/general transcription machinery at the indicated promoter regions in a wildtype strain (black), adr1Δ (pink), cat8Δ (blue) and adr1Δcat8Δ (green). Binding is expressed as the percent of the wildtype derepressed value (set to 100%) after normalization to the TEL negative control locus. Error bars represent the standard deviation of biological replicates (two or more). Data is shown at 4 hours of derepression. H: ChIP analysis for Taf1 in repressing (grey bars) and derepressing (four hours, black bars) conditions. Error bars represent technical replicates. Inset: ChIP analysis by PCR at for Taf1 at ACT1. The protein(s) assayed for in each case is listed in Table 2.

Mentions: We first assayed binding of RNA polymerase II (Pol II). A 5-fold increase on average over the amount of pol II detected at the promoter in repressing conditions was generally observed upon derepression. The amount of pol II at each of the glucose-regulated promoters in repressing conditions was not elevated compared to a region of the telomere not expected to be bound by pol II (additional negative control regions included the coding region of the Pol I structural gene and the ADH2 ORF yielded the same results). This was true for all of the coactivators we studied (with the exception of TFIID, see below) so data was shown only for derepressed conditions. For mutant strains, the results were expressed as the percent of the factor bound compared to 100% binding in the wild-type strain, both in derepressing conditions. Pol II binding, based on three biological samples, was generally consistent with expression data (compare Fig. 1A and Table 1). The notable exception was the greater than wildtype occupancy at FBP1 in adr1Δ strain. Although Adr1 is not the dominant factor at this gene in terms of expression, it does bind to this promoter [9] and has a minor affect on expression. The increased levels of Pol II in this background may indicate that Adr1 does have a small role and that its deletion requires compensation by increased levels of coactivators.


Adr1 and Cat8 mediate coactivator recruitment and chromatin remodeling at glucose-regulated genes.

Biddick RK, Law GL, Young ET - PLoS ONE (2008)

Recruitment profiles under derepressed conditions.A–G: ChIP analysis for coactivators/general transcription machinery at the indicated promoter regions in a wildtype strain (black), adr1Δ (pink), cat8Δ (blue) and adr1Δcat8Δ (green). Binding is expressed as the percent of the wildtype derepressed value (set to 100%) after normalization to the TEL negative control locus. Error bars represent the standard deviation of biological replicates (two or more). Data is shown at 4 hours of derepression. H: ChIP analysis for Taf1 in repressing (grey bars) and derepressing (four hours, black bars) conditions. Error bars represent technical replicates. Inset: ChIP analysis by PCR at for Taf1 at ACT1. The protein(s) assayed for in each case is listed in Table 2.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2175534&req=5

pone-0001436-g001: Recruitment profiles under derepressed conditions.A–G: ChIP analysis for coactivators/general transcription machinery at the indicated promoter regions in a wildtype strain (black), adr1Δ (pink), cat8Δ (blue) and adr1Δcat8Δ (green). Binding is expressed as the percent of the wildtype derepressed value (set to 100%) after normalization to the TEL negative control locus. Error bars represent the standard deviation of biological replicates (two or more). Data is shown at 4 hours of derepression. H: ChIP analysis for Taf1 in repressing (grey bars) and derepressing (four hours, black bars) conditions. Error bars represent technical replicates. Inset: ChIP analysis by PCR at for Taf1 at ACT1. The protein(s) assayed for in each case is listed in Table 2.
Mentions: We first assayed binding of RNA polymerase II (Pol II). A 5-fold increase on average over the amount of pol II detected at the promoter in repressing conditions was generally observed upon derepression. The amount of pol II at each of the glucose-regulated promoters in repressing conditions was not elevated compared to a region of the telomere not expected to be bound by pol II (additional negative control regions included the coding region of the Pol I structural gene and the ADH2 ORF yielded the same results). This was true for all of the coactivators we studied (with the exception of TFIID, see below) so data was shown only for derepressed conditions. For mutant strains, the results were expressed as the percent of the factor bound compared to 100% binding in the wild-type strain, both in derepressing conditions. Pol II binding, based on three biological samples, was generally consistent with expression data (compare Fig. 1A and Table 1). The notable exception was the greater than wildtype occupancy at FBP1 in adr1Δ strain. Although Adr1 is not the dominant factor at this gene in terms of expression, it does bind to this promoter [9] and has a minor affect on expression. The increased levels of Pol II in this background may indicate that Adr1 does have a small role and that its deletion requires compensation by increased levels of coactivators.

Bottom Line: We determined the individual contributions of Cat8 and Adr1 on the expression of a cohort of glucose-repressed genes and found three broad categories: genes that need both activators for full derepression, genes that rely mostly on Cat8 and genes that require only Adr1.These promoter-specific contributions are also apparent in the chromatin remodeling that accompanies derepression: ADH2 requires both Adr1 and Cat8, whereas, at FBP1, significant remodeling occurs with Cat8 alone.Thus, at many of the glucose-repressed genes, Cat8 and Adr1 appear to have interchangeable roles and promoter architecture may dictate the roles of these activators.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, University of Washington, Seattle, Washington, United States of America.

ABSTRACT

Background: Adr1 and Cat8 co-regulate numerous glucose-repressed genes in S. cerevisiae, presenting a unique opportunity to explore their individual roles in coactivator recruitment, chromatin remodeling, and transcription.

Methodology/principal findings: We determined the individual contributions of Cat8 and Adr1 on the expression of a cohort of glucose-repressed genes and found three broad categories: genes that need both activators for full derepression, genes that rely mostly on Cat8 and genes that require only Adr1. Through combined expression and recruitment data, along with analysis of chromatin remodeling at two of these genes, ADH2 and FBP1, we clarified how these activators achieve this wide range of co-regulation. We find that Adr1 and Cat8 are not intrinsically different in their abilities to recruit coactivators but rather, promoter context appears to dictate which activator is responsible for recruitment to specific genes. These promoter-specific contributions are also apparent in the chromatin remodeling that accompanies derepression: ADH2 requires both Adr1 and Cat8, whereas, at FBP1, significant remodeling occurs with Cat8 alone. Although over-expression of Adr1 can compensate for loss of Cat8 at many genes in terms of both activation and chromatin remodeling, this over-expression cannot complement all of the cat8Delta phenotypes.

Conclusions/significance: Thus, at many of the glucose-repressed genes, Cat8 and Adr1 appear to have interchangeable roles and promoter architecture may dictate the roles of these activators.

Show MeSH
Related in: MedlinePlus