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Changes in genome-wide occupancy of core transcriptional regulators during stress

ABSTRACT: Organisms respond to heat stress by reprogramming gene expression. Here we show that genome-wide reprogramming involves enhanced assembly of the TFIID and SAGA regulatory pathways at heat induced genes, and disassembly of the TFIID pathway at heat-repressed genes. While TFIID and SAGA are recruited to heat-induced genes, only SAGA appears to be associated with achieving maximal induction. Mot1, an ATP-dependent inhibitor of the TATA binding protein TBP, assembles at heat-induced SAGA-regulated genes, but functions to attenuate rather than promote activation. Changes in promoter occupancy of bromodomain factor Bdf1 are tightly linked to changes in TFIID occupancy, which further supports the notion that the two work together. Bdf1 is inhibitory to a number of SAGA-regulated genes and dissociates when these genes are activated, suggesting that Bdf1 normally blocks transcription complex assembly at these genes. These linkages in reprogramming of factor occupancy at promoters provide direct evidence for two functionally distinct transcription assembly pathways, and reveal unexpected cross-talk between the pathways. Keywords = Chromatin Immunoprecipitation Keywords = Microarray Keywords = TBP Keywords: other

ORGANISM(S): Saccharomyces cerevisiae

PROVIDER: GSE1373 | GEO | 2004/11/03

SECONDARY ACCESSION(S): PRJNA90137

REPOSITORIES: GEO

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Project description:TATA-binding protein (TBP) is central to the regulation of transcription initiation. Recruitment of TBP to target genes can be positively regulated by one of two basal transcription factor complexes: SAGA or TFIID. Negative regulation of TBP promoter association can be performed by Mot1 or the NC2 complex. Recent evidence suggest that Mot1, NC2, and TBP form a DNA-dependent protein complex. Here, we compare the functions of Mot1 and NC2beta during basal and activated transcription using the anchor-away technique for conditional nuclear depletion. Genome-wide expression analysis indicates that both proteins regulate a highly similar set of genes (r2=0.8). Upregulated genes were enriched for SAGA occupancy, while downregulated genes preferred TFIID binding. Mot1p and NC2beta depletion during heat shock resulted in a failure to downregulate gene expression after initial activation, which was accompanied by increased TBP and RNA pol II promoter occupancies. Depletion of Mot1p or NC2beta displayed preferential synthetic lethality with the TBP-interaction module of SAGA. Our results suggest that Mot1 and NC2beta cooperate in vivo to regulate TBP function, and that they are involved in maintaining basal expression levels as well as in resetting gene expression after induction by stress. In this study 3 S. cerevisiae strains were grown with and without addition of rapamycin. For each experimental factor two independent colonies were inoculated in Complete Synthetic Medium (CSM) with 2% glucose. Overnight cultures were diluted to 0.3 OD600 in 50 ml medium and grown to 1 OD600. Cultures were then grown for 60 minutes in the absence or presence of rapamycin at 1ug/ml (LC laboratories). Cells were harvested by centrifugation at 4000 rpm for 3 minutes and were frozen in liquid nitrogen. Amplified cRNA samples were labeled with either cy3 or cy5 and put on microarray together with an oppositely labeled common reference sample consisting of cRNA of untreated wildtype strain.

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Changes in genome-wide occupancy of core transcriptional regulators during stress

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