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| Status |
Public on Jun 01, 2012 |
| Title |
GS003: Tight cooperation between Mot1p and NC2β in regulating genome-wide transcription, repression of transcription following heat shock induction and genetic interaction with SAGA. |
| Organism |
Saccharomyces cerevisiae |
| Experiment type |
Expression profiling by array
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| Summary |
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.
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| Overall design |
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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| Contributor(s) |
Spedale G, Meddens C, Koster M, Ko CW, van Hooff SR, Holstege FC, Timmers HM, Pijnappel WP |
| Citation(s) |
21976730 |
| |
| Submission date |
Jun 22, 2011 |
| Last update date |
Mar 26, 2013 |
| Contact name |
Marian Groot Koerkamp |
| Organization name |
Princess Maxima Center for Pediatric Oncology
|
| Department |
Research
|
| Lab |
Drostlab
|
| Street address |
Heidelberglaan 25
|
| City |
Utrecht |
| State/province |
Utrecht |
| ZIP/Postal code |
3584 CS |
| Country |
Netherlands |
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| Platforms (1) |
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| Samples (12)
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| Relations |
| BioProject |
PRJNA143855 |
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