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Series GSE8862 Query DataSets for GSE8862
Status Public on Dec 01, 2007
Title RSC Regulates Nucleosome Positioning at Pol II Genes and Density at Pol III Genes
Organism Saccharomyces cerevisiae
Experiment type Genome binding/occupancy profiling by genome tiling array
Summary Nucleosomes restrict the access of transcription factors to chromatin. RSC is a SWI/SNF-family chromatin-remodeling complex from yeast that repositions and ejects nucleosomes in vitro. Here, we examined these activities and their importance in vivo. We utilized array-based methods to examine nucleosome occupancy and positioning at more than 200 locations in the genome following the controlled destruction of the catalytic subunit of RSC, Sth1. Loss of RSC function caused pronounced and general reductions in transcription from Pol I, II, and III genes. At Pol III genes, Sth1 loss conferred a general gain in nucleosome density and an accompanying reduction in RNA Pol III occupancy. In contrast, we observed primarily single nucleosome changes, including movement, at Pol II promoters. Importantly, a greater number of changes were observed near the transcription start sites of RSC-occupied promoters than non-occupied promoters. These changes are distinct from those due to general loss of transcription. Thus, RSC action affects both nucleosome density and positioning in vivo, but applies these remodeling modes differently at Pol II and Pol III genes.
Keywords: ChIP-chip, nucleosome, mononucleosome, RSC, transcription
 
Overall design To examine chromatin remodeling that was dependent upon RSC, we generated a customized microarray platform that focused on known RSC targets. This array (GPL5637) sampled 218 specific loci in the genome at relatively high resolution to monitor nucleosome occupancy. We first confirmed the design of the array by performing ChIP against two subunits of RSC. Since the Sth1 degron allele used in our studies is degraded under specific growth conditions, we also performed RSC ChIP under mock degradation conditions (mock-after). We next analyzed the chromatin structure of RSC target genes in the absence of RSC function. In these experiments, we compared microarray results between two strains: the Degron strain, which allows for RSC inactivation, and the Control strain, which prevents RSC inactivation. We analyzed chromatin structure using two approaches. First, general histone occupancy was examined by performing ChIP against histone H4. Second, we hybridized DNA from mononucleosomes that were released by micrococcal nuclease digestion to examine single nucleosome events, including movement, gain, and loss. We also examined the occupancy of RNA Polymerase III by ChIP. Finally, we examined single nucleosome occupancies by mononucleosome hybridization in a RNA Polymerase II mutant, rpb1-1, at both permissive and non-permissive temperatures.
 
Contributor(s) Parnell TJ, Huff JT, Cairns BR
Citation(s) 18059476
Submission date Aug 23, 2007
Last update date Sep 19, 2012
Contact name Timothy J Parnell
E-mail(s) timothy.parnell@hci.utah.edu
Organization name Huntsman Cancer Institute
Street address 2000 Circle of Hope
City Salt Lake City
State/province UT
ZIP/Postal code 84112
Country USA
 
Platforms (1)
GPL5637 Saccharomyces cerevisiae RSC Specific 22K 8pack
Samples (50)
GSM224551 Rsc3-ChIP_Mock-Before_replicate1
GSM224552 Rsc3-ChIP_Mock-Before_replicate2
GSM224553 Rsc3-ChIP_Mock-Before_replicate3
Relations
BioProject PRJNA102223

Download family Format
SOFT formatted family file(s) SOFTHelp
MINiML formatted family file(s) MINiMLHelp
Series Matrix File(s) TXTHelp

Supplementary file Size Download File type/resource
GSE8862_RAW.tar 23.4 Mb (http)(custom) TAR (of TXT)
Processed data included within Sample table
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