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Series GSE245521 Query DataSets for GSE245521
Status Public on Nov 07, 2023
Title Genome-wide Regulation of Pol II, FACT, and Spt6 Occupancies by RSC in Saccharomyces cerevisiae
Organism Saccharomyces cerevisiae
Experiment type Genome binding/occupancy profiling by high throughput sequencing
Summary RSC (remodels the structure of chromatin) is an essential ATP-dependent chromatin remodeling complex in Saccharomyces cerevisiae. The catalytic subunit of RSC, Sth1 uses its ATPase activity to slide or remove nucleosomes. RSC has been shown to regulate the width of the nucleosome-depleted regions (NDRs) by sliding the flanking nucleosomes away from NDRs. As such the nucleosomes encroach NDRs when RSC is depleted and leads to transcription initiation defects. In this study, we examined the effects of the catalytic-dead Sth1 on transcription and compared them to the effects observed during acute and rapid Sth1 depletion by auxin-induced degron strategy. We found that rapid depletion of Sth1 reduces recruitment of TBP and Pol II in highly transcribed genes, as would be expected considering its role in regulating chromatin structure at promoters. In contrast, cells harboring the catalytic-dead Sth1 exhibited a severe reduction in TBP binding, but surprisingly, also displayed a substantial accumulation in Pol II occupancies within coding regions. After depleting endogenous Sth1 in the catalytic dead mutant, we observed a further increase in Pol II occupancies, suggesting that the inactive Sth1 contributed to the observed accumulation of Pol II in coding regions. Notwithstanding the Pol II increase, the ORF occupancies of histone chaperones FACT and Spt6 were significantly reduced in the mutant. These results suggest a potential role for RSC in recruiting/retaining these chaperones in coding regions. Pol II accumulation despite substantial reductions in TBP, FACT, and Spt6 occupancies in the catalytic-dead mutant could be indicative of severe transcription elongation and termination defects. Such defects would be consistent with studies showing that RSC is recruited to coding regions in a transcription-dependent manner. Thus, these findings imply a role for RSC in transcription elongation and termination processes, in addition to its established role in transcription initiation.
 
Overall design The S. cerevisiae cultures spiked in with 10% of S. pombe cells were formaldehyde crosslinked and subjected to sonication to obtain chromatin. Thus obtained chromatin was subjected to chromatin immunoprecipitation using antibodies against RNA Pol II (Pol II), TATA-binding protein, and histone chaperone Spt6 and Spt16. The ChIPed DNA was subjected to paired-end sequencing.
 
Contributor(s) Govind C
Citation missing Has this study been published? Please login to update or notify GEO.
Submission date Oct 16, 2023
Last update date Nov 08, 2023
Contact name Chhabi Govind
E-mail(s) govind@oakland.edu
Organization name Oakland University
Department Biological Sciences
Street address 333 Science and Engineering Building
City Rochester
State/province mi
ZIP/Postal code 48085
Country USA
 
Platforms (1)
GPL13821 Illumina HiSeq 2000 (Saccharomyces cerevisiae)
Samples (32)
GSM7844424 Pol II ChIP_Rep1_Sth1-AID/Sth1-KR+Aux+Met [Pol II_cn5_S5]
GSM7844425 Pol II ChIP_Rep2_Sth1-AID/Sth1-KR+Aux+Met [Pol II_cn6_S6]
GSM7844426 Pol II ChIP_Rep1_Sth1-AID/Sth1-KR [Pol II_cn7_S7]
Relations
BioProject PRJNA1028656

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Supplementary file Size Download File type/resource
GSE245521_RAW.tar 1.1 Gb (http)(custom) TAR (of BW)
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Raw data are available in SRA
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