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Status |
Public on Dec 31, 2021 |
Title |
The DNA polymerase subunit Pol32 and RNase H1 are required for stress granule formation through R-loop regulation |
Organism |
Saccharomyces cerevisiae |
Experiment type |
Expression profiling by high throughput sequencing
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Summary |
Strong cellular stress causes wide-spread perturbations in the transcriptome and in several types of DNA/RNA interactions, and through incompletely understood pathways to formation of cytoplasmic stress granules (SGs). We have investigated the relationships between strong transcriptional induction, RNA:DNA hybrid stretches (R-loops), and SG formation under severe hyperosmotic and glucose stress. Several mutations affecting DNA processing proteins, including the DNA polymerase subunit Pol32, confer SG formation defects. Severe stress increased R loop levels globally. We found that facilitating removal of R-loops by RNase H overexpression, with activity towards RNA:DNA hybrids, accelerated and enhanced transcriptional induction of stress-activated genes. Thus, overexpression of RNase H1, but not RNase H2, reduced R-loops globally around the 1 h mark. Remarkably, it also reduced SG formation. We performed a genome-wide analysis of the induction or repression kinetics of gene expression under severe stress conditions. RNase H1 overexpression reduced R-loops locally in highly transcribed stress-affected genes, as expected. Notably, it also increased expression of several stress-induced genes. Conversely, in cells where R-loops are not efficiently resolved, transcriptional induction of the same genes under stress was muted and occurred with a delay. Thus, in pol32∆ mutants, where SG accumulation is delayed, R-loop levels are elevated, and induction of stress genes suppressed. The pol32∆ mutants are also refractory to the effects of RNase H1 overexpression on stress gene induction, R-loop resolution, and SG formation, indicating that Pol32 may act downstream of Rnh1 in the regulation of these three processes. These findings demonstrate an unexpected link between R-loops and formation of SGs. Together, these observations indicate that under stress, strong transcriptional induction of specific genes or genomic regions causes R-loop accumulation, which then requires RNase H1 activity for resolution. If unresolved, the accumulated R-loops impede continued stress-induced transcription, and delay or prevent SG formation.
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Overall design |
Samples were collected every 30 min after onset of 1.5 M KCl stress, up to 2 h. Samples were collected from two independent replicate experiment time series Total number of samples = 10. Control samples were collected at the 0 h time point.
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Contributor(s) |
Kim H, Sunnerhagen P |
Citation missing |
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Submission date |
Nov 12, 2018 |
Last update date |
Dec 31, 2021 |
Contact name |
Per Sunnerhagen |
E-mail(s) |
per.sunnerhagen@cmb.gu.se
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Phone |
+46317863830
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Organization name |
University of Gothenburg
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Department |
Chemistry and Molecular Biology
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Street address |
Box 462
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City |
Gothenburg |
ZIP/Postal code |
40530 |
Country |
Sweden |
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Platforms (1) |
GPL13821 |
Illumina HiSeq 2000 (Saccharomyces cerevisiae) |
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Samples (10)
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Relations |
BioProject |
PRJNA505169 |
SRA |
SRP168403 |
Supplementary file |
Size |
Download |
File type/resource |
GSE122423_Expression_data_Kim_et_al.csv.gz |
6.6 Mb |
(ftp)(http) |
CSV |
SRA Run Selector |
Raw data are available in SRA |
Processed data are available on Series record |
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