The mRNA degradation factor Xrn1 regulates transcription elongation in parallel to Ccr4
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PROVIDER: PRJNA524296 | ENA |
REPOSITORIES: ENA
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Project description:During the last decade several examples of coordination between gene transcription and mRNA degradation have been reported. mRNA imprinting by Rpb4 and 7 subunits of RNA polymerase II (RNAPII) and by the Ccr4-Not complex allows controlling its fate during transcription. Transcription regulation by mRNA degradation factors like Xrn1 constitutes a feedback loop that contributes to mRNA homeostasis. Mechanistic details of these phenomena are unclear. Most studies involve measurement of mRNA decay rates, usually by stressing procedures such as transcriptional shut-off or incorporation of modified nucleotides that can lead to biased results. In this work we have used the easily repressible yeast GAL1 gene to perform a genetic analysis of mRNA synthesis and degradation under physiological conditions. We combined this experimental approach with computational multi-agent modelling, testing different possibilities of Xrn1 and Ccr4-Not action in gene transcription. This double strategy brought us to conclude that Xrn1 regulates RNAPII backtracking in a Ccr4-independent manner. We validated this conclusion measuring TFIIS genome-wide recruitment to elongating RNAPII molecules. We found that xrn1∆ and ccr4∆ exhibited very different patterns of TFIIS/RNPAII which confirmed their differential role in controlling transcription elongation.
2019-07-17 | GSE127179 | GEO
Project description: Not available
| S-EPMC6765136 | biostudies-literature
Project description:mRNA homeostasis is favored by crosstalk between transcription and degradation machineries. Both the Ccr4-Not and the Xrn1-decaysome complexes have been described to influence transcription. While Ccr4-Not has been shown to directly stimulate transcription elongation, the information available on how Xrn1 influences transcription is scarce and contradictory. In this study we have addressed this issue by mapping RNA polymerase II (RNA pol II) at high resolution, using CRAC and BioGRO-seq techniques in Saccharomyces cerevisiae. We found significant effects of Xrn1 perturbation on RNA pol II profiles across the genome. RNA pol II profiles at 5’ exhibited significant reductions in slope that were compatible with decreased elongation rates. This phenomenon was also observed soon after the nuclear pool of Xrn1 was depleted. Nucleosome mapping confirmed drastically altered chromatin dynamics. We also found an accumulation of arrested RNA pol II at the 3’ end, immediately upstream of polyadenylation sites of most genes, with particular incidence in those functionally related to regulatory processes. Lack of Xrn1 provoked alterations in RNA pol II CTD phosphorylation and increased recruitment of 3’ pre-mRNA processing factors. However, accumulation of RNA pol II at 3’ ends was rather related to the nucleosomal configuration of those regions.
2021-05-24 | GSE153036 | GEO
Project description:mRNA homeostasis is favored by crosstalk between transcription and degradation machineries. Both the Ccr4-Not and the Xrn1-decaysome complexes have been described to influence transcription. While Ccr4-Not has been shown to directly stimulate transcription elongation, the information available on how Xrn1 influences transcription is scarce and contradictory. In this study we have addressed this issue by mapping RNA polymerase II (RNA pol II) at high resolution, using CRAC and BioGRO-seq techniques in Saccharomyces cerevisiae. We found significant effects of Xrn1 perturbation on RNA pol II profiles across the genome. RNA pol II profiles at 5’ exhibited significant reductions in slope that were compatible with decreased elongation rates. This phenomenon was also observed soon after the nuclear pool of Xrn1 was depleted. Nucleosome mapping confirmed drastically altered chromatin dynamics. We also found an accumulation of arrested RNA pol II at the 3’ end, immediately upstream of polyadenylation sites of most genes, with particular incidence in those functionally related to regulatory processes. Lack of Xrn1 provoked alterations in RNA pol II CTD phosphorylation and increased recruitment of 3’ pre-mRNA processing factors. However, accumulation of RNA pol II at 3’ ends was rather related to the nucleosomal configuration of those regions.
2021-05-24 | GSE153035 | GEO
Project description:The rates of mRNA synthesis and degradation determine cellular mRNA levels and can be monitored by comparative Dynamic Transcriptome Analysis (cDTA) that uses non-perturbing metabolic RNA labeling. Here we present cDTA data for 46 yeast strains lacking genes involved in mRNA degradation and metabolism. In these strains, changes in mRNA degradation rates are generally compensated by changes in mRNA synthesis rates, resulting in a buffering of mRNA levels. We show that buffering of mRNA levels requires the RNA exonuclease Xrn1. The buffering is rapidly established when mRNA synthesis is impaired, but is delayed when mRNA degradation is impaired, apparently due to Xrn1-dependent transcription repressor induction. Cluster analysis of the data defines the general mRNA degradation machinery, reveals different substrate preferences for the two mRNA deadenylase complexes Ccr4-Not and Pan2-Pan3, and unveils an interwoven cellular mRNA surveillance network. For this purpose, cDTA was carried out as described in Sun et.al. Genome Res. 2012 (DOI:10.1101/gr.130161.111). S. cerevisiae cultures were grown at 30¡ãC in 50 ml aliquots of YPD medium. S. pombe cultures were grown at 32¡ãC in YES medium. For inhibitor perturbation, BY4741 cells (OD600=0.1) were inoculated from a saturated overnight culture in two 100 ml aliquots of YPD liquid medium, incubated at 30¡ãC and grown to early-log phase (OD600=0.8). Cells were labeled for 6 minutes (sample 0 min). Then cycloheximide was added and cells were labeled for 10 minutes and 60 minutes. Cells were harvested and treated as described in Sun et.al. Genome Res. 2012 (DOI:10.1101/gr.130161.111). 1,10-Phenanthroline was added to the culture to a final concentration of 25 _g/ml as described (Dori-Bachash et al., 2012), and labeled for 6 minutes after 18 minutes treatment. Cells were treated as above. For the anchor-away analysis, S. cerevisiae cells (OD600=0.1) were cultured in two 200 ml aliquots of YPD containing 1_g/ml rapamycin and incubated at 30¡ãC untill OD600 reached 0.8. Due to the instability of rapamycin, we added 1_g/ml rapamycin every two hours. Then a 20 ml sample was taken and labeled with 4sU. The cells were then treated as described in Sun et.al. Genome Res. 2012 (DOI:10.1101/gr.130161.111).
2013-10-18 | E-MTAB-1525 | biostudies-arrayexpress
Project description:Xrn1 is a cytoplasmic 5’-3’ exoribonuclease responsible for degradation of multiple types of RNA. Data obtained on yeast model described Xrn1 function in degradation of messenger (m)RNA, Rapid transfer (t)RNA Decay (RTD) of hypomodified and unstable tRNA transcripts as well as maturation of 25S ribosomal (r)RNA and small nucleolar (sno)RNAs. Our group discovered previously that deletion of Xrn1 gene results in stabilization of a group of long non-coding RNAs which are often transcribed antisense to protein-coding genes and may regulate their expression on transcriptional level. However much less is known about human (h)Xrn1 function. Data available from human cell lines confirm involvement of hXrn1 in degradation of mRNA, initiator tRNA methionine and some miRNA. The high sequence similarity of Xrn1 between yeast and man suggests that also its function should be well conserved for hXrn1. Therefore, in this experiment using inducible shRNA we want to define hXrn1 targets by analyzing RNAs that are stabilized after Xrn1 silencing with focus on both coding and non-coding transcripts.
2020-01-14 | GSE114605 | GEO
Project description:To determine the effects of inactivation of both the nosense-mediated mRNA decay pathway and the general 5' to 3' decay pathway on yeast mRNA decay, we compared the expression profiles of the wild-type, xrn1, xrn1 upf1, xrn1 nmd2, and xrn1 upf3 strains.
2011-11-08 | E-MEXP-27 | biostudies-arrayexpress
Project description:Control of messenger RNA (mRNA) decay rate is intimately connected to translation elongation, but the spatial coordination of these events is poorly understood. The Ccr4-Not complex initiates mRNA decay through deadenylation and activation of decapping. We used a combination of cryo–electron microscopy, ribosome profiling, and mRNA stability assays to examine the recruitment of Ccr4-Not to the ribosome via specific interaction of the Not5 subunit with the ribosomal E-site in Saccharomyces cerevisiae. This interaction occurred when the ribosome lacked accommodated A-site transfer RNA, indicative of low codon optimality. Loss of the interaction resulted in the inability of the mRNA degradation machinery to sense codon optimality. Our findings elucidate a physical link between the Ccr4-Not complex and the ribosome and provide mechanistic insight into the coupling of decoding efficiency with mRNA stability.
2020-04-16 | GSE144250 | GEO
Project description:Control of messenger RNA (mRNA) decay rate is intimately connected to translation elongation, but the spatial coordination of these events is poorly understood. The Ccr4-Not complex initiates mRNA decay through deadenylation and activation of decapping. We used a combination of cryo–electron microscopy, ribosome profiling, and mRNA stability assays to examine the recruitment of Ccr4-Not to the ribosome via specific interaction of the Not5 subunit with the ribosomal E-site in Saccharomyces cerevisiae. This interaction occurred when the ribosome lacked accommodated A-site transfer RNA, indicative of low codon optimality. Loss of the interaction resulted in the inability of the mRNA degradation machinery to sense codon optimality. Our findings elucidate a physical link between the Ccr4-Not complex and the ribosome and provide mechanistic insight into the coupling of decoding efficiency with mRNA stability.
2020-04-16 | GSE131408 | GEO
Project description:To define the translational landscape of Xrn1-sensitive lncRNAs in yeast, we performed Ribo-Seq in WT and upf1 mutant cells, in native conditions or upon treatment with translation elongation inhibitor (cycloheximide).
2024-02-20 | GSE203278 | GEO