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Xrn1 influence on gene transcription results from the combination of general effects on elongating RNA pol II and gene-specific chromatin configuration [RNA-seq]

ABSTRACT: 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.

ORGANISM(S): Saccharomyces cerevisiae

PROVIDER: GSE153036 | GEO | 2021/05/24

REPOSITORIES: GEO

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Xrn1 influence on gene transcription results from the combination of general effects on elongating RNA pol II and gene-specific chromatin configuration [Mnase-seq]

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.

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The Not4 ubiquitin ligase utilizes its conserved RNA binding domains to regulate global proteostasis and RNA polymerase II-dependent transcription

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Project description:Determination of 3' or 5' intragenic RNA pol II occupancy. Maintaining the proper mRNA levels is a key aspect in the regulation of gene expression. The balance between mRNA synthesis and decay determines these levels. Using a whole-genome analysis, we demonstrate that most yeast mRNAs are degraded by the 5'-to-3' pathway (the "decaysome"), as proposed previously. Unexpectedly, the level of these mRNAs is highly robust to perturbations in this major pathway, as defects in various decaysome components lead to transcription down-regulation. Moreover, these components shuttle between the cytoplasm and the nucleus, in a manner dependent on proper mRNA degradation. In the nucleus, they associate with chromatin - preferentially ~30bp upstream of transcription start-sites - and directly stimulate transcription initiation and elongation. Hence, the decaysome has a dual role in maintaining mRNA levels. Significantly, proper import of some decaysome components seems to play a key role in coupling the two functions. The gene expression process is therefore circular, whereby the hitherto first and last stages are interconnected. This study focuses on the distribution of RNA pol II in the 3' or 5' region of 377 S. cerevisiae ORFs. S. cerevisiae cells grown in YPD to exponential phase were subjected to chromatin immunoprecipitation (ChIP) with anti-Rpb3 antibody. Data were normalized using whole cell extract hybridized on the same array. Home-made macroarrays containing 300 bp from both the 5' or 3' ends of each ORF were used.

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Distribution of total RNA polymerase II along the 5'/3' regions in xrn1 mutants

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