Mapping of dsRNA in yeast using reconstituted RNAi pathway
Ontology highlight
ABSTRACT:
ORGANISM(S): Saccharomyces cerevisiae
PROVIDER: GSE64090 | GEO | 2015/08/11
SECONDARY ACCESSION(S): PRJNA270085
REPOSITORIES: GEO
Similar Datasets
Project description:Small RNA produced by Dicer (Dcr1) are used to map dsRNA in wild-type strain and a xrn1-delta mutant of S. cerevisiae, inactivated for the cytoplasmic 5'-3' RNA decay pathway. Small RNA sequencing in wild-type and xrn1-delta strains of S. cerevisiae, with or without reconstituted RNAi pathway.
2015-08-11 | E-GEOD-64090 | biostudies-arrayexpress
Project description:Antisense (as)lncRNAs are extensively degraded by the nuclear exosome and the cytoplasmic exoribonuclease Xrn1 in the budding yeast Saccharomyces cerevisiae, lacking RNA interference (RNAi). Whether the ribonuclease III Dicer affects aslncRNAs in close RNAi-capable relatives remains unknown. Using genome-wide RNA profiling, here we show that aslncRNAs are primarily targeted by the exosome and Xrn1 in the RNAi-capable budding yeast Naumovozyma castellii, Dicer only affecting Xrn1-sensitive lncRNAs (XUTs) levels in Xrn1-deficient cells. The dcr1 and xrn1 mutants display synergic growth defects, indicating that Dicer becomes critical in the absence of Xrn1. Small RNA sequencing showed that Dicer processes aslncRNAs into small RNAs, with a preference for asXUTs. Consistently, Dicer localizes into the cytoplasm. Finally, we observed an expansion of the exosome-sensitive antisense transcriptome in N. castellii compared to S. cerevisiae, suggesting that the presence of cytoplasmic RNAi has reinforced the nuclear RNA surveillance machinery to temper aslncRNAs expression. Our data provide fundamental insights into aslncRNAs metabolism and open perspectives into the possible evolutionary contribution of RNAi in shaping the aslncRNAs transcriptome.
2019-08-21 | GSE129269 | GEO
Project description:Antisense (as)lncRNAs are extensively degraded by the nuclear exosome and the cytoplasmic exoribonuclease Xrn1 in the budding yeast Saccharomyces cerevisiae, lacking RNA interference (RNAi). Whether the ribonuclease III Dicer affects aslncRNAs in close RNAi-capable relatives remains unknown. Using genome-wide RNA profiling, here we show that aslncRNAs are primarily targeted by the exosome and Xrn1 in the RNAi-capable budding yeast Naumovozyma castellii, Dicer only affecting Xrn1-sensitive lncRNAs (XUTs) levels in Xrn1-deficient cells. The dcr1 and xrn1 mutants display synergic growth defects, indicating that Dicer becomes critical in the absence of Xrn1. Small RNA sequencing showed that Dicer processes aslncRNAs into small RNAs, with a preference for asXUTs. Consistently, Dicer localizes into the cytoplasm. Finally, we observed an expansion of the exosome-sensitive antisense transcriptome in N. castellii compared to S. cerevisiae, suggesting that the presence of cytoplasmic RNAi has reinforced the nuclear RNA surveillance machinery to temper aslncRNAs expression. Our data provide fundamental insights into aslncRNAs metabolism and open perspectives into the possible evolutionary contribution of RNAi in shaping the aslncRNAs transcriptome.
2019-08-21 | GSE129232 | GEO
Project description:Antisense (as)lncRNAs are extensively degraded by the nuclear exosome and the cytoplasmic exoribonuclease Xrn1 in the budding yeast Saccharomyces cerevisiae, lacking RNA interference (RNAi). Whether the ribonuclease III Dicer affects aslncRNAs in close RNAi-capable relatives remains unknown. Using genome-wide RNA profiling, here we show that aslncRNAs are primarily targeted by the exosome and Xrn1 in the RNAi-capable budding yeast Naumovozyma castellii, Dicer only affecting Xrn1-sensitive lncRNAs (XUTs) levels in Xrn1-deficient cells. The dcr1 and xrn1 mutants display synergic growth defects, indicating that Dicer becomes critical in absence of Xrn1. Small RNA sequencing showed that Dicer processes aslncRNAs into small RNAs, with a preference for asXUTs. Consistently, Dicer localizes into the cytoplasm. Finally, we observed an expansion of the exosome-sensitive antisense transcriptome in N. castellii compared to S. cerevisiae, suggesting that the presence of cytoplasmic RNAi has reinforced the nuclear RNA surveillance machinery to temper aslncRNAs expression. Our data provide fundamental insights into aslncRNAs metabolism and open perspectives into the possible evolutionary contribution of RNAi in shaping the aslncRNAs transcriptome.
2019-08-21 | GSE90041 | GEO
Project description:Using a system expressing dsRNA molecules and luciferase reporters, we explored factors contributing to RNAi inefficiency in mouse embryonic stem cells (ESCs) and NIH3T3 fibroblasts that could explain previous contradictions concerning mammalian RNAi. Low and poorly processive Dicer activity combined with insufficiently abundant substrates appear to be the primary reason for RNAi inefficiency.
2019-02-10 | GSE126324 | GEO
Project description:To investigate the role of cytoplasmic helicases in the decay of Xrn1-sensitive lncRNAs, we performed RNA-Seq in WT, ecm32-delta, ski2-delta, slh1-delta, dbp1-delta and dhh1-delta yeast cells.
2023-07-27 | GSE235553 | GEO
Project description:The Dicer ribonuclease III (RNase III) enzymes process long double-stranded RNA (dsRNA) into the small interfering RNAs (siRNAs) that direct RNA interference. Here, we describe the structure and activity of a catalytically active fragment of Kluyveromyces polysporus Dcr1, which represents the noncanonical Dicers found in budding yeast. The crystal structure reveals a homodimer that resembles bacterial RNase III but includes a novel N-terminal domain and newly identified catalytic residues conserved throughout eukaryotic RNase III enzymes. Biochemical analyses show that Dcr1 dimers bind cooperatively along the dsRNA substrate and cleave at precise intervals based on the distance between consecutive active sites. Thus, unlike canonical Dicers, which successively remove siRNA duplexes from the dsRNA termini, Dcr1 initiates processing in the interior and works outward. The distinct mechanism of budding-yeast Dicers establishes a novel paradigm for natural protein-based molecular rulers and imparts substrate preferences with ramifications for biological function. High-throughput sequencing of small RNAs generated by in vitro processing of long dsRNA using Kluyveromyces polysporus Dicer
2011-05-11 | E-GEOD-29168 | biostudies-arrayexpress
Project description:The Dicer ribonuclease III (RNase III) enzymes process long double-stranded RNA (dsRNA) into the small interfering RNAs (siRNAs) that direct RNA interference. Here, we describe the structure and activity of a catalytically active fragment of Kluyveromyces polysporus Dcr1, which represents the noncanonical Dicers found in budding yeast. The crystal structure reveals a homodimer that resembles bacterial RNase III but includes a novel N-terminal domain and newly identified catalytic residues conserved throughout eukaryotic RNase III enzymes. Biochemical analyses show that Dcr1 dimers bind cooperatively along the dsRNA substrate and cleave at precise intervals based on the distance between consecutive active sites. Thus, unlike canonical Dicers, which successively remove siRNA duplexes from the dsRNA termini, Dcr1 initiates processing in the interior and works outward. The distinct mechanism of budding-yeast Dicers establishes a novel paradigm for natural protein-based molecular rulers and imparts substrate preferences with ramifications for biological function.
2011-05-11 | GSE29168 | GEO
Project description:Background: RNA silencing pathways play critical roles in gene regulation, virus infection, and transposon control. RNA interference (RNAi) is mediated by small interfering RNAs (siRNAs), which are liberated from double stranded (ds) RNA precursors by Dicer and direct the RNA-induced silencing complex (RISC) to target transcripts. Recent efforts have uncovered important principles governing small RNA (smRNA) sorting into RISC, yet mechanisms defining substrate selection by Dicer proteins remain uncharacterized. Methodology: To better characterize Dicer-2 substrates in Drosophila, we examined the antiviral RNAi response, which generates virus-derived siRNAs from viral RNA. Using high-throughput sequencing, we found that diverse viruses were uniquely targeted; substrates included dsRNA replication intermediates and intramolecular RNA stem loops. smRNA distribution patterns from viral and synthetic dsRNA precursors were highly reproducible, and machine learning techniques identified characteristics of precursor molecules and smRNA duplexes important in determining relative smRNA abundance. Significance: To our knowledge, this study provides the first description of the rules governing Dicer-2 substrate selection, which has important implications for exogenous RNA silencing technologies and the development of smRNA-based antiviral therapeutics. virus-derived siRNA (vsiRNA) expression comparison between control and 4 different virus-infected cells in control as well as 5 different RNAi pathway protein knock-downs in Drosophila dl1 cells
2013-01-31 | E-GEOD-28957 | biostudies-arrayexpress
Project description:Sequence determinants of dsRNA processing by DICER
| PRJNA836488 | ENA