Sequence determinants of dsRNA processing by DICER
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA836488 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR19143000.fastq.gz | Fastqsanger.gz | |||
| SRR19143001.fastq.gz | Fastqsanger.gz | |||
| SRR19143014.fastq.gz | Fastqsanger.gz | |||
| SRR19143015.fastq.gz | Fastqsanger.gz | |||
| SRR19143016.fastq.gz | Fastqsanger.gz |
Items per page: 5 1 - 5 of 30 |
Similar Datasets
Project description:Sequence determinants of dsRNA processing by DICER [Rescue Experiment]
| PRJNA836498 | ENA
Project description:Sequence determinants of dsRNA processing by DICER [Massively Parallel Assay]
| PRJNA836500 | ENA
Project description:RNA silencing relies on specific and efficient processing of dsRNA by DICER which produces microRNAs (miRNAs) and small interfering RNAs (siRNAs). However, our current knowledge of DICER’s specificity is restricted to the secondary structures of its substrates: a dsRNA than 22 bp with a 2-nt 3′ overhang. We recently found evidence pointing to additional sequence-dependent determinant(s) beyond these features. To systematically interrogate the features of precursor miRNAs (pre-miRNAs), we carried out massively parallel assays with over a million pre-miRNA variants. Our analyses revealed a highly conserved cis-acting element, termed the “GYM” motif (paired G, paired pyrimidine, and mismatched C or A) at the cleavage site, which strongly promotes processing at a specific position. We find that the C-terminal double-stranded RNA binding domain (dsRBD) of DICER recognizes the GYM motif. Mutation of the dsRBD alters pre-miRNA cleavage sites and impairs processing in a motif-dependent fashion, which in turn affects the miRNA repertoire in cells. Consistently, integrating the GYM motif into short hairpin RNA (shRNA) or Dicer substrate siRNA (DsiRNA) potentiates RNA interference.
2022-12-16 | GSE202533 | GEO
Project description:RNA silencing relies on specific and efficient processing of dsRNA by DICER which produces microRNAs (miRNAs) and small interfering RNAs (siRNAs). However, our current knowledge of DICER’s specificity is restricted to the secondary structures of its substrates: a dsRNA than 22 bp with a 2-nt 3′ overhang. We recently found evidence pointing to additional sequence-dependent determinant(s) beyond these features. To systematically interrogate the features of precursor miRNAs (pre-miRNAs), we carried out massively parallel assays with over a million pre-miRNA variants. Our analyses revealed a highly conserved cis-acting element, termed the “GYM” motif (paired G, paired pyrimidine, and mismatched C or A) at the cleavage site, which strongly promotes processing at a specific position. We find that the C-terminal double-stranded RNA binding domain (dsRBD) of DICER recognizes the GYM motif. Mutation of the dsRBD alters pre-miRNA cleavage sites and impairs processing in a motif-dependent fashion, which in turn affects the miRNA repertoire in cells. Consistently, integrating the GYM motif into short hairpin RNA (shRNA) or Dicer substrate siRNA (DsiRNA) potentiates RNA interference.
2022-12-16 | GSE202530 | GEO
Project description:Small interfering RNAs (siRNAs) are the key components for RNA interference (RNAi), a conserved RNA silencing mechanism in many eukaryotes. In Drosophila, an RNase III enzyme Dicer-2 (Dcr-2), aided by its cofactor Loquacious-PD (Loqs-PD), plays major role in generating 21 base-pair (bp) siRNA duplexes from long double-stranded RNAs (dsRNAs). The ATP hydrolysis by the helicase domain of Dcr-2 is critical to the successful processing of a long dsRNA into consecutive siRNA duplexes. Here we report the cryo-EM structures of Dcr-2/Loqs-PD in the apo state and in multiple processing states of a 50-bp dsRNA substrate. The structures elucidated interactions between Dcr-2 and Loqs-PD, and significant conformational changes of Dcr-2 during a dsRNA processing cycle. The N-terminal helicase and DUF283 domains undergo conformational change upon initial dsRNA binding, forming an ATP binding pocket and a 5’-phosphate binding pocket. The overall conformation of Dcr-2/Loqs-PD is relatively rigid during translocating along the dsRNA in the presence of ATP, while the interaction between DUF283 and RIIIDb domains prevents non-specific cleavage during translocation by blocking the access of dsRNA to the RNase active center. Additional ATP-dependent conformational changes are required to form an active-dicing state and precisely cleave the dsRNA into 21-bp siRNA duplex confirmed by the structure in the post-dicing state. Collectively, this study revealed the molecular mechanism for the full-cycle of ATP-dependent dsRNA processing by Dcr-2/Loqs-PD.
2022-04-21 | PXD033365 |
Project description:This SuperSeries is composed of the SubSeries listed below.
2022-12-16 | GSE202535 | GEO
Project description:A hallmark of RNA silencing is a class of ~22 nt RNAs which are processed from dsRNA precursor by Dicer. Accurate processing by Dicer is critical for the functionality of microRNAs (miRNAs). According to the current model, Dicer measures the length by anchoring the 3' overhang of the dsRNA terminus. Here we find that human Dicer binds to the 5' end of RNA and utilizes the 5' end as an additional reference point for cleavage site selection (5' counting rule). We further identify a novel motif (5'-pocket) in Dicer, which recognizes the 5' end of RNA. By analyzing miRNA population from 5'-pocket mutant Dicer expressing Dicer-null mES, we provide that the 5' pocket is significant for Dicer processing in vivo.
2011-07-13 | GSE27903 | GEO
Project description:Although many in vitro studies have helped us understand how Dicer-2 is able to discriminate between different dsRNA substrate termini, much less is known about how this translates to the in vivo recognition of viral dsRNA. Indeed, Dicer-2 associates with several dsRNA-binding proteins (dsRBPs), which can modify its specificity for a substrate, however it remains unknown how Dicer-2 is able to recognize the protected termini of viral dsRNAs. Therefore, in order to study how the ribonucleoprotein network of Dicer-2 impacts antiviral immunity, we used an IP-MS approach to identify interactants of several fly lines expressing different versions of GFP:Dicer-2. By combining the global analysis of the Dicer-2 interactome with different line-specific analyses, we were able to both obtain a global overview of the partners of Dicer-2 in vivo, and study how this interactome was modulated by different factors such as the infection and/or the presence of different point mutations on the helicase or RNase III domains of GFP:Dicer-2. This allowed the identification of several new Dicer-2 interactants as well as new pro- and antiviral factors that had an impact on DCV infection. In addition, this work provides a resource composed of several candidates, available to the scientific community that can now be investigated further to gain a better understanding of the proteins involved in Dicer-2-mediated antiviral RNAi.
2025-04-15 | PXD038898 | Pride
Project description:A hallmark of RNA silencing is a class of ~22 nt RNAs which are processed from dsRNA precursor by Dicer. Accurate processing by Dicer is critical for the functionality of microRNAs (miRNAs). According to the current model, Dicer measures the length by anchoring the 3' overhang of the dsRNA terminus. Here we find that human Dicer binds to the 5' end of RNA and utilizes the 5' end as an additional reference point for cleavage site selection (5' counting rule). We further identify a novel motif (5'-pocket) in Dicer, which recognizes the 5' end of RNA. By analyzing miRNA population from 5'-pocket mutant Dicer expressing Dicer-null mES, we provide that the 5' pocket is significant for Dicer processing in vivo. Examination of small RNA profiles from Dicer-null mouse embryonic stem cells transfected with either wild-type or 5' pocket mutant Dice expression plasmids.
2011-07-12 | E-GEOD-27903 | biostudies-arrayexpress
Project description:5’-isomiRs expand the repertoire of miRNA targets. However, how they are generated is not well understood. Here, we showed that, for some miRNAs in mammalian cells, 5’-isomiRs are generated by alternative Dicer processing by using miRNA offset RNAs (moRs) to determine Drosha cleavage sites in cells. In addition, we showed that in miR-203, alternative Dicer processing is regulated by a conserved sliding-bulge structure at the Dicer processing site, which allows the pre-miRNA molecule to fold into two different structures that are processed differently by Dicer. So far no RNA motif that slides to change conformation and alter a protein–RNA interaction has been reported. Thus, our study revealed a novel RNA motif that regulates 5’-isomiR generation in some miRNAs. It might also contribute to regulating protein–RNA interactions in other biological processes, since it takes only one point mutation to generate the sliding bulge, and there are a large number of different RNAs in the cell.
2016-09-01 | GSE80772 | GEO