Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Small RNA-seq from FPLC-fractionated leaf and flower crude extracts

ABSTRACT: Background: RNA interference (RNAi) is an indispensable regulatory mechanism governing developmental processes and stress responses via sequence-specific control of target RNAs mediated by the action of small, 20–24-nt-long, non-coding regulatory (s)RNAs such as micro (mi) and small interfering (si) RNAs. Biogenesis and sorting of miRNAs into ARGONAUTE (AGO) proteins are intensively investigated, however very few information is available about the presence and distribution of distinct sRNA pools in plant cells. Results: High-throughput sequencing of size-separated sRNA pools of plant crude extracts revealed that the majority of the canonical miRNAs were associated with high molecular weight RNA-induced silencing complexes co-migrating with AGO1 (HMW RISC). In contrast, the majority of 24-nt-long siRNAs were found in association with low molecular weight complexes co-migrating with AGO4 (LMW RISC). Intriguingly, we identified a large set of sRNAs in the cytoplasm, including mature miRNA sequences, in the low molecular size range corresponding to protein-unbound sRNAs. By comparing the RISC-loaded and protein-unbound pools of miRNAs, we identified miRNAs with highly different loading efficiencies. Investigation of some selected miRNAs in a transient expression system validated this finding. We also showed that the availability of RISCs is a limiting factor determining the loading efficiency of miRNAs. Conclusion: Our data reveal the existence of a regulatory checkpoint, likely controlled by information carried by the diverse miRNA precursors, determining the RISC-loading efficiencies of various miRNAs by sorting only a subset of the produced miRNAs into the biologically active RISCs.

ORGANISM(S): Arabidopsis thaliana

PROVIDER: GSE130431 | GEO | 2019/07/29

REPOSITORIES: GEO

ACCESS DATA

Json Xml

Similar Datasets

A universal method for the rapid isolation of all known classes of functional small RNAs

Project description:Diverse classes of regulatory small (s)RNAs operate via ARGONAUTE-family proteins within RNA-induced-silencing-complexes (RISCs). Based the conserved biochemical properties intrinsic to all ARGONAUTEs, we have developed a universal, 15-min benchtop extraction procedure allowing simultaneous purification of all classes of RISC-associated sRNAs, without prior knowledge of the sample’s -intrinsic ARGONAUTE repertoires. Optimized as a user-friendly kit, the method –coined “TraPR” for Trans-kingdom, rapid, affordable Purification of RISCs– operates irrespectively of the organism, tissue, cell type or bio-fluid of interest, and scales to minute amounts of input material. The method is highly suited for direct sRNA deep-sequencing, with TraPR-generated libraries being qualitatively at least on-par with those obtained via gold-standard procedures that require immunoprecipitations and/or lengthy polyacrylamide gel excisions. TraPR considerably improves the quality and consistency of sRNA sample preparation including from notoriously difficult-to-handle tissues/bio-fluids such as starchy storage roots and mammalian plasma, and regardless of RNA contaminants or samples’ RNA-degradation status.

2021-09-08 | PXD015395 | Pride

The distribution of individual miRNA species between high-molecular weight and low-molecular weight RISC complexes by small RNA sequencing in thyroid cancer cell lines

Project description:Thyroid cancer is the most prevalent malignancy of the endocrine system. We and others have shown that several microRNAs, which are ~21-24nt post-transcriptional gene regulators, are aberrantly expressed in anaplastic thyroid cancer (ATC) and papillary thyroid cancer (PTC) tissues and cell lines. In the cell, miRNAs are bound to Argonaute (AGO) proteins as low molecular weight RNA Induced Silencing Complexes (LMW-RISCs) that can then assemble into high molecular weight RISCs (HMW-RISCs) that also include additional proteins and mRNA. In this study, we sought to analyze the association of miRNAs across RISC complexity in ATC and PTC. For both ATC and PTC lines, miRNAs were enriched in HMW-RISC and LMW-RISC fractions compared with intermediate fractions or very low molecular weight (AGO-poor) fractions. Furthermore, 60% of all miRNAs were more abundant in LMW-RISC versus HMW- RISC fractions by ~2-4 fold. Surprisingly, miR-21-5p, one of the most abundant miRNAs in both ATC and PTC lines, was consistently one the least abundant miRNAs in HMW-RISC and the most enriched miRNA in LMW-RISC fractions. These findings suggest that miR-21 may be uniquely distributed in RISCs relative to other miRNAs. Furthermore, the methodology described here is a useful way to assess the relative magnitude of miR-21association with HMW and LMW-RISCs and may help to reveal the true roles of this miRNA in thyroid cancer development, progression, and treatment.

2020-02-28 | GSE146015 | GEO

A universal, benchtop, gel-free method for the rapid and simultaneous isolation of all known classes of functional small RNAs

Project description:All known silencing small (s)RNAs operate via ARGONAUTE(AGO)-family proteins within RNA-induced-silencing-complexes (RISCs). Based on AGOs conserved biochemical properties, we have developed a universal, 15-min benchtop extraction procedure allowing simultaneous purification of all classes of RISC-associated sRNAs known to date, without prior knowledge of the samples-intrinsic AGO repertoires. Optimized into a user-friendly kit, the method –coined “TraPR” for Trans-kingdom, rapid, affordable Purification of RISCs– operates irrespectively of the organism, tissue, cell type or bio-fluid of interest, including from minute amounts of input material. The method is highly suited for direct sRNA deep-sequencing, with TrAPR-generated libraries being qualitatively and quantitatively at least on-par with those obtained via gold-standard procedures involving tedious polyacrylamide gel excisions. TraPR considerably improves the quality and consistency of sRNA sample preparation including from notoriously difficult-to-handle tissues/bio-fluids such as starchy storage roots and mammalian plasma, and regardless of RNA contaminants or samples’ RNA-degradation status.

2020-07-01 | GSE134516 | GEO

XRN2 Suppresses Aberrant Entry of tRNA Trailers into Argonaute in Humans and Arabidopsis

Project description:MicroRNAs (miRNAs) are a well-characterized class of small RNAs (sRNAs) that regulate gene expression post-transcriptionally. miRNAs function within a complex milieu of other sRNAs of similar size and abundance, with the best characterized being tRNA fragments or tRFs. The mechanism by which the RNA-induced silencing complex (RISC) selects for specific sRNAs over others is not entirely understood in human cells. Several highly expressed tRNA trailers (tRF-1s) are strikingly similar to microRNAs in length but are generally excluded from the microRNA effector pathway. This exclusion provides a paradigm for identifying mechanisms of RISC selectivity. Here, we show that 5' to 3' exoribonuclease XRN2 contributes to human RISC selectivity. Although highly abundant, tRF-1s are highly unstable and degraded by XRN2 which blocks tRF-1 accumulation in RISC. We also find that XRN mediated degradation of tRF-1s and subsequent exclusion from RISC is conserved in plants. Our findings reveal a conserved mechanism that prevents aberrant entry of a class of highly produced sRNAs into Ago2.

2023-04-24 | GSE184124 | GEO

Comprehensive Analysis of RDR1/2/6-independent Small RNAs in Arabidopsis thaliana

Project description:Plant small RNAs (sRNAs) regulate key physiological mechanisms through post-transcriptional and transcriptional silencing of gene expression. sRNAs fall into two major categories: those that are reliant on RNA Dependent RNA Polymerases (RDRs) for biogenesis and those that aren’t. Known RDR-dependent sRNAs include phased and repeat-associated short interfering RNAs, while known RDR-independent sRNAs are primarily microRNAs (miRNAs) and other hairpin-derived sRNAs. In this study, we produced and analyzed sRNA-seq libraries produced from rdr1/rdr2/rdr6 triple mutant plants. We found 58 previously annotated MIRNA loci that were dependent on RDR function, casting doubt on their classification as MIRNAs. We also found 38 RDR-independent sRNA clusters that were not MIRNAs or otherwise hairpin-derived. These 38 sRNA loci have novel biogenesis mechanisms, and frequently arise from protein-coding genes. Altogether, our analysis suggests that these 38 sRNA loci represent one or more new types of sRNA loci in Arabidopsis thaliana.

2018-04-09 | GSE105262 | GEO

Differentially expressed small RNAs in Arabidopsis galls formed by Meloidogyne javanica

Project description:Root-knot nematodes (RKNs) induce inside the vascular cylinder the giant cells (GCs) imbibed into a gall. Gene-repression in early developing GCs could be facilitated by small RNAs (sRNA) as miRNAs. 24nt-sRNAs, rasiRNAs and 21-22nt-sRNAs can also mediate epigenetic mechanisms. Three sRNA libraries from 3dpi galls and three from uninfected root segments were sequenced following Illumina-Solexa technology. Three sRNA libraries from 3dpi galls and three from uninfected root segments were sequenced

2015-11-19 | E-GEOD-71563 | biostudies-arrayexpress

Neisseria meningitides small RNA's regulate Citric acid cycle activity under Feast and Famine conditions

Project description:We identified twin small non-coding RNAs regulating tricarboxylic acid (TCA) cycle activity in Neisseria meningitidis. Expression of TCA cycle enzymes was elevated in sRNA deletion mutants. Direct interaction between sRNAs and the ribosomal entry sites on target mRNAs was demonstrated. Expression of the sRNAs was down-regulated in cells grown in poor medium with glucose as the sole carbon source but not without lrp, indicating that sRNA expression is controlled by the stringent response. N. meningitidis, over-expressing the sRNAs replicated in blood, but not in human cerebrospinal fluid. In addition, Lrp synthesis was inhibited by direct interaction between the sRNA and the 5’ UTR of lrp. sRNAs control adaptation of N. meningitidis to variation in nutrient supply in different niches of its host.

2017-03-22 | PXD000891 | Pride

Specific small-RNA signatures in the amygdala at premotor and motor stages of Parkinson’s disease revealed by deep sequencing analysis

Project description:Motivation: Most computational tools for small non-coding RNAs (sRNA) sequencing data analysis focus in microRNAs (miRNAs), overlooking other types of sRNAs that show multi-mapping hits. Here, we have developed a pipeline to non-redundantly quantify all types of sRNAs, and extract patterns of expression in biologically defined groups. We have used our tool to characterize and profile sRNAs in post-mortem brain samples of control individuals and Parkinson’s disease (PD) cases at early-premotor and late-symptomatic stages.Results: Clusters of co-expressed sRNAs mapping onto tRNAs significantly separated premotor and motor cases from controls. A similar result was obtained using a matrix of miRNAs slightly varying in sequence (isomiRs). The present framework revealed sRNA alterations at premotor stages of PD, which might reflect initial pathogenic perturbations. This tool may be useful to discover sRNA expression patterns linked to different biological conditions.

2017-04-01 | GSE97285 | GEO

Differentially expressed small RNAs in Arabidopsis galls formed by Meloidogyne javanica

2015-11-19 | GSE71563 | GEO

Comparative analysis of small RNAs released by the filarial nematode Litomosoides sigmodontis in vitro and in vivo

Project description:BACKGROUND: The release of small non-coding RNAs (sRNAs) has been reported in parasitic nematodes, trematodes and cestodes of medical and veterinary importance. However, little is known regarding the diversity and composition of sRNAs released by different lifecycle stages and the portion of sRNAs that persist in host tissues during filarial infection. This information is relevant to understanding potential roles of sRNAs in parasite-to-host communication, as well as to inform on the location within the host and time point at which they can be detected. METHODOLOGY & PRINCIPAL FINDINGS: We have used small RNA (sRNA) sequencing analysis to identify sRNAs in replicate samples of the excretory-secretory (ES) products of developmental stages of the filarial nematode Litomosoides sigmodontis in vitro and compare this to the parasite-derived sRNA detected in host tissues. We show that all L. sigmodontis developmental stages release RNAs in vitro, including ribosomal RNA fragments, 5’-derived tRNA fragments (5’-tRFs) and, to a lesser extent, microRNAs (miRNAs). The gravid adult females (gAF) produce the largest diversity and abundance of miRNAs in the ES compared to the adult males or microfilariae. Analysis of sRNAs detected in serum and macrophages from infected animals reveals that the class of parasite miRNAs are preferentially detected in vivo, compared to their low levels in the ES products, and identifies miR-92-3p and miR-71-5p as L. sigmodontis miRNAs that are stably detected in host cells in vivo. CONCLUSIONS: Our results suggest that gravid adult female worms secrete the largest diversity of extracellular sRNAs compared to adult males or microfilariae. We further show differences in the parasite sRNA biotype distribution detected in vitro versus in vivo. We identify macrophages as one reservoir for parasite sRNA during infection, and confirm the presence of parasite miRNAs and tRNAs in host serum during patent infection.

2019-11-18 | GSE140538 | GEO

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data