Dicer-independent RNA-directed DNA methylation in Arabidopsis [small RNA-seq 1]
Ontology highlight
ABSTRACT:
ORGANISM(S): Arabidopsis thaliana
PROVIDER: GSE70913 | GEO | 2016/03/17
SECONDARY ACCESSION(S): PRJNA289906
REPOSITORIES: GEO
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Project description:In Arabidopsis thaliana, four different DICER-LIKE (DCL) proteins have distinct, but partially overlapping functions in the biogenesis of microRNAs (miRNAs) and small interfering RNAs (siRNAs) from longer, non-coding precursor RNAs. To analyze the impact of different components of the small RNA (sRNA) biogenesis machinery on the transcriptome, we subjected dcl and other mutants impaired in sRNA biogenesis to whole-genome tiling array analysis. We compared both protein-coding genes and noncoding transcripts, including most pri-miRNAs, in two tissues and several stress conditions. We discovered distinct effects of dcl1, hyl1 and se mutations on the transcriptome, as well as a number of common genes affected in dcl1 and dcl2 dcl3 dcl4 triple mutants. Our results furthermore suggest that the DCL1 is not only involved in miRNA action, but can also contribute to silencing of certain transposons, apparently through an effect on DNA methylation. Together, our findings contribute to the knowledge of both specialization and overlap between different RNA silencing pathways.
2010-05-08 | GSE21685 | GEO
Project description: Not available
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Project description:In Arabidopsis thaliana, four different DICER-LIKE (DCL) proteins have distinct, but partially overlapping functions in the biogenesis of microRNAs (miRNAs) and small interfering RNAs (siRNAs) from longer, non-coding precursor RNAs. To analyze the impact of different components of the small RNA (sRNA) biogenesis machinery on the transcriptome, we subjected dcl and other mutants impaired in sRNA biogenesis to whole-genome tiling array analysis. We compared both protein-coding genes and noncoding transcripts, including most pri-miRNAs, in two tissues and several stress conditions. We discovered distinct effects of dcl1, hyl1 and se mutations on the transcriptome, as well as a number of common genes affected in dcl1 and dcl2 dcl3 dcl4 triple mutants. Our results furthermore suggest that the DCL1 is not only involved in miRNA action, but can also contribute to silencing of certain transposons, apparently through an effect on DNA methylation. Together, our findings contribute to the knowledge of both specialization and overlap between different RNA silencing pathways. Wild type and dcl1-100, se-3, hyl1-2, rdr6-15 ans dcl2,3,4 mutants were grown on MS or soil at 23M-BM-0C and LL. 7 day old seedlings (grown on MS plates) and inflorescences from 28 (>40) day old plants (dcl1) were used for RNA extraction. For stress treatments, seedlings had been grown for 10 days on solid MS medium at 21M-BM-0C, they were transferred to liquid MS medium containing no additives (mock control), 200 mM NaCl (salt stress), 300 mM mannitol (osmotic stress) or 100 M-NM-<M ABA. For cold and heat stress, seedlings were transferred to pre-chilled or pre-warmed liquid MS medium and incubated at 8M-BM-0C and 30M-BM-0C, respectively. Samples were taken after 1 h and 12 h of continuous stress treatment. RNA was extracted from whole seedlings. All RNA samples were converted into double stranded DNA targets that were hybridized to whole genome tiling arrays (Affymetrix Arabidopis Tiling1.0RM-BM-.). All hybridizations were performed with 3 biological replicates.
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Project description:Here we describe the identification and regulation of a novel dsRNA virus in Colletotrichum higginsianum. High throughput sequencing of small RNAs and strand-specific RNA-seq was performed on single gene knock-out mutants created for each RNAi component gene: rdr1, rdr2, rdr3, dcl1, dcl2, ago1, and ago2, and the double mutant: ∆dcl1∆dcl2. De novo assembly of the ∆dcl1 RNA-seq data identified two contigs that represented the forward and reverse strands of an uncharacterized dsRNA virus, designated here as Colletotrichum higginsianum non-segmented dsRNA virus 1 (ChNRV1). We found increased presence of the viral RNA in the RNA-seq datasets of the ∆dcl1, ∆dcl1dcl2, and ∆ago1 strains, suggesting that these genes are required for control of the virus. We show that viral small RNAs co-immunoprecipitate with a 6xFLAG-3xHIS-tagged AGO1 protein by sequencing the small RNAs from immunoprecipitated fractions. Additionally, analyses of the small RNA datasets from the RNAi mutants revealed control of the virus through small RNA-mediated silencing required both AGO1 and DCL1.
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Project description:Here we describe the identification and regulation of a novel dsRNA virus in Colletotrichum higginsianum. High throughput sequencing of small RNAs and strand-specific RNA-seq was performed on single gene knock-out mutants created for each RNAi component gene: rdr1, rdr2, rdr3, dcl1, dcl2, ago1, and ago2, and the double mutant: ∆dcl1∆dcl2. De novo assembly of the ∆dcl1 RNA-seq data identified two contigs that represented the forward and reverse strands of an uncharacterized dsRNA virus, designated here as Colletotrichum higginsianum non-segmented dsRNA virus 1 (ChNRV1). We found increased presence of the viral RNA in the RNA-seq datasets of the ∆dcl1, ∆dcl1dcl2, and ∆ago1 strains, suggesting that these genes are required for control of the virus. We show that viral small RNAs co-immunoprecipitate with a 6xFLAG-3xHIS-tagged AGO1 protein by sequencing the small RNAs from immunoprecipitated fractions. Additionally, analyses of the small RNA datasets from the RNAi mutants revealed control of the virus through small RNA-mediated silencing required both AGO1 and DCL1.
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Project description:Here we describe the identification and regulation of a novel dsRNA virus in Colletotrichum higginsianum. High throughput sequencing of small RNAs and strand-specific RNA-seq was performed on single gene knock-out mutants created for each RNAi component gene: rdr1, rdr2, rdr3, dcl1, dcl2, ago1, and ago2, and the double mutant: ∆dcl1∆dcl2. De novo assembly of the ∆dcl1 RNA-seq data identified two contigs that represented the forward and reverse strands of an uncharacterized dsRNA virus, designated here as Colletotrichum higginsianum non-segmented dsRNA virus 1 (ChNRV1). We found increased presence of the viral RNA in the RNA-seq datasets of the ∆dcl1, ∆dcl1dcl2, and ∆ago1 strains, suggesting that these genes are required for control of the virus. We show that viral small RNAs co-immunoprecipitate with a 6xFLAG-3xHIS-tagged AGO1 protein by sequencing the small RNAs from immunoprecipitated fractions. Additionally, analyses of the small RNA datasets from the RNAi mutants revealed control of the virus through small RNA-mediated silencing required both AGO1 and DCL1.
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Project description:We sequenced Endogenous short RNAs in Mucor circinelloides fungus grown in standard liquid culture. Short RNAs were profiled in wild type, Dicer-like 1 mutant (dcl1-), Dicer-like 2 mutant (dcl2-) and double Dicer mutant (dcl1-/dcl2-) strains. We identified many loci that produced less short RNAs in the dcl2- strain suggesting that DCL2 is the major protein generating short RNAs in Mucor circinelloides.
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