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Expression data from 5-week-old Arabidopsis

ABSTRACT: AGO4 plays an important role in RNA-directed DNA methylation (RdDM). RdDM on specific genomic loci have the potential to silence the nearby protein coding gene. We used wild-type La-er and ago4-1 mutant Arabidopsis to identify AGO4-regulated genes.

ORGANISM(S): Arabidopsis thaliana

PROVIDER: GSE18928 | GEO | 2010/12/20

SECONDARY ACCESSION(S): PRJNA121035

REPOSITORIES: GEO

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Expression data from 5-week-old Arabidopsis

Project description:AGO4 plays an important role in RNA-directed DNA methylation (RdDM). RdDM on specific genomic loci have the potential to silence the nearby protein coding gene. We used wild-type La-er and ago4-1 mutant Arabidopsis to identify AGO4-regulated genes. Five-weeks wild-type and ago4-1 mutant RNA were used for Affymetrix microarrays. Differentially expressed genes were identified to study the role of AGO4 in regulating plant development.

2010-12-20 | E-GEOD-18928 | biostudies-arrayexpress

RNA polymerase V targets transcriptional silencing components to promoters of protein-coding genes

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Arabidopsis thaliana

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AGO4 slicer activity reveals the passenger strand function of plant 23 nt siRNAs and the post-slicing retention of target RNAs mediating RNA-directed DNA methylation [WGBS]

Project description:In plants, transcriptional silencing by RNA-directed DNA methylation (RdDM) is mediated by ARGONAUTE 4 (AGO4) and 24 nt short-interfering RNAs (siRNAs) that are generated in parallel with 23 nt RNAs of unknown function. We show that 23 nt RNAs serve as the passenger strands of 23/24 nt duplexes loaded into AGO4. The 24 nt siRNAs then guide AGO4 slicing of the passenger strands, generating 11 and 12 nt cleavage products. Unexpectedly, we find that the 12 nt products remain associated with the guide strand-AGO4 complexes. Long noncoding RNAs generated at RdDM loci are similarly sliced and retained by AGO4. These results suggest a model in which RNA POLYMERASE V transcripts at target loci are sliced repeatedly as transcription elongation proceeds, sequentially releasing AGO4-siRNA-scaffold RNA complexes that independently recruit the RdDM machinery. Consistent with this hypothesis, plant lines expressing wild-type versus slicing-defective AGO4 show quantitative variation in cytosine methylation and siRNA levels within RdDM loci.

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AGO4 slicer activity reveals the passenger strand function of plant 23 nt siRNAs and the post-slicing retention of target RNAs mediating RNA-directed DNA methylation [smallRNA-seq]

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Whole genome bisulfite sequencing of two/three-week-old Arabidopsis thaliana plants of the genotypes Col-0, nrpe1-11 and ros1-4.

Project description:This study used the mutants nrpe1-11 and ros1-4 to assess the impact of RNA-directed DNA methylation (RdDM) and ROS1 on the Arabidopsis thaliana (ecotype Col-0) methylome.

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Redundant AGO-hook platforms and AGO4-DNA interaction in polV-directed RdDM

Project description:Small RNA-dependent pairing of AGO/PIWI-containing effector complexes to chromatin-bound nascent transcripts has been proposed as a universal mechanism for guiding RNA-mediated TGS in eukaryotes. Likewise, Pol V-dependent transcripts have been implicated in the targeting of AGO4 to chromatin in RNA-directed DNA methylation (RdDM) in plants. Here, we show that the AGO hook platforms of PolV and SPT5L, another component of the transcriptional complex, are functionally redundant yet essential for RdDM at a genome-wide level. Synthesis of Pol V transcripts is uncoupled from AGO4 recruitment in AGO hook-minus plants, challenging the prevailing RNA-based mechanism of AGO4 targeting to chromatin in RdDM. Transcription is essential to lock the PolV transcription complex into a stable and productive DNA-bound state potentiating interactions of AGO4 with DNA. Consistent with this idea, laser UV-assisted crosslinking shows specific AGO4-DNA interaction at RdDM loci, suggesting a revised model for Pol V-mediated DNA methylation in plants, which explains the exquisite specificity of methylation.

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AGO6 functions in RNA-mediated transcriptional gene silencing in shoot and root meristems in Arabidopsis thaliana

Project description:RNA-directed DNA methylation (RdDM) is a small interfering RNA (siRNA)-mediated epigenetic modification that contributes to transposon silencing in plants. RdDM requires a complex transcriptional machinery that includes specialized RNA polymerases, named Pol IV and Pol V, as well as chromatin remodelling proteins, transcription factors, RNA binding proteins, and other plant-specific proteins whose functions are not yet clarified. In Arabidopsis thaliana, DICER-LIKE3 and members of the ARGONAUTE4 group of AROGONAUTE (AGO) proteins are involved, respectively, in generating and using 24-nt siRNAs that trigger methylation and transcriptional gene silencing (TGS) of homologous promoter sequences. AGO proteins act in silencing effector complexes by anchoring the 3â and 5â ends of the guide siRNAs at their N-terminal PAZ domain and MID domain, respectively. In addition, many AGO proteins cleave complementary target RNAs through an endonuclease (âslicerâ) activity in their C-terminal PIWI domain. AGO4 is the main AGO protein implicated in the RdDM pathway. Here we report the identification of the related AGO6 in a forward genetic screen for mutants defective in RdDM and TGS in shoot and root apical meristems in Arabidopsis thaliana. The identification of AGO6, and not AGO4, in our screen is consistent with the primary expression of AGO6 in shoot and root growing points and the preferential association of Pol V with AGO6. Examination of siRNA abundance in the trasngenic wild type plant (contains trigger and silencer transgenes) and the ago6-4 mutant.

2011-12-01 | E-GEOD-28537 | biostudies-arrayexpress

Analysis of the Argonaute 4-associated non-coding RNA in Arabidopsis thaliana sheds novel insights into gene regulation through RNA-directed DNA methylation

Project description:RNA-directed DNA methylation (RdDM) is a de novo DNA methylation mechanism in plants that plays a fundamental role in plant defence against invasive DNA and in maintaining genome stability by silencing transposons and repetitive sequences. Using nuclear RNA immunoprecipitation, we constructed a highly enriched library and obtained sequences of ncRNAs specifically associated with Argonaute 4 (AGO4), a key component of RNA-directed DNA methylation. A negative immunoprecipation (IP) library was prepared using IP RNA from tissues that don't express the target protein. A nuclear RNA library was also prepared using nuclei isolated RNA.

2016-07-01 | GSE48617 | GEO

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