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Arabidopsis thaliana

ABSTRACT: Functional analysis of the Arabidopsis thaliana Pol V largest subunit, NRPE1, carboxy-terminal domain by whole genome bisulfite sequencing

PROVIDER: PRJNA360690 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR5163930_1.fastq.gz	Fastqsanger.gz
	SRR5163930_2.fastq.gz	Fastqsanger.gz
	SRR5163940_1.fastq.gz	Fastqsanger.gz
	SRR5163940_2.fastq.gz	Fastqsanger.gz
	SRR5163931_1.fastq.gz	Fastqsanger.gz

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Long non-coding RNA produced by RNA polymerase V determines boundaries of heterochromatin

Project description:We report genome-wide detection of long noncoding RNA (lncRNA) generate by Pol V in transcriptional gene silencing in Arabidopsis thaliana. We further show that most of these transcripts are bound by AGO4. RNA immunoprecipitation followed by high-throughput sequencing (RIP-seq) RNA immunoprecipitation with an anti-AGO4 antibody was performed in one biological replicate and included Col-0, nrpe1, and ago4-1; RNA immunoprecipitation with an anti-NRPE1 (largest subunit of Pol V) was performed in two replicates; replicate 1 included Col-0 and nrpe1, replicated two included Col-0, nrpe1, ago4-1, and idn2-1

2016-10-24 | E-GEOD-70290 | biostudies-arrayexpress

The DDR complex facilitates the genome-wide association of RNA Polymerase V to promoters and evolutionarily young transposons

Project description:The plant-specific DNA-dependent RNA polymerase V (Pol V) evolved from Pol II to function in an RNA-directed DNA methylation pathway. Here, we have identified targets of Pol V in Arabidopsis thaliana on a genome-wide scale using ChIP-seq of NRPE1, the largest catalytic subunit of Pol V. We found that Pol V is enriched at promoters and evolutionarily recent transposons. This localization pattern is highly correlated with Pol V-dependent DNA methylation and small RNA accumulation. We also show that genome-wide association of Pol V with chromatin is dependent on all members of a putative chromatin-remodeling complex termed DDR. Our study presents the first genome-wide view of Pol V occupancy and sheds light on the mechanistic basis of Pol V localization. Furthermore, these findings suggest a role for Pol V and RNA-directed DNA methylation in genome surveillance and in responding to genome evolution. For wild type plants (ecotype Columbia) and nrpe1 mutants whole-genome small RNA (sRNA-seq) and bisulfite sequencing (BS-seq) was performed. In addition whole genome chromatin immunoprecipitation (ChIP-seq) was performed on wild type (ecotype Columbia) plants as a negative control with experimentals consiting of wild type plants carrying a C-terminally epitope tagged (2XFLAG) NRPE1, as well as the NRPE1-FLAG construct in drd1, dms3, and rdm1 mutant backgrounds.

2012-08-05 | E-GEOD-39247 | biostudies-arrayexpress

Arabidopsis thaliana

Project description:sRNA expression in the Pol V largest subunit mutant, nrpe1-11

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PolII Rpb3 ChIP-seq data from D. melanogaster

Project description:We performed Pol II ChIP-seq experiments with 3rd instar larva salivary glands material from male, female and MSL2 RNAi D.melanogaster, as well as from S2 cell lines material. The antibody used recognizes the Rpb3 subunit of RNA Pol II irrespective of the phosphorylation state of the Rpb1 carboxy-terminal domain, therefore capturing Pol II during all stages of transcription. Corrected processed data has been made available following Vaquerizas et al (2013). [M-^SResponse to Comments on M-^SDrosophila Dosage Compensation Involves Enhanced Pol II Recruitment to Male X-Linked PromotersM-^T, Juan M. Vaquerizas, Florence M. G. Cavalli, Thomas Conrad, Asifa Akhtar, Nicholas M. Luscombe, Science 340 (6130): 273 (2013)]

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The DDR complex facilitates the genome-wide association of RNA Polymerase V to promoters and evolutionarily young transposons

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A SWI/SNF nucleosome remodeling complex acts in non-coding RNA-mediated transcriptional silencing

Project description:RNA-mediated transcriptional silencing prevents deleterious effects of transposon activity and controls the expression of protein-coding genes. It involves long non-coding RNAs (lncRNAs)1, which in Arabidopsis thaliana are produced by a specialized RNA Polymerase V (Pol V)2. lncRNAs guide Argonaute-siRNA complexes to specific genomic loci and mediate the establishment of DNA methylation3,4. The mechanism by which lncRNAs affect chromatin structure and mRNA production remains mostly unknown. Here we identify the SWI/SNF nucleosome remodeling complex as a component of the RNA-mediated transcriptional silencing pathway. We found that SWI3, an essential subunit of the SWI/SNF complex, physically interacts with a lncRNA-binding IDN2 protein5,6. RNA-mediated DNA methylation and transcriptional silencing was compromised in the swi3 mutant. Moreover, targets of SWI/SNF significantly overlapped with genes controlled by Pol V, which shows that the physical interaction reflects a functional relationship. We further found that non-coding transcription by Pol V affects nucleosome positioning on silenced regions. We propose that lncRNAs mediate transcriptional silencing by guiding the SWI/SNF complex and establishing positioned nucleosomes on specific genomic loci. We further propose that guiding ATP-dependent chromatin remodeling complexes may be a more general function of lncRNAs. RNA-seq of 3 samples (Col-0, nrpe1 [a mutant defective in the largest subunit of Pol V and unable to produce lncRNA], swi3b, idn2) with 3 biological replicates.

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Project description:The Arabidopsis thaliana multi-subunit RNA Polymerase IV largest subunit DeCL domain was functionally assessed with an NRPD1 DeCL domain deletion construct.

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Assessment of the Pol IV largest subunit, NRPD1, DeCL domain deletion construct to rescue Pol IV-dependent DNA methylation in the nrpd1-3 mutant (Bisulfite-Seq).

Project description:The Arabidopsis thaliana multi-subunit RNA Polymerase IV largest subunit DeCL domain was functionally assessed with an NRPD1 DeCL domain deletion construct.

2017-07-17 | GSE95823 | GEO

Functional analysis of the Arabidopsis thaliana Pol V largest subunit, NRPE1, carboxy-terminal domain by whole genome bisulfite sequencing

Project description:RNA Polymerase V transcription recruits siRNA-Argonaute protein complexes to chromatin, thereby specifying sites of RNA-directed DNA methylation (RdDM) and transcriptional gene silencing in plants. The Pol V largest subunit, NRPE1, has an extensive carboxyl-terminal domain (CTD) that is dispensable for catalytic activity in vitro, yet essential in vivo. A CTD subdomain, DeCL, named for its similarity to a chloroplast protein, DEFECTIVE CHLOROPLASTS AND LEAVES, is required for Pol V function at virtually all loci, similar to mutants defective for Pol V recruitment. Deletions removing three other CTD subdomains affect overlapping subsets of loci, similar to mutants lacking proteins that bind Pol V or its transcripts. A yeast two-hybrid screen for CTD-interactors identified the 3-prime -> 5-prime exoribonuclease, RRP6L1 as an interactor with the DeCL subdomain and the adjacent QS subdomain, named for its numerous glutamine-serine (QS) repeats. These RRP6L1-binding subdomains immediately follow the Argonaute-binding subdomain. Experimental evidence indicates that RRP6L1 trims the 3-prime ends of Pol V transcripts sliced by ARGONAUTE 4 (AGO4), suggesting a model whereby the adjacent CTD subdomains enable the spatial and temporal coordination of AGO4 and RRP6L1 RNA processing activities.

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sRNA expression in the Pol V largest subunit mutant, nrpe1-11

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