ChIP-seq of DRD1, DMS3 and RDM1 in Arabidopsis
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ABSTRACT:
PROVIDER: PRJNA538858 | ENA |
REPOSITORIES: ENA
Similar Datasets
Project description:We performed FLAG ChIP-seq of transgenic lines expressing FLAG-tagged DRD1, DMS3 and RDM1, as well as Col0 as negative control.
2019-07-09 | GSE130290 | GEO
Project description:RNA-directed DNA methylation (RdDM) plays an essential role in transposable element (TE) silencing in plants. In the Arabidopsis RdDM pathway, the DDR complex containing DRD1, DMS3, and RDM1, is necessary for recruiting Pol V to transcribe scaffold RNA. Although the role of DDR is known, the mechanism by which the DDR complex is regulated remains unexplored. Here, we demonstrate that the Anaphase Promoting Complex/Cyclosome (APC/C) monitors the assembly of the DDR complex by targeting DMS3 for degradation. We show that a subset of Pol V-dependent RdDM loci are de-repressed in apc/c mutants, accompanied by defective recruitment and transcription of Pol V. APC/C targets DMS3 for ubiquitination and degradation in a D box-dependent manner, and the D-box-mutated DMS3 fails to complement the dms3 mutant. Competitive binding assays shows that the dosage of DMS3 is critical for the assembly of the DDR complex, and in vivo gel filtration analysis shows that the assembly of both DDR and Pol V is compromised in the apc8 mutant. These findings uncover a safeguard role of APC/C-mediated DMS3 degradation in the assembly of the DDR complex, and provide a direct link between selective protein degradation and RdDM.
2019-01-31 | GSE106970 | GEO
Project description:RNA-directed DNA methylation (RdDM) plays an essential role in transposable element (TE) silencing in plants. In the Arabidopsis RdDM pathway, the DDR complex containing DRD1, DMS3, and RDM1, is necessary for recruiting Pol V to transcribe scaffold RNA. Although the role of DDR is known, the mechanism by which the DDR complex is regulated remains unexplored. Here, we demonstrate that the Anaphase Promoting Complex/Cyclosome (APC/C) monitors the assembly of the DDR complex by targeting DMS3 for degradation. We show that a subset of Pol V-dependent RdDM loci are de-repressed in apc/c mutants, accompanied by defective recruitment and transcription of Pol V. APC/C targets DMS3 for ubiquitination and degradation in a D box-dependent manner, and the D-box-mutated DMS3 fails to complement the dms3 mutant. Competitive binding assays shows that the dosage of DMS3 is critical for the assembly of the DDR complex, and in vivo gel filtration analysis shows that the assembly of both DDR and Pol V is compromised in the apc8 mutant. These findings uncover a safeguard role of APC/C-mediated DMS3 degradation in the assembly of the DDR complex, and provide a direct link between selective protein degradation and RdDM.
2019-01-31 | GSE106971 | GEO
Project description:Chemical cross-linking coupled to mass spectrometry was used to study the A. thaliana DDR complex consisting of DMS3, RMD1 and a peptide from the interaction region of DRD1. Cross-linking was performed using different cross-linking chemistries: disuccinimidyl suberate (DSS) and a combination of adipic acid dihydrazide (ADH) and the coupling reagent, DMTMM.
2019-07-15 | PXD013470 | Pride
Project description:Estrogen Receptor alpha (ERα) is a key driver of most breast cancers, and it is the target of endocrine therapies used in the clinic to treat women with ERα positive (ER+) breast cancer. The two methods ChIP-seq (chromatin immunoprecipitation coupled with deep sequencing) and RIME (Rapid Immunoprecipitation of Endogenous Proteins) have greatly improved our understanding of ERα function during breast cancer progression and in response to anti-estrogens. A critical component of both ChIP-seq and RIME protocols is the antibody that is used to pull down the bait protein. To date, most of the ChIP-seq and RIME experiments for the study of ERα have been performed using the sc-543 antibody from Santa Cruz Biotechnology. However, this antibody has been discontinued, thereby severely impacting the study of ERα in normal physiology as well as diseases such as breast cancer and ovarian cancer. Here, we compare the sc-543 antibody with other commercially available antibodies, and we show that 06-935 (EMD Millipore) and ab3575 (Abcam) antibodies can successfully replace the sc-543 antibody for ChIP-seq and RIME experiments.
2019-03-21 | PXD012930 | Pride
Project description:We identified in a forward genetic screen a novel factor required for RNA-directed DNA methylation (IWR1-like transcription factor). The aim of the study was to identify natural targets of this factor and to compare it to targets of drd1 another major key factor required for the RNA silencing process in plants. We performed transcriptome analysis to identify targets of this factor. For this we used whole plants at seedling stage, isolated total RNA and then used the standard Affymetrix pipeline/chemistry. The mutated transcription factor, dms4-1 was compared to its respective wildtype (EDT, Col-O background). For microarray analysis THREE biological replicates were run.<br><br>For transcriptome comparison we analyzed another factor required for the process RNA-directed DNA methylation in plants, drd1 also identified in a forward genetic screen and previously characterized in depth (Kanno et al. Current Biology, 14, 801-805, (2004), Huettel et al. EMBO J. 25, 2828-2836 (2006)). For transcriptome analysis two alleles of drd1 (drd1-1 and drd1-6) with two biological replicates each was compared to the respective wildtype DT, which served as starting material in this forward genetic screen (DT, Col-0 background). For drd1 mutants and DT wildtype analysis 10 day old seedlings were used as starting matieral while for dms4-1 and EDT 3 week old seedlings were analyzed.
2010-01-10 | E-MEXP-2251 | biostudies-arrayexpress
Project description:The Anaphase Promoting Complex/Cyclosome degrades DMS3 to maintain RNA-directed DNA Methylation in Arabidopsis [lncRNA-seq]
| PRJNA418706 | ENA
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
Project description:Using translational profiling we obtained RNA-Seq data from total RNA present in the dorsal striatum of Drd1-EGFP/Rpl10a (CP73) mice administed cocaine
2019-12-17 | GSE142142 | GEO
Project description:The Anaphase Promoting Complex/Cyclosome degrades DMS3 to maintain RNA-directed DNA Methylation in Arabidopsis [small RNA-seq]
| PRJNA418705 | ENA