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Series GSE201233 Query DataSets for GSE201233
Status Public on Feb 17, 2023
Title 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]
Organism Arabidopsis thaliana
Experiment type Non-coding RNA profiling by high throughput sequencing
Summary 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.
Overall design Small RNA sequencing of RNAs copurifying with wild-type or slicing-defective AGO4 proteins stably expressed in ago4-4 null-mutant Arabidopsis plants.
Contributor(s) Wang F, Huang H, Pikaard C
Citation(s) 36746605
Submission date Apr 21, 2022
Last update date Feb 18, 2023
Contact name Craig S. Pikaard
Organization name Howard Hughes Medical Institute, Indiana University
Department Department of Biology and Department of Molecular and Cellular Biochemistry
Street address 915 E. Third Street
City Bloomington
State/province Indiana
ZIP/Postal code 47405
Country USA
Platforms (1)
GPL19580 Illumina NextSeq 500 (Arabidopsis thaliana)
Samples (6)
GSM6053228 sRNA-seq, mock immunoprecipitation, biol rep 1
GSM6053229 sRNA-seq, wild-type AGO4 immunoprecipitation, biol rep 1
GSM6053230 sRNA-seq, slicing-defective AGO4 immunoprecipitation, biol rep 1
This SubSeries is part of SuperSeries:
GSE201235 Enzymatic reactions of AGO4 in RNA-directed DNA methylation: siRNA duplex loading, passenger strand elimination, target RNA slicing, and sliced target retention
BioProject PRJNA830475

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Supplementary file Size Download File type/resource
GSE201233_RAW.tar 70.5 Mb (http)(custom) TAR (of TXT)
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Raw data are available in SRA
Processed data provided as supplementary file

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