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| Status |
Public on Feb 25, 2017 |
| Title |
Functional analysis of the Arabidopsis thaliana Pol V largest subunit, NRPE1, carboxy-terminal domain by whole genome bisulfite sequencing |
| Organism |
Arabidopsis thaliana |
| Experiment type |
Methylation profiling by high throughput sequencing
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| Summary |
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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| Overall design |
NRPE1 constructs with targeted carboxy-terminal domain deletions were assessed for their ability to complement nrpe1-11 mutants. Controls include Col-0 wild-type plants, nrpe1-11 mutant plants, and nrpe1-11 mutants expressing a full length NRPE1 transgene.
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| Contributor(s) |
Wendte JM, Pikaard CS |
| Citation missing |
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| Submission date |
Jan 10, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Jered Wendte |
| Organization name |
Indiana University
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| Street address |
915 E. 3rd Street
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| City |
Bloomington |
| State/province |
IN |
| ZIP/Postal code |
47405 |
| Country |
USA |
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| Platforms (1) |
| GPL19580 |
Illumina NextSeq 500 (Arabidopsis thaliana) |
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| Samples (26)
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| This SubSeries is part of SuperSeries: |
| GSE93360 |
Functional analysis of the RNA polymerase V CTD |
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| Relations |
| BioProject |
PRJNA360690 |
| SRA |
SRP096354 |
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