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| Status |
Public on Apr 01, 2015 |
| Title |
Small RNA sequencing of transgenes silencing by paramutation in C. elegans |
| Organism |
Caenorhabditis elegans |
| Experiment type |
Non-coding RNA profiling by high throughput sequencing
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| Summary |
In the nematode Caenorhabditis elegans, different small RNA-dependent gene silencing mechanisms act in the germline to initiate transgenerational gene silencing. Piwi-interacting RNAs (piRNAs) can initiate transposon and gene silencing by acting upstream of endogenous short interfering RNAs (siRNAs), which engage a nuclear RNA interference (RNAi) pathway to trigger transcriptional gene silencing. Once gene silencing has been established, it can be stably maintained over multiple generations without the requirement of the initial trigger and is also referred to as RNAe or paramutation. This heritable silencing depends on the integrity of the nuclear RNAi pathway. However, the exact mechanism by which silencing is maintained across generations is not understood.Here we demonstrate that silencing of piRNA targets involves the production of two distinct classes of small RNAs with different genetic requirements. The first class, secondary siRNAs, are localized close to the direct target site for piRNAs. Nuclear import of the secondary siRNAs by the Argonaute HRDE-1 leads to the production of a distinct class of small RNAs that map throughout the transcript, which we term tertiary siRNAs. Both classes of small RNAs are necessary for full repression of the target gene and can be maintained independently of the initial piRNA trigger. Consistently, we observed a form of paramutation associated with tertiary siRNAs. Once paramutated, a tertiary siRNA generating allele confers dominant silencing in the progeny regardless of its own transmission, suggesting germline-transmitted siRNAs are sufficient for multigenerational silencing.
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| Overall design |
C. elegans strains containing transgenes silenced by piRNAs were crossed to strains with transgenes with similar sequences but without piRNA target sites, to investigate the spreading of silencing between transgenes mediated by small RNAs. Mutant backgrounds were used to investigate the genetic requirements for this process.
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| Contributor(s) |
Sarkies P, Miska E |
| Citation missing |
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| Submission date |
Feb 26, 2015 |
| Last update date |
Nov 30, 2021 |
| Contact name |
Peter Sarkies |
| E-mail(s) |
[email protected]
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| Organization name |
Imperial College London
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| Department |
MRC Clinical Sciences Centre
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| Street address |
Du Cane Road
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| City |
London |
| ZIP/Postal code |
W12 0NN |
| Country |
United Kingdom |
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| Platforms (2) |
| GPL15716 |
Illumina MiSeq (Caenorhabditis elegans) |
| GPL18730 |
Illumina HiSeq 1500 (Caenorhabditis elegans) |
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| Samples (24)
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| Relations |
| BioProject |
PRJNA276571 |
| SRA |
SRP055556 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE66344_RAW.tar |
273.2 Mb |
(http)(custom) |
TAR (of FA) |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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