NRDE-2 (nuclear RNAi defective-2) is a conserved nuclear protein of the NRDE2 family, with an N-terminal NRDE-2 domain, an SR-rich region and HAT-like (half-a-TPR) repeats. It is a core downstream factor of the C. elegans nuclear RNAi pathway. siRNA-loaded nuclear Argonautes (NRDE-3 in somatic cells, HRDE-1 in the germline) recruit NRDE-2 to nascent transcripts complementary to their guide RNAs. NRDE-2 is then needed for NRDE-1 to associate with pre-mRNA and chromatin, for siRNA-directed H3K9 and H3K27 trimethylation at target loci, and for inhibition of RNA polymerase II during transcription elongation, which silences target pre-mRNA co-transcriptionally. When antisense ribosomal siRNAs are present, NRDE-2 also moves to the nucleolus and inhibits RNA polymerase I elongation on pre-rRNA. NRDE-2 is required for multigenerational RNAi inheritance and germline immortality. The human ortholog NRDE2 binds and inhibits the exosome cofactor MTR4 in nuclear speckles. Whether worm NRDE-2 keeps this exosome-regulatory activity has not been tested.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005515 protein binding | IPI PMID:20543824 Small regulatory RNAs inhibit RNA polymerase II during the e... | REMOVE | Summary: NRDE-2 co-immunoprecipitates with the nuclear Argonaute NRDE-3. The association is weak (about 0.5% of the cellular NRDE-2 pool) and depends on NRDE-3 nuclear localization. Reason: Generic protein binding is uninformative. The NRDE-3 association is real, but the paper does not show a specific molecular activity (for example a direct adaptor function) that would justify a more informative MF term. The biology is captured by the process annotations. Supporting Evidence: PMID:20543824 In support of this hypothesis, we observed a weak, but reproducible, association between NRDE-3 and NRDE-2 |
| GO:0005634 nucleus | IDA PMID:20543824 Small regulatory RNAs inhibit RNA polymerase II during the e... | ACCEPT | Summary: Functional GFP::NRDE-2 localizes predominantly to the nucleus. Reason: Core location for a co-transcriptional silencing factor. Supporting Evidence: PMID:20543824 A fusion gene between GFP and NRDE-2 (GFP∷NRDE-2), which was sufficient to rescue nrde-2(-) mutant phenotypes (Fig. 1b), localized predominantly to the nucleus (Fig. 1d). |
| GO:0005634 nucleus | IEA GO_REF:0000044 | ACCEPT | Summary: UniProt subcellular-location mapping to nucleus. Reason: Consistent with experimental data. |
| GO:0005730 nucleolus | IEA GO_REF:0000044 | KEEP AS NON CORE | Summary: UniProt mapping to nucleolus, based on the risiRNA study. NRDE-2 accumulates in the nucleolus when antisense ribosomal siRNAs are present. Reason: Experimentally supported, but conditional. Nucleolar accumulation occurs when risiRNAs accumulate and is linked to silencing of pre-rRNA by RNAP I. Supporting Evidence: PMID:34365510 In the presence of risiRNAs, NRDE-2 accumulated in the nucleolus and colocalized with RNA polymerase I. |
| GO:0016607 nuclear speck | IEA GO_REF:0000044 | KEEP AS NON CORE | Summary: UniProt mapping to nuclear speck, transferred by similarity from human NRDE2. Reason: Human NRDE2 is mainly in nuclear speckles, and worm NRDE-2 is nuclear and keeps the SR-rich and HAT-like regions. Speckle localization has not been shown for the worm protein, so this is kept as plausible but non-core. Supporting Evidence: PMID:30842217 Unexpectedly, NRDE2 mainly localizes in nuclear speckles, where it inhibits MTR4 recruitment and RNA degradation, and thereby ensures efficient mRNA nuclear export. |
| GO:0016607 nuclear speck | ISS GO_REF:0000024 | KEEP AS NON CORE | Summary: ISS transfer of nuclear speckle localization from human NRDE2. Reason: Same reasoning as the IEA row. Plausible from orthology but not yet observed in C. elegans. Supporting Evidence: PMID:30842217 Unexpectedly, NRDE2 mainly localizes in nuclear speckles, where it inhibits MTR4 recruitment and RNA degradation, and thereby ensures efficient mRNA nuclear export. |
| GO:0031047 regulatory ncRNA-mediated gene silencing | IMP PMID:20543824 Small regulatory RNAs inhibit RNA polymerase II during the e... | ACCEPT | Summary: nrde-2 is required for siRNA-mediated silencing of nuclear-localized RNAs. This acts co-transcriptionally, through inhibition of RNAP II elongation. Reason: Correct and suitably general. NRDE-2 silencing combines co-transcriptional RNAP II inhibition and heterochromatin deposition, and this parent term covers both. Supporting Evidence: PMID:20543824 We conclude that siRNAs-direct a NRDE-2/3-dependent inhibition of RNAP II activity that occurs during the elongation phase of transcription. |
| GO:0031048 regulatory ncRNA-mediated heterochromatin formation | IBA GO_REF:0000033 | ACCEPT | Summary: NRDE2-family IBA for ncRNA-mediated heterochromatin formation. Supported in worm by NRDE-2-dependent, siRNA-directed H3K9me3. Reason: Core process, well supported experimentally in C. elegans. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN000336449 · PTN000336449 SUPPORTS TRANSFER NRDE2-family node. Worm NRDE-2 is itself experimentally required for siRNA-directed H3K9me3. Supporting Evidence: PMID:21901112 Endo-siRNA-driven H3K9me requires the nuclear RNAi pathway including the Argonaute (Ago) NRDE-3, the conserved nuclear RNAi factor NRDE-2, as well as NRDE-1 and NRDE-4. |
| GO:0031048 regulatory ncRNA-mediated heterochromatin formation | IMP PMID:22231482 Amplification of siRNA in Caenorhabditis elegans generates a... | ACCEPT | Summary: nrde-2 mutants lose the dsRNA-triggered H3K9me3 footprint at targeted loci. Reason: Direct experimental support for NRDE-2 in RNAi-directed heterochromatin formation. Supporting Evidence: PMID:22231482 nrde-2 mutant animals showed a 6-fold loss in H3K9me3 response to dsRNA compared to eri-1 and wild type strains |
| GO:0035194 regulatory ncRNA-mediated post-transcriptional gene silencing | IMP PMID:21901112 A pre-mRNA-associating factor links endogenous siRNAs to chr... | MODIFY | Summary: nrde-2 mutants are defective in nuclear RNAi (for example lir-1 RNAi) and in the recruitment of NRDE-1 to pre-mRNA. The silencing is nuclear and co-transcriptional, not post-transcriptional. Reason: The genetic requirement is solid, but NRDE-2 acts in the nucleus on nascent transcripts and chromatin. It inhibits RNAP II elongation and promotes H3K9me3 rather than destabilizing mature mRNA or blocking its translation, as the post-transcriptional silencing definition requires. The general parent regulatory ncRNA-mediated gene silencing describes this better. Proposed replacements: regulatory ncRNA-mediated gene silencing Supporting Evidence: PMID:21901112 NRDE-3 and NRDE-2 are required for the association of NRDE-1 with pre-mRNA and chromatin. PMID:20543824 These results define NRDE-2 as a component of the nuclear RNAi machinery and demonstrate that metazoan siRNAs can silence nuclear-localized RNAs co-transcriptionally. |
| GO:0048255 mRNA stabilization | ISS GO_REF:0000024 | MARK AS OVER ANNOTATED | Summary: ISS from human NRDE2, which stabilizes mRNA by inhibiting MTR4 and the nuclear exosome. Reason: The human evidence is solid, but no C. elegans study has shown that NRDE-2 stabilizes mRNAs. Every characterized worm function of NRDE-2 is in siRNA-directed silencing. A manual similarity transfer of this specific outcome, with no worm evidence, goes beyond what is known. The broader IBA (negative regulation of RNA catabolic process) is kept as non-core. Supporting Evidence: PMID:30842217 Together, our data pinpoint NRDE2 as a nuclear exosome negative regulator that ensures mRNA stability and nuclear export. |
| GO:1902369 negative regulation of RNA catabolic process | IBA GO_REF:0000033 | KEEP AS NON CORE | Summary: NRDE2-family IBA for negative regulation of RNA catabolic process, based on exosome inhibition by human (and probably yeast) NRDE2. Reason: The PAINT node places MTR4/exosome inhibition at the base of the NRDE2 family, and there is no worm-specific evidence that it was lost. It is kept as a plausible ancestral activity. It is not the characterized function of worm NRDE-2, so it is non-core. Propagation Review Root cause: NO FAILURE NON CORE Sources checked: PANTHER:PTN000336449 · PTN000336449 SUPPORTS TRANSFER NRDE2-family node. The MTR4-inhibition data come from human NRDE2 and have not been tested for the worm protein. Supporting Evidence: PMID:30842217 NRDE2 forms a 1:1 complex with MTR4, a nuclear exosome cofactor critical for exosome recruitment, via a conserved MTR4-interacting domain (MID). |
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Download this section (compressed HTML)Q: Does C. elegans NRDE-2 keep the MTR4-interacting domain and the MTR-4/exosome-inhibitory activity of human NRDE2? If so, is that activity part of how it silences nascent transcripts?
Q: What is the direct molecular activity by which NRDE-2 links Argonaute recruitment to NRDE-1 loading, H3K9me3 and RNAP II elongation arrest?
Experiment: Test in vitro whether recombinant worm NRDE-2 binds MTR-4. In vivo, test whether NRDE-2 mutants lacking the putative MTR4-interacting region keep nuclear RNAi and H3K9me3 at germline target genes.
nrde-2 encodes a protein containing a conserved domain of unknown function (DUF 1740), a serine/arginine-rich domain, and a half-a-tetratricopeptide (HAT)-like domain that is conserved in plants, fungi, insect, and mammals; NRDE-2 functions with NRDE-3 in the nuclear RNAi pathway to regulate gene expression via inhibition of RNA polymerase II during the elongation phase of transcription and via an NRDE-2-dependent enrichment of histone H3K9 methylation at genomic sites targeted by RNAi; an NRDE-2::GFP fusion protein localizes to the nucleus.
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