RDE-3 (also called MUT-2) is a C. elegans ribonucleotidyltransferase of the polymerase beta nucleotidyltransferase superfamily that acts as a poly(UG) polymerase. It adds long, non-templated, perfectly alternating uridine and guanosine tails (pUG tails) to the 3' ends of mRNA fragments targeted by RNA interference and to transposon RNAs. pUG tails with more than 16 perfectly alternating U and G nucleotides turn the attached RNA fragment into a template for RNA-directed RNA polymerases such as RRF-1, which synthesize secondary 22G siRNAs. Cycles of pUG RNA-templated siRNA synthesis and siRNA-directed pUGylation sustain dsRNA-triggered transgenerational silencing in the germline. RDE-3 localizes to perinuclear germline Mutator foci with RRF-1 and the RDE-8 endonuclease. It is required for siRNA accumulation, RNAi in all tissues, transposon silencing, and fertility and viability at high temperature. RDE-3 also pUGylates Orsay virus RNAs to support antiviral RNAi.
Definition: Catalysis of the template-independent addition of alternating uridine and guanosine monophosphate residues to the 3' end of an RNA molecule, forming a poly(UG) tail.
Justification: RDE-3/MUT-2 adds perfectly alternating UG tails, which no existing child of template-free RNA nucleotidyltransferase activity describes. RNA uridylyltransferase activity covers only the U additions.
Parent term: template-free RNA nucleotidyltransferase activity
Supporting Evidence:
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0009792 embryo development ending in birth or egg hatching | IMP PMID:15723801 A member of the polymerase beta nucleotidyltransferase super... | KEEP AS NON CORE | Summary: rde-3 null mutants are inviable and sterile at high temperature (PMID:15723801). Reason: A real mutant phenotype, but a temperature-dependent, pleiotropic consequence of losing pUG RNA-driven small-RNA silencing. RDE-3 has no known direct role in embryogenesis. Supporting Evidence: PMID:15723801 Genetic analysis of presumptive null alleles indicates that rde-3 is required for siRNA accumulation and for efficient RNAi in all tissues, and it is essential for fertility and viability at high temperatures. |
| GO:0035194 regulatory ncRNA-mediated post-transcriptional gene silencing | IMP PMID:10535731 The rde-1 gene, RNA interference, and transposon silencing i... | ACCEPT | Summary: rde-3 was among the RNAi-deficient (Rde) mutants recovered by Tabara et al. and is associated with transposon mobilization (PMID:10535731). Reason: RDE-3 does part of the work of RNAi: it pUGylates target mRNA fragments, converting them into RdRP templates (PMID:32499657). Supporting Evidence: PMID:10535731 Interestingly, several, but not all, RNAi-deficient strains exhibit mobilization of the endogenous transposons. PMID:32499657 pUG tails promote gene silencing by recruiting RNA-dependent RNA polymerases, which use pUG-tailed RNAs (pUG RNAs) as templates to synthesize small interfering RNAs (siRNAs). |
| GO:0035194 regulatory ncRNA-mediated post-transcriptional gene silencing | IMP PMID:15723801 A member of the polymerase beta nucleotidyltransferase super... | ACCEPT | Summary: Presumptive null rde-3 alleles abolish siRNA accumulation and efficient RNAi in all tissues (PMID:15723801). Reason: Core process. The mechanism was later shown to be pUG tailing of RNAi target fragments (PMID:32499657). Supporting Evidence: PMID:15723801 Genetic analysis of presumptive null alleles indicates that rde-3 is required for siRNA accumulation and for efficient RNAi in all tissues, and it is essential for fertility and viability at high temperatures. PMID:32499657 Here we show that, in its natural context in C. elegans, RDE-3 adds pUG tails to targets of RNA interference, as well as to transposon RNAs. |
| GO:0045132 meiotic chromosome segregation | IMP PMID:10535731 The rde-1 gene, RNA interference, and transposon silencing i... | KEEP AS NON CORE | Summary: Meiotic chromosome segregation defect (high incidence of males) in rde-3 mutants, from Tabara et al. (PMID:10535731; abstract only in cache). Reason: Curator IMP from full text we cannot read; deferred to. Mutator-class mutants commonly show X-chromosome nondisjunction, but RDE-3 is a pUG polymerase with no known direct role in chromosome segregation. The phenotype is most likely an indirect consequence of defective small-RNA silencing. Supporting Evidence: PMID:10535731 several, but not all, RNAi-deficient strains exhibit mobilization of the endogenous transposons |
| GO:0050265 RNA uridylyltransferase activity | IBA GO_REF:0000033 | ACCEPT | Summary: PAINT IBA for RNA uridylyltransferase activity at the TUTase/PAP-like nucleotidyltransferase node (PTN000263330). Reason: RDE-3 does transfer UMP to RNA 3' ends, but as half of an alternating UG polymer: it is a poly(UG) polymerase (PMID:30988468, PMID:32499657). The term is correct but incomplete because no GO term covers poly(UG) polymerase activity. A new term is proposed (proposed_new_terms). Supporting Evidence: PMID:30988468 a poly(UG) polymerase, Caenorhabditis elegans MUT-2, that adds alternating uridine and guanosine nucleotides to form poly(UG) tails PMID:32499657 Here we show that, in its natural context in C. elegans, RDE-3 adds pUG tails to targets of RNA interference, as well as to transposon RNAs. |
| GO:1990074 polyuridylation-dependent mRNA catabolic process | IBA GO_REF:0000033 | MARK AS OVER ANNOTATED | Summary: PAINT IBA for polyuridylation-dependent mRNA catabolism, seeded by S. pombe Cid1/Cid16 and mammalian TUT4/TUT7. Reason: In the donor enzymes, oligo(U) tails mark mRNAs for decay. RDE-3 instead adds pUG tails that convert mRNA fragments into templates for RdRP-driven siRNA synthesis and transgenerational silencing (PMID:32499657). The tailed RNA acts as a silencing intermediate, not a catabolic substrate. RDE-3-directed silencing does lower target mRNA levels, so the annotation is not entirely wrong, but it describes the ancestral TUTase function and not the diverged one in this lineage. Propagation Review Root cause: PROPAGATION BAD Failure modes: FUNCTIONAL DIVERGENCE Sources checked: PANTHER:PTN000263330 · PTN000263330 SUPPORTS SOURCE BUT NOT TARGET Node seeded by Cid1-type and TUT4/7 uridylyltransferases whose oligo(U) tails trigger mRNA decay. RDE-3 is a diverged poly(UG) polymerase whose tails make silencing templates. Supporting Evidence: PMID:32499657 pUG tails promote gene silencing by recruiting RNA-dependent RNA polymerases, which use pUG-tailed RNAs (pUG RNAs) as templates to synthesize small interfering RNAs (siRNAs). PMID:32499657 pUG tails could either mark mRNA fragments for degradation or convert mRNA fragments into active mediators of gene silencing. |
| GO:1990633 mutator focus | IDA PMID:22713602 MUT-16 promotes formation of perinuclear mutator foci requir... | ACCEPT | Summary: RDE-3/MUT-2 is one of the mutator proteins that localize to perinuclear Mutator foci (PMID:22713602). Reason: Core location. pUG RNAs colocalize with RDE-3 in Mutator foci, which are likely sites of pUGylation (PMID:32499657). Supporting Evidence: PMID:22713602 Here we show that each of the six mutator proteins localizes to punctate foci at the periphery of germline nuclei. PMID:32499657 The fact that enzyme and enzyme product both localize to Mutator foci suggests that Mutator foci may be sites of RNA pUGylation. |
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Download this section (compressed HTML)Q: Does RDE-3 have intrinsic UG alternation in vitro, or does the alternation depend on partner proteins or template-like secondary structure in the growing tail?
Q: Should GO create a poly(UG) polymerase activity term under template-free RNA nucleotidyltransferase activity (GO:0098680)?
Experiment: In vitro pUGylation assays with recombinant RDE-3, comparing wild type and catalytic-site mutants and varying UTP/GTP ratios.
Hypothesis: RDE-3 alone adds perfectly alternating UG tails to RNA 3' ends.
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