RRF-1 is a C. elegans cellular RNA-directed RNA polymerase (RdRP) that synthesizes secondary small interfering RNAs (22G-RNAs). Its products are unprimed, 5'-triphosphorylated antisense RNAs made on target mRNA templates, and they are loaded into worm-specific Argonautes (WAGOs) to amplify and sustain silencing. RRF-1 is the RdRP of somatic RNAi and, in the germline, the RdRP of the Mutator pathway: it colocalizes with MUT-16 in perinuclear Mutator foci, where it is thought to use poly(UG)-tailed mRNA fragments made by RDE-3 as templates, and it binds UG-repeat RNA. RRF-1 associates with the Dicer-related helicase DRH-3 in RdRP complexes and acts partly redundantly with the germline RdRP EGO-1. It is required for exogenous RNAi, for transposon and endogenous gene silencing.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0001172 RNA-templated transcription | IEA GO_REF:0000108 | ACCEPT | Summary: Inter-ontology IEA from the RdRP activity annotation. Reason: RRF-1 synthesizes RNA on mRNA-derived templates in a cell-free system (PMID:18007599), which is RNA-templated transcription. Supporting Evidence: PMID:18007599 In this cell-free system, RdRP activity acts on mRNA-derived templates to produce small RNAs. |
| GO:0003968 RNA-directed RNA polymerase activity | IBA GO_REF:0000033 | ACCEPT | Summary: PAINT IBA for the eukaryotic RdRP family (PTN000554964). RRF-1 itself (WBGene00004508) is among the descendant evidence. Reason: RRF-1's own experimental RdRP evidence (PMID:18007599) is part of the grounding of this node. That is expected, not circular. The IBA adds that the activity is inherited across the family. Supporting Evidence: PMID:18007599 The RRF-1 complex is predominantly responsible for the RdRP activity, and synthesizes secondary-type siRNA molecules in a Dicer-independent manner. |
| GO:0003968 RNA-directed RNA polymerase activity | IDA PMID:18007599 In vitro analyses of the production and activity of secondar... | ACCEPT | Summary: In a C. elegans cell-free system the RRF-1 complex was responsible for most RdRP activity and synthesized secondary-type siRNAs (PMID:18007599). Reason: Direct biochemical evidence. This is the core molecular function. Supporting Evidence: PMID:18007599 The RRF-1 complex is predominantly responsible for the RdRP activity, and synthesizes secondary-type siRNA molecules in a Dicer-independent manner. |
| GO:0003968 RNA-directed RNA polymerase activity | IDA PMID:24684932 MUT-14 and SMUT-1 DEAD box RNA helicases have overlapping ro... | ACCEPT | Summary: Mutator complexes purified from worms, which contain RRF-1, synthesize siRNAs in vitro (PMID:24684932). Reason: Independent biochemical support for RdRP activity of the RRF-1-containing mutator complex (PMID:24684932, PMID:22713602). Supporting Evidence: PMID:24684932 mutator complexes containing the mutant protein retain the ability to synthesize siRNAs in vitro PMID:22713602 The RdRP RRF-1 colocalizes with MUT-16 at Mutator foci, suggesting a role for Mutator foci in siRNA amplification. |
| GO:0003968 RNA-directed RNA polymerase activity | IEA GO_REF:0000120 | ACCEPT | Summary: Combined IEA (InterPro IPR007855 RdRP domain; EC 2.7.7.48). Reason: Consistent with the direct biochemical evidence (PMID:18007599). Supporting Evidence: PMID:18007599 The RRF-1 complex is predominantly responsible for the RdRP activity, and synthesizes secondary-type siRNA molecules in a Dicer-independent manner. |
| GO:0003968 RNA-directed RNA polymerase activity | ISS PMID:10704412 EGO-1 is related to RNA-directed RNA polymerase and function... | ACCEPT | Summary: ISS from the RdRP family relationship first described for EGO-1, which noted RRF-1 as a related C. elegans RdRP (PMID:10704412). Reason: Sequence-based inference later confirmed biochemically (PMID:18007599). Supporting Evidence: PMID:10704412 The predicted EGO-1 protein was found to be related to the tomato RNA-directed RNA polymerase (RdRP) PMID:18007599 The RRF-1 complex is predominantly responsible for the RdRP activity, and synthesizes secondary-type siRNA molecules in a Dicer-independent manner. |
| GO:0005515 protein binding | IPI PMID:18007599 In vitro analyses of the production and activity of secondar... | MODIFY | Summary: IPI with DRH-3 (Q93413) from the RRF-1 complex purification (PMID:18007599). Reason: The interaction with the Dicer-related helicase DRH-3 is real and functionally important: DRH-3 is a core component of RdRP complexes (PMID:19800275). Generic protein binding carries no functional information. The informative term for the same interaction, DEAD/H-box RNA helicase binding, is already annotated from the same paper. Proposed replacements: DEAD/H-box RNA helicase binding Supporting Evidence: PMID:19800275 DRH-3 is a core component of RNA-dependent RNA polymerase (RdRP) complexes essential for several distinct 22G-RNA systems. |
| GO:0005634 nucleus | IDA PMID:18007599 In vitro analyses of the production and activity of secondar... | KEEP AS NON CORE | Summary: Nuclear localization from the Aoki et al. cell-free/biochemical study (PMID:18007599; abstract only in cache). Reason: Curator IDA from full text that we cannot read; deferred to. RRF-1's characterized site of action in the germline is the cytoplasmic perinuclear Mutator focus (PMID:22713602), so nuclear localization is not core. Supporting Evidence: PMID:22713602 The RdRP RRF-1 colocalizes with MUT-16 at Mutator foci, suggesting a role for Mutator foci in siRNA amplification. |
| GO:0005737 cytoplasm | IDA PMID:18007599 In vitro analyses of the production and activity of secondar... | ACCEPT | Summary: Cytoplasmic localization (PMID:18007599). Reason: Consistent with the perinuclear cytoplasmic Mutator foci where RRF-1 concentrates in germ cells (PMID:22713602). Supporting Evidence: PMID:22713602 The RdRP RRF-1 colocalizes with MUT-16 at Mutator foci, suggesting a role for Mutator foci in siRNA amplification. |
| GO:0016779 nucleotidyltransferase activity | IDA PMID:18007599 In vitro analyses of the production and activity of secondar... | MODIFY | Summary: IDA for nucleotidyltransferase activity from the RdRP assays (PMID:18007599). Reason: Correct but much less informative than the activity actually shown, which is RNA-templated RNA synthesis. RNA-directed RNA polymerase activity is the appropriate term. Proposed replacements: RNA-directed RNA polymerase activity Supporting Evidence: PMID:18007599 The RRF-1 complex is predominantly responsible for the RdRP activity, and synthesizes secondary-type siRNA molecules in a Dicer-independent manner. |
| GO:0017151 DEAD/H-box RNA helicase binding | IPI PMID:18007599 In vitro analyses of the production and activity of secondar... | KEEP AS NON CORE | Summary: IPI with DRH-3 (WBGene00008400), a DEAH/D-box helicase, in the RRF-1 complex (PMID:18007599). Reason: The RRF-1-DRH-3 association defines the RdRP complex (PMID:19800275). It is a valid binding annotation but describes complex membership rather than RRF-1's own catalytic function. Supporting Evidence: PMID:19800275 DRH-3 is a core component of RNA-dependent RNA polymerase (RdRP) complexes |
| GO:0030422 siRNA processing | IBA GO_REF:0000033 | ACCEPT | Summary: PAINT IBA for siRNA processing (PTN000554964). The GO definition includes amplification of siRNA by RdRP. Reason: RRF-1 is the RdRP that synthesizes secondary siRNAs during RNAi and in the Mutator pathway (PMID:18007599, PMID:22713602). It performs the amplification step. Supporting Evidence: PMID:18007599 The RRF-1 complex is predominantly responsible for the RdRP activity, and synthesizes secondary-type siRNA molecules in a Dicer-independent manner. PMID:22713602 The RdRP RRF-1 colocalizes with MUT-16 at Mutator foci, suggesting a role for Mutator foci in siRNA amplification. |
| GO:0030422 siRNA processing | IMP PMID:18007599 In vitro analyses of the production and activity of secondar... | ACCEPT | Summary: rrf-1 mutant extracts lack most secondary-siRNA-producing RdRP activity (PMID:18007599). Reason: Core process, supported by both mutant and biochemical evidence. Supporting Evidence: PMID:18007599 The RRF-1 complex is predominantly responsible for the RdRP activity, and synthesizes secondary-type siRNA molecules in a Dicer-independent manner. |
| GO:0031380 nuclear RNA-directed RNA polymerase complex | IBA GO_REF:0000033 | MARK AS OVER ANNOTATED | Summary: PAINT IBA from S. pombe Rdp1 (RDRC). The term is defined as the RdRP complex required for RNAi-mediated heterochromatin assembly. Reason: RRF-1 does form an RdRP complex with the Dicer-related helicase DRH-3 (PMID:18007599, PMID:19800275). But the characterized RRF-1 complex acts in cytoplasmic perinuclear Mutator foci (PMID:22713602), and RRF-1 is not one of the RdRPs needed for H3K9me2 assembly on unpaired meiotic DNA, which requires only EGO-1 (PMID:16271877). The nuclear, heterochromatin-assembly complex defined by this term fits EGO-1 better than RRF-1. Propagation Review Root cause: PROPAGATION BAD Failure modes: COMPARTMENT OR COMPLEX MISMATCH Sources checked: PANTHER:PTN000554964 · PTN000554964 SUPPORTS SOURCE BUT NOT TARGET Eukaryotic RdRP node seeded by S. pombe Rdp1, whose RDRC acts in nuclear heterochromatin assembly. RRF-1 keeps the RdRP-helicase complex but acts in cytoplasmic Mutator foci; in C. elegans the heterochromatin role belongs to EGO-1. Supporting Evidence: PMID:16271877 Among C. elegans RdRPs, we find that only EGO-1 is required for H3K9me2 enrichment on unpaired chromosomal regions during meiosis. PMID:22713602 The RdRP RRF-1 colocalizes with MUT-16 at Mutator foci, suggesting a role for Mutator foci in siRNA amplification. |
| GO:1990633 mutator focus | IDA PMID:22713602 MUT-16 promotes formation of perinuclear mutator foci requir... | ACCEPT | Summary: RRF-1 colocalizes with MUT-16 at perinuclear Mutator foci in germ cells (PMID:22713602). Reason: Defining localization. Mutator foci are sites of siRNA amplification and of pUG RNA accumulation (PMID:32499657). Supporting Evidence: PMID:22713602 The RdRP RRF-1 colocalizes with MUT-16 at Mutator foci, suggesting a role for Mutator foci in siRNA amplification. PMID:32499657 Indeed, both the endonuclease RDE-8, which cleaves mRNAs targeted by dsRNA12, and the RdRP RRF-117 also localize to Mutator foci, further suggesting that pUG RNA/siRNA cycling occurs in Mutator foci. |
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Download this section (compressed HTML)Q: Does purified RRF-1 initiate synthesis directly on the pUG tail of a pUG-tailed RNA, as the UG-repeat binding data suggest, and what fixes the 22-nt product length?
Q: RRF-1 has three UniProtKB entries (G5ECM1, G3MU40, Q7JLA0). Should GO annotations be consolidated on the isoform a entry G5ECM1?
Experiment: Reconstitute RRF-1 (with and without DRH-3/EKL-1) and test RdRP activity on pUG-tailed versus untailed mRNA fragments.
Hypothesis: pUG tails recruit RRF-1 and promote de novo initiation of 22G-RNA synthesis on the attached mRNA fragment.
rrf-1 encodes an RNA-directed RNA polymerase (RdRP) homolog required for somatic (but not germline) RNAi; RRF-1 is also required for 'transitive interference'.
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