Dcr1 is the sole Dicer (ribonuclease III family endoribonuclease) of fission yeast and the enzyme that initiates the RNA interference (RNAi) pathway. It processes double-stranded RNA precursors, arising largely from pericentromeric repeat transcripts and amplified by the RNA-dependent RNA polymerase complex (RDRC/Rdp1), into ~21-23 nt siRNA duplexes that are loaded onto the single Argonaute protein (Ago1) to form the RITS complex. The protein has a modular architecture: an N-terminal DExD/H-box ATP-binding helicase domain (with an MRTGAGKT ATP-binding motif and a DECH box), a helicase C-terminal domain, a Dicer dsRNA-binding/dimerization fold, tandem catalytic RNase III domains (RIIIa and RIIIb) that coordinate Mg2+/Mn2+, and a C-terminal extended double-stranded RNA-binding domain (dsRBD) bearing a novel CHCC zinc-binding motif that binds one Zn2+ and is required for nuclear retention of the protein. Through siRNA biogenesis Dcr1 is essential for RNAi-directed heterochromatin assembly (H3K9 methylation by Clr4) at centromeres, the silent mating-type locus, and telomeres, and hence for accurate chromosome segregation. Beyond canonical RNAi, Dcr1 has roles in co-transcriptional gene silencing at euchromatic loci, in releasing RNA polymerases (Pol I/II/III) at sites of transcription-replication collision to maintain genome stability, and in cellular quiescence (G0). Dcr1 is predominantly nuclear, associating with chromatin and the nuclear periphery, with a smaller cytoplasmic pool.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005737 cytoplasm | IBA GO_REF:0000033 | KEEP AS NON CORE | Summary: Dcr1 has a documented cytoplasmic pool, but its core RNAi/heterochromatin function occurs in the nucleus where it is retained via its zinc-binding C-terminal domain. Cytoplasmic localization is real but represents a contextual/secondary compartment (e.g. upon heat stress when Dicer is deported to the cytoplasm). Reason: Cytoplasmic localization is experimentally supported but is not the principal site of Dcr1's core function; nuclear retention is required for heterochromatin assembly. Supporting Evidence: PMID:21847092 disruption of zinc coordination renders Dcr1 mainly cytoplasmic and leads to remarkable changes in gene expression and loss of heterochromatin assembly |
| GO:0003723 RNA binding | IBA GO_REF:0000033 | MARK AS OVER ANNOTATED | Summary: Dcr1 binds RNA, including double-stranded RNA substrates that it cleaves. RNA binding is accurate but the more specific double-stranded RNA binding term (also annotated) better captures the molecular function. Reason: Generic RNA binding is subsumed by the more informative double-stranded RNA binding annotation (GO:0003725, IDA). Proposed replacements: double-stranded RNA binding Supporting Evidence: PMID:21847092 binds rather strongly to a regular A-form RNA helix (Kd=0.8 ΞΌM, with two dsRBDs bound per 24 bp dsRNA helix) |
| GO:0004525 ribonuclease III activity | IBA GO_REF:0000033 | ACCEPT | Summary: Core molecular function. Dcr1 is a ribonuclease III that cleaves dsRNA into ~23 nt siRNAs, demonstrated directly in vitro from immunoprecipitated enzyme. Reason: Phylogenetic inference is corroborated by direct experimental assay of dsRNase activity. Supporting Evidence: PMID:12482946 Dcr1p degraded double-stranded RNA into approximately 23 nucleotide fragments in vitro |
| GO:0005634 nucleus | IBA GO_REF:0000033 | ACCEPT | Summary: Core localization. Dcr1 is predominantly nuclear, retained there by its zinc-binding C-terminal domain, and this nuclear pool is required for RNAi-directed heterochromatin assembly. Reason: Phylogenetic inference corroborated by experimental nuclear localization and functional requirement for nuclear retention. Supporting Evidence: PMID:21847092 coordination of zinc by the C-terminus of Dcr1 is required for its nuclear retention and thus RNAi-mediated heterochromatin assembly |
| GO:0030422 siRNA processing | IBA GO_REF:0000033 | ACCEPT | Summary: Core biological process. Dcr1 generates the siRNAs that drive RNAi in fission yeast, processing dsRNA into ~21-23 nt small RNAs loaded onto Ago1. Reason: Well established as the sole siRNA-generating Dicer in S. pombe; supported by phylogeny and experiment. Supporting Evidence: PMID:27738016 which are cleaved by Dcr1 to small RNAs (21β23nt) |
| GO:0003677 DNA binding | IEA GO_REF:0000002 | MARK AS OVER ANNOTATED | Summary: The extended dsRBD of Dcr1 binds double-stranded DNA in vitro, but this generic DNA binding term is over-general and is better represented by the specific double-stranded DNA binding annotation; moreover dsDNA binding is dispensable for function. Reason: Generic DNA binding (InterPro-derived) is less informative than the experimentally supported double-stranded DNA binding term and overstates the role of nucleic acid binding by the dsRBD. Proposed replacements: double-stranded DNA binding Supporting Evidence: PMID:21847092 the extended dsRBD of Dcr1 showed a fairly strong affinity to a regular B-form DNA helix (Kd=1.5 ΞΌM) |
| GO:0004525 ribonuclease III activity | IEA GO_REF:0000002 | ACCEPT | Summary: Core molecular function, here supported electronically from the RNase III domain signature; consistent with direct experimental demonstration of dsRNase activity. Reason: InterPro2GO inference is correct and corroborated by experimental evidence. Supporting Evidence: PMID:12482946 tandem C-terminal ribonuclease (RNase) III signatures |
| GO:0005524 ATP binding | IEA GO_REF:0000002 | ACCEPT | Summary: Dcr1 contains a DExD/H-box helicase ATP-binding domain with a canonical ATP-binding Walker A-type motif (MRTGAGKT) and a DECH box, supporting ATP binding. Reason: Supported by conserved domain architecture and the identified ATP-binding motif. Supporting Evidence: PMID:12482946 an N-terminal helicase domain containing an ATP-binding site motif 32 M RTGAGKT 39 and a 145 DECH 148 box |
| GO:0005634 nucleus | IEA GO_REF:0000044 | ACCEPT | Summary: Core localization; nuclear localization is experimentally established and functionally important. Reason: Consistent with experimental nuclear localization and requirement for nuclear retention in RNAi. Supporting Evidence: PMID:21847092 coordination of zinc by the C-terminus of Dcr1 is required for its nuclear retention and thus RNAi-mediated heterochromatin assembly |
| GO:0005737 cytoplasm | IEA GO_REF:0000044 | KEEP AS NON CORE | Summary: Cytoplasmic localization is supported but is a secondary/contextual compartment relative to the core nuclear RNAi function. Reason: Real cytoplasmic pool but not the principal functional site. Supporting Evidence: PMID:21847092 disruption of zinc coordination renders Dcr1 mainly cytoplasmic and leads to remarkable changes in gene expression and loss of heterochromatin assembly |
| GO:0006396 RNA processing | IEA GO_REF:0000002 | MARK AS OVER ANNOTATED | Summary: Generic RNA processing is correct but uninformative; the specific siRNA processing term better captures Dcr1's role. Reason: Over-general parent term; superseded by siRNA processing (GO:0030422). Proposed replacements: siRNA processing |
| GO:0010467 gene expression | IEA GO_REF:0000117 | MARK AS OVER ANNOTATED | Summary: Extremely general term derived from a machine-learning model; not informative about Dcr1's specific role in gene silencing. Reason: Too general to be useful; the gene's role is in gene silencing, captured by more specific terms. |
| GO:0016787 hydrolase activity | IEA GO_REF:0000002 | MARK AS OVER ANNOTATED | Summary: Over-general parent of the specific ribonuclease III / RNA endonuclease activities that are separately annotated. Reason: Subsumed by GO:0004525 ribonuclease III activity. Proposed replacements: ribonuclease III activity |
| GO:0016891 RNA endonuclease activity producing 5'-phosphomonoesters, hydrolytic mechanism | IEA GO_REF:0000002 | ACCEPT | Summary: Accurate mechanistic molecular function for an RNase III enzyme, which leaves 5'-phosphate/3'-OH ends; consistent with the demonstrated dsRNA-cleaving activity. Reason: Mechanistically correct refinement of the catalytic activity, supported by RNase III domain assignment and in vitro cleavage. Supporting Evidence: PMID:12482946 Dcr1p degraded double-stranded RNA into approximately 23 nucleotide fragments in vitro |
| GO:0031047 regulatory ncRNA-mediated gene silencing | IEA GO_REF:0000117 | ACCEPT | Summary: Accurate higher-level process: Dcr1 mediates ncRNA (siRNA)-directed gene silencing. Captured more specifically by the siRNA/heterochromatin and CTGS terms. Reason: Correct parent process consistent with Dcr1's role in siRNA-directed silencing. Supporting Evidence: PMID:12193640 double-stranded RNA arising from centromeric repeats targets formation and maintenance of heterochromatin through RNAi |
| GO:0006363 termination of RNA polymerase I transcription | IMP PMID:25417108 Dicer promotes transcription termination at sites of replica... | KEEP AS NON CORE | Summary: Dcr1 promotes release of polymerases, including from antisense transcription of rRNA genes normally transcribed by Pol I, at sites of replication stress. This is a real but non-canonical, Dcr1-specific role distinct from its core siRNA-generating function. Reason: Experimentally supported genome-stability role, but peripheral to the central RNAi/siRNA function. Supporting Evidence: PMID:25417108 from antisense transcription of rRNA and tRNA genes, which are normally transcribed by Pol I and Pol III |
| GO:0006386 termination of RNA polymerase III transcription | IMP PMID:25417108 Dicer promotes transcription termination at sites of replica... | KEEP AS NON CORE | Summary: Dcr1 promotes Pol release from antisense transcription of tRNA genes (normally Pol III-transcribed) at replication-stress loci; a non-canonical genome-stability role. Reason: Supported but peripheral relative to the core siRNA/heterochromatin function. Supporting Evidence: PMID:25417108 from antisense transcription of rRNA and tRNA genes, which are normally transcribed by Pol I and Pol III |
| GO:0030466 silent mating-type cassette heterochromatin formation | EXP PMID:16246721 The nucleation and maintenance of heterochromatin by a histo... | ACCEPT | Summary: Core biological process. RNAi (including Dcr1) directs heterochromatin nucleation at the silent mating-type region in parallel with the Clr3 HDAC pathway. Reason: Established RNAi-directed silencing site; experimentally supported. Supporting Evidence: PMID:16246721 acts in a distinct pathway parallel to RNAi-directed heterochromatin nucleation to recruit Clr4 and mediate H3K9 methylation at the silent mating-type region and centromeres |
| GO:0033562 co-transcriptional gene silencing by RNA interference machinery | IMP PMID:17512405 RNAi-dependent and -independent RNA turnover mechanisms cont... | ACCEPT | Summary: Core biological process. Dcr1 is required for RNAi-mediated co-transcriptional gene silencing of centromeric repeats and contributes at the mating-type locus. Reason: Well-supported mechanism by which RNAi silences heterochromatic transcripts. Supporting Evidence: PMID:17512405 the RNAi pathway is required for heterochromatin-dependent silencing of transgene insertions at centromeric repeats and acts together with other pathways to silence transgenes at the silent mating-type locus |
| GO:0140727 siRNA-mediated pericentric heterochromatin formation | IMP PMID:12193640 Regulation of heterochromatic silencing and histone H3 lysin... | ACCEPT | Summary: Core biological process. dcr1 deletion causes accumulation of centromeric repeat transcripts, loss of H3K9 methylation and impaired centromere function, establishing Dcr1 as essential for siRNA-directed pericentric heterochromatin. Reason: Foundational genetic evidence for RNAi-directed pericentromeric heterochromatin. Supporting Evidence: PMID:12193640 This is accompanied by transcriptional de-repression of transgenes integrated at the centromere, loss of histone H3 lysine-9 methylation, and impairment of centromere function |
| GO:0140727 siRNA-mediated pericentric heterochromatin formation | EXP PMID:15615848 RNA-dependent RNA polymerase is an essential component of a ... | ACCEPT | Summary: Core biological process, corroborated by the self-enforcing RNAi loop in which Dicer, Argonaute and Rdp1 couple siRNA generation to heterochromatin assembly at centromeres. Reason: Independent experimental support for the siRNA-directed pericentromeric heterochromatin role. Supporting Evidence: PMID:15615848 factors involved in the RNA interference (RNAi) pathway including Argonaute, Dicer, and RNA-dependent RNA polymerase are required for heterochromatin assembly at centromeric repeats and the silent mating-type region |
| GO:0070317 negative regulation of G0 to G1 transition | IMP PMID:27738016 RNA interference is essential for cellular quiescence. | KEEP AS NON CORE | Summary: RNAi (including Dcr1) is essential for cellular quiescence; dcr1Ξ cells lose viability at G0 entry and during quiescence maintenance. This is a genuine but pleiotropic, non-core role downstream of the heterochromatin/Pol-release functions. Reason: Experimentally supported quiescence/cell-cycle role, secondary to the core RNAi function. Supporting Evidence: PMID:27738016 RNAi mutants lose viability at G0 entry and are unable to maintain long-term quiescence |
| GO:0004525 ribonuclease III activity | IDA PMID:12482946 Dicer is required for chromosome segregation and gene silenc... | ACCEPT | Summary: Core molecular function, directly demonstrated: immunoprecipitated Dcr1 cleaved dsRNA into ~23 nt fragments in vitro, absent from dcr1Ξ. Reason: Direct biochemical demonstration of the catalytic RNase III activity. Supporting Evidence: PMID:12482946 an β23-nt band was produced by Dcr1-HA immunoprecipitates when incubated with the human let-7 microRNA precursor (Fig. 3 B , lane 3), but not by immunoprecipitates prepared from dcr1 Ξ cells |
| GO:0005634 nucleus | EXP PMID:21847092 An extended dsRBD with a novel zinc-binding motif mediates n... | ACCEPT | Summary: Core localization; experimentally established nuclear localization requiring the zinc-binding C-terminus. Reason: Direct experimental evidence for nuclear localization and its functional importance. Supporting Evidence: PMID:21847092 coordination of zinc by the C-terminus of Dcr1 is required for its nuclear retention and thus RNAi-mediated heterochromatin assembly |
| GO:0000785 chromatin | IDA PMID:21151114 Dicer associates with chromatin to repress genome activity i... | ACCEPT | Summary: Dcr1 physically associates with chromatin (DamID), including euchromatic genes, ncRNA genes and retrotransposon LTRs, where association is sufficient to trigger silencing. Chromatin association is integral to its co-transcriptional silencing role. Reason: Direct DamID evidence that Dcr1 associates with chromatin to effect silencing. Supporting Evidence: PMID:21151114 the RNAi proteins Dcr1 and Rdp1 physically associate with some euchromatic genes, noncoding RNA genes and retrotransposon long terminal repeats |
| GO:0006369 termination of RNA polymerase II transcription | IMP PMID:25417108 Dicer promotes transcription termination at sites of replica... | KEEP AS NON CORE | Summary: Dcr1 specifically promotes release of Pol II from 3' ends of highly transcribed genes at sites of replication stress, independently of other RNAi components and of siRNA biogenesis. A genuine genome-stability role, distinct from the core siRNA function. Reason: Experimentally supported Dcr1-specific Pol II release, peripheral to canonical RNAi silencing. Supporting Evidence: PMID:25417108 Dcr1, but not other components of the canonical RNAi pathway, promotes the release of Pol II from the 3? end of highly transcribed genes |
| GO:0005634 nucleus | IDA PMID:25543137 Dicer and Hsp104 function in a negative feedback loop to con... | ACCEPT | Summary: Core localization; nuclear pool consistent with heterochromatin function. Reason: Direct evidence corroborating nuclear localization (relocalizes to cytoplasmic inclusions only when Hsp104 is absent). Supporting Evidence: PMID:25543137 In the absence of Hsp104, Dicer accumulates in cytoplasmic inclusions and heterochromatin becomes unstable at elevated temperatures |
| GO:0005737 cytoplasm | IDA PMID:25543137 Dicer and Hsp104 function in a negative feedback loop to con... | KEEP AS NON CORE | Summary: A cytoplasmic pool/inclusions are observed (notably when Hsp104 is absent or under heat stress); secondary to the principal nuclear function. Reason: Supported but contextual cytoplasmic localization, not the core functional compartment. Supporting Evidence: PMID:25543137 In the absence of Hsp104, Dicer accumulates in cytoplasmic inclusions and heterochromatin becomes unstable at elevated temperatures |
| GO:0033562 co-transcriptional gene silencing by RNA interference machinery | IMP PMID:25543137 Dicer and Hsp104 function in a negative feedback loop to con... | ACCEPT | Summary: Core biological process; Dcr1 maintains RNAi-mediated co-transcriptional silencing and heterochromatin stability. Reason: Supported; consistent with the established CTGS role of Dcr1. Supporting Evidence: PMID:25543137 In the absence of Hsp104, Dicer accumulates in cytoplasmic inclusions and heterochromatin becomes unstable at elevated temperatures |
| GO:0005634 nucleus | IDA PMID:22895252 A novel RNAi protein, Dsh1, assembles RNAi machinery on chro... | ACCEPT | Summary: Core localization; Dcr1 acts in the nucleus on chromatin with RDRC and Dsh1 to amplify heterochromatic siRNA. Reason: Supported nuclear site of action consistent with siRNA amplification on chromatin. Supporting Evidence: PMID:22895252 Dsh1 interacts with Dcr1 and RDRC and couples the reactions of both proteins to the effective production of siRNA in vivo |
| GO:0140727 siRNA-mediated pericentric heterochromatin formation | IMP PMID:22895252 A novel RNAi protein, Dsh1, assembles RNAi machinery on chro... | ACCEPT | Summary: Core biological process. Dcr1, with RDRC and the novel factor Dsh1, amplifies pericentromeric siRNA required for stable heterochromatin formation. Reason: Supported role in siRNA amplification driving pericentric heterochromatin. Supporting Evidence: PMID:22895252 siRNA is amplified by the RNA-dependent RNA polymerase complex (RDRC) and the endoribonuclease Dcr1, which synthesizes dsRNA and processes the dsRNA, respectively. The amplification is essential for stable heterochromatin formation |
| GO:1990188 euchromatin binding | IDA PMID:22431512 RNAi keeps Atf1-bound stress response genes in check at nucl... | KEEP AS NON CORE | Summary: Dcr1 associates with euchromatic, Atf1-bound stress-response genes at nuclear pores, where RNAi-mediated CTGS keeps them in check. A genuine but specialized association relative to the core pericentromeric role. Reason: Experimentally supported euchromatin association, but a specialized/contextual activity. Supporting Evidence: PMID:22431512 RNAi components interact with chromatin at nuclear pores to keep stress response genes in check |
| GO:0000791 euchromatin | IDA PMID:22431512 RNAi keeps Atf1-bound stress response genes in check at nucl... | KEEP AS NON CORE | Summary: Dcr1 is active at euchromatic stress-response genes (CTGS at nuclear pores); a specialized localization beyond its core heterochromatic sites. Reason: Supported but specialized euchromatic site of action. Supporting Evidence: PMID:22431512 RNAi-mediated CTGS represses stress-inducible genes by degrading mRNAs under noninduced conditions |
| GO:0005721 pericentric heterochromatin | IDA PMID:21151114 Dicer associates with chromatin to repress genome activity i... | ACCEPT | Summary: Core localization. Dcr1 associates with pericentromeric chromatin, the principal site of RNAi-directed heterochromatin assembly. Reason: Supported by physical chromatin association data and consistent with the central RNAi function at centromeres. Supporting Evidence: PMID:21151114 the RNA interference (RNAi) pathway is required to generate small interfering RNAs (siRNAs) that mediate heterochromatic silencing of centromeric repeats |
| GO:0005829 cytosol | IDA PMID:22431512 RNAi keeps Atf1-bound stress response genes in check at nucl... | KEEP AS NON CORE | Summary: A cytosolic pool is reported (and Dicer is exported to the cytoplasm under chronic heat stress); secondary to the core nuclear function. Reason: Supported but contextual cytosolic localization. Supporting Evidence: PMID:22431512 a Dicer thermoswitch deports Dicer to the cytoplasm, thereby disrupting CTGS and enabling expression of genes implicated in the acquisition of thermotolerance |
| GO:0033562 co-transcriptional gene silencing by RNA interference machinery | IMP PMID:21151114 Dicer associates with chromatin to repress genome activity i... | ACCEPT | Summary: Core biological process. Physical association of Dcr1 with chromatin is sufficient to trigger a co-transcriptional silencing response. Reason: Strong experimental support (DamID + silencing assays) for the CTGS role. Supporting Evidence: PMID:21151114 Physical association of RNAi with chromatin is sufficient to trigger a silencing response but not to assemble heterochromatin |
| GO:0033562 co-transcriptional gene silencing by RNA interference machinery | IMP PMID:22431512 RNAi keeps Atf1-bound stress response genes in check at nucl... | ACCEPT | Summary: Core biological process; RNAi-mediated CTGS by Dcr1 represses stress-inducible genes at nuclear pores under non-induced conditions. Reason: Independent experimental support for the CTGS role of Dcr1. Supporting Evidence: PMID:22431512 RNAi-mediated CTGS represses stress-inducible genes by degrading mRNAs under noninduced conditions |
| GO:0034399 nuclear periphery | IDA PMID:22431512 RNAi keeps Atf1-bound stress response genes in check at nucl... | KEEP AS NON CORE | Summary: Dcr1 acts at the nuclear periphery / nuclear pores, where RNAi components engage Atf1-bound stress genes. A specific sub-nuclear localization supporting CTGS. Reason: Supported sub-nuclear localization; a specialized site within the broader nuclear function. Supporting Evidence: PMID:22431512 RNAi components interact with chromatin at nuclear pores to keep stress response genes in check |
| GO:0003690 double-stranded DNA binding | IDA PMID:21847092 An extended dsRBD with a novel zinc-binding motif mediates n... | KEEP AS NON CORE | Summary: The extended dsRBD binds B-form dsDNA in vitro (Kd ~1.5 uM). This binding is real but dispensable for Dcr1 function (RNAi/heterochromatin), so it is a non-core property. Reason: Direct in vitro binding demonstrated, but shown to be dispensable for proper functioning of Dcr1. Supporting Evidence: PMID:21847092 the extended dsRBD of Dcr1 showed a fairly strong affinity to a regular B-form DNA helix (Kd=1.5 ΞΌM) |
| GO:0003725 double-stranded RNA binding | IDA PMID:21847092 An extended dsRBD with a novel zinc-binding motif mediates n... | ACCEPT | Summary: Core molecular function consistent with Dicer biology: the dsRBD binds A-form dsRNA strongly (Kd ~0.8 uM). Note the C-terminal dsRBD's dsRNA binding is dispensable for function, but Dcr1 as a whole binds and processes dsRNA substrates. Reason: Directly demonstrated dsRNA binding; central to recognition/processing of dsRNA substrates. Supporting Evidence: PMID:21847092 binds rather strongly to a regular A-form RNA helix (Kd=0.8 ΞΌM, with two dsRBDs bound per 24 bp dsRNA helix) |
| GO:0008270 zinc ion binding | IDA PMID:21847092 An extended dsRBD with a novel zinc-binding motif mediates n... | ACCEPT | Summary: Core structural molecular function. The C-terminal extended dsRBD coordinates one Zn2+ via a novel CHCC motif (C1275, H1312, C1350, C1352); zinc coordination is required for nuclear retention and thus RNAi-mediated heterochromatin assembly. Reason: Directly demonstrated by NMR structure/AAS/MS and functionally validated by SHSS mutants. Supporting Evidence: PMID:21847092 the Dcr1 C-terminal domain containing the dsRBD and the C33 indeed include one zinc ion per molecule |
| GO:0010629 negative regulation of gene expression | IMP PMID:17981703 Involvement of Dcr1 in post-transcriptional regulation of ge... | KEEP AS NON CORE | Summary: Dcr1 contributes to post-transcriptional regulation of certain proteins (e.g. glycolytic enzymes), consistent with negative regulation of gene expression. This is supported but a general/peripheral aspect relative to the core silencing functions. Reason: Supported but a general, secondary regulatory role; the specific silencing mechanisms are captured by CTGS/heterochromatin terms. Supporting Evidence: PMID:17981703 compatible with the existence of a post-transcriptional gene regulatory mechanism involving Dcr1 or a Dcr1 complex |
| GO:0005737 cytoplasm | HDA PMID:16823372 ORFeome cloning and global analysis of protein localization ... | KEEP AS NON CORE | Summary: High-throughput (ORFeome GFP) localization detected a cytoplasmic signal; a real but secondary compartment relative to the core nuclear function. Reason: HDA-level evidence for a cytoplasmic pool; not the principal functional site. Supporting Evidence: PMID:16823372 ORFeome cloning and global analysis of protein localization in the fission yeast Schizosaccharomyces pombe |
| GO:0005829 cytosol | HDA PMID:16823372 ORFeome cloning and global analysis of protein localization ... | KEEP AS NON CORE | Summary: High-throughput localization reported a cytosolic pool; secondary to the core nuclear RNAi function. Reason: HDA-level evidence for a cytosolic pool; contextual rather than core. Supporting Evidence: PMID:16823372 ORFeome cloning and global analysis of protein localization in the fission yeast Schizosaccharomyces pombe |
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Download this section (compressed HTML)Q: Is the transcription-termination / Pol-release activity of Dcr1 a direct biochemical function of the enzyme (e.g. cleavage of nascent RNA at collision sites) or an indirect consequence of chromatin association, given it is independent of siRNA biogenesis?
Q: What is the nuclear retention factor recognized by the conserved surface of the extended dsRBD, and how does it integrate Dcr1 into the chromatin-bound RNAi machinery?
Experiment: Use a catalytically dead RNase III Dcr1 (point mutations in RIIIa/RIIIb Mg2+-binding residues) versus zinc-coordination mutants to separate the contribution of siRNA production from the chromatin/Pol-release functions genome-wide (ChIP-seq of Pol I/II/III, H3K9me2, and small-RNA-seq).
Experiment: Reconstitute Dcr1-RDRC-Dsh1 on nucleosomal templates in vitro to test whether Dcr1 can promote polymerase release independently of siRNA generation, directly probing the Dcr1-specific termination activity.
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