{"gene":"WDR74","run_date":"2026-06-11T09:02:06","timeline":{"discoveries":[{"year":2008,"finding":"The AAA-ATPase Rix7 (yeast ortholog of NVL2) is required for the energy-dependent release of Nsa1 (yeast ortholog of WDR74) from a discrete late-nucleolar pre-60S particle; in rix7 mutants, Nsa1 cannot dissociate from pre-60S particles and aberrantly accumulates in the cytoplasm associated with aberrant 60S subunits. Rix7 interacts genetically with Nsa1 and is targeted to the Nsa1-defined preribosomal particle.","method":"Genetic epistasis, in vivo localization (fluorescence microscopy), co-immunoprecipitation, yeast mutant analysis","journal":"The Journal of cell biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal genetic and biochemical evidence in yeast ortholog, multiple orthogonal methods (genetics, localization, co-IP), independently foundational study","pmids":["18559667"],"is_preprint":false},{"year":2013,"finding":"Nsa1 (WDR74 ortholog) associates with a late-nucleolar pre-60S particle that also contains the DEAD-box RNA helicase Mak5; mutant alleles of MAK5, NOP1, and NOP4 bypass the essential requirement for Nsa1, placing Nsa1 in a pathway involving these factors. Dominant-negative Rix7 retains its growth defect even in the absence of Nsa1, indicating Rix7 has additional nuclear substrates beyond Nsa1.","method":"Genetic epistasis (bypass suppressor screen), co-immunoprecipitation, synthetic lethality screens, yeast mutant analysis","journal":"PloS one","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal genetic methods (bypass suppression, synthetic lethality, dominant-negative analysis) in yeast ortholog, replicated with prior work","pmids":["24312670"],"is_preprint":false},{"year":2015,"finding":"WDR74 is a component of the MTR4-exosome complex in the nucleolus; the AAA-ATPase NVL2 (human ortholog of Rix7) uses its ATPase activity to dissociate WDR74 from this complex. Knockdown of WDR74 decreases 60S ribosome levels in human cells.","method":"Proteomic screen (mass spectrometry), co-immunoprecipitation, ATPase-deficient NVL2 mutant analysis, siRNA knockdown with ribosome fractionation","journal":"Biochemical and biophysical research communications","confidence":"High","confidence_rationale":"Tier 2 / Moderate — co-IP with mass spectrometry, ATPase mutant validation, loss-of-function with defined ribosome phenotype; single lab but multiple orthogonal methods","pmids":["26456651"],"is_preprint":false},{"year":2017,"finding":"WDR74 is required for early pre-rRNA cleavage within ITS1 in the 60S ribosome biogenesis pathway; knockdown of WDR74 causes significant defects in this cleavage step. ATPase-deficient NVL2 prevents dissociation of WDR74 from the MTR4-exosome complex, causing partial migration of WDR74 from the nucleolus to nucleoplasm and an increased interaction between WDR74 and MTR4 in the nucleoplasm, which also produces the same early ITS1 processing defect.","method":"siRNA knockdown with pre-rRNA processing analysis (Northern blot), ATPase-deficient NVL2 mutant, in situ proximity ligation assay, subcellular fractionation/localization","journal":"Biochemical and biophysical research communications","confidence":"High","confidence_rationale":"Tier 2 / Moderate — loss-of-function with specific molecular phenotype (ITS1 cleavage), mutant validation, proximity ligation assay; single lab, multiple orthogonal methods","pmids":["29107693"],"is_preprint":false},{"year":2018,"finding":"The full-length structure of yeast Nsa1 (WDR74 ortholog) was determined using a hybrid X-ray crystallography / SAXS approach: the N-terminal WD40 domain was solved by X-ray crystallography, and the disordered C-terminus was modeled by SAXS with rigid body and ab initio modeling, revealing the quaternary structure of the entire protein.","method":"X-ray crystallography (WD40 domain), SAXS (full-length solution structure), ab initio and rigid body modeling","journal":"Journal of visualized experiments : JoVE","confidence":"High","confidence_rationale":"Tier 1 / Moderate — experimental structure determination using two complementary structural methods on purified recombinant protein; single study but rigorous hybrid approach","pmids":["29364241"],"is_preprint":false},{"year":2018,"finding":"WDR74 functions as a transcriptional coactivator for Smad proteins in the canonical TGF-β signaling pathway; WDR74 directly interacts with Smad proteins and enhances TGF-β-mediated phosphorylation and nuclear accumulation of Smad2 and Smad3, leading to stronger transcriptional responses.","method":"Co-immunoprecipitation (direct interaction with Smad proteins), Western blot (Smad2/3 phosphorylation), nuclear fractionation, gain- and loss-of-function assays with TGF-β reporter","journal":"Journal of genetics and genomics = Yi chuan xue bao","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP for direct interaction, phosphorylation assay, nuclear accumulation measurement; single lab with multiple methods","pmids":["30594465"],"is_preprint":false},{"year":2019,"finding":"WDR74 promotes nuclear β-catenin accumulation and activates downstream Wnt-responsive genes in lung cancer cells; gain- and loss-of-function studies showed WDR74 regulates cell proliferation, cell cycle, chemoresistance, and aggressiveness via the Wnt/β-catenin signaling pathway.","method":"Gain- and loss-of-function (overexpression and knockout), Western blot (β-catenin nuclear accumulation), Wnt reporter assays, xenograft mouse model","journal":"Cancer letters","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — functional KO/OE with defined pathway placement and in vivo validation; single lab, multiple methods","pmids":["31838084"],"is_preprint":false},{"year":2020,"finding":"WDR74 modulates RPL5 protein levels, which in turn regulates MDM2 activity and protects p53 from MDM2-mediated ubiquitination and degradation; WDR74 thus controls the RPL5-MDM2-p53 pathway to promote melanoma cell proliferation, apoptosis resistance, and metastasis.","method":"iTRAQ proteomic screening, gain- and loss-of-function approaches, Western blot (RPL5, MDM2, p53 ubiquitination), in vivo xenograft and metastasis models","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — proteomic identification plus functional validation with multiple molecular readouts; single lab","pmids":["32005977"],"is_preprint":false},{"year":2011,"finding":"Wdr74 is essential for blastocyst formation in mouse preimplantation development; Wdr74 knockdown causes embryos to arrest at the morula stage with activated Trp53-dependent apoptosis and global reduction of RNA polymerase I, II, and III transcripts. Blocking Trp53 function rescues blastocyst formation in Wdr74-deficient embryos, placing Wdr74 upstream of Trp53-dependent apoptosis.","method":"RNAi knockdown in mouse embryos, RT-qPCR (RNA Pol I/II/III transcripts), genetic epistasis (Trp53 rescue), embryo phenotypic analysis","journal":"PloS one","confidence":"High","confidence_rationale":"Tier 2 / Strong — RNAi loss-of-function with specific molecular phenotype plus genetic epistasis rescue establishing pathway position; multiple orthogonal methods","pmids":["21799883"],"is_preprint":false},{"year":2024,"finding":"WDR74 functions as part of a pre-ribosomal subcomplex termed the 'WDR74 module', consisting of WDR74, RPF1, MAK16, and RRP1; each component of this module is mutually required for interaction of the others with MTR4, and all components are required for accurate pre-rRNA cleavage during 60S biogenesis. Impaired NVL2-mediated release of WDR74 from the MTR4-exosome complex prevents MTR4 from recruiting PICT1, an MTR4 adaptor required for 3'-end maturation of 5.8S rRNA.","method":"Co-immunoprecipitation combined with mass spectrometry, siRNA knockdown, pre-rRNA processing analysis, interaction mapping","journal":"Biochemical and biophysical research communications","confidence":"High","confidence_rationale":"Tier 2 / Moderate — co-IP/MS identification of the subcomplex, combined with loss-of-function showing mutual dependency and specific rRNA processing defects; single lab, multiple orthogonal methods","pmids":["39706051"],"is_preprint":false},{"year":2022,"finding":"WDR74 interacts with Smad2/3 in macrophages (co-immunoprecipitation) and promotes TGF-β/Smad pathway activation; WDR74 overexpression increases Smad2/3 phosphorylation and promotes M2 macrophage polarization and ECM production in a diabetic foot ulcer mouse model. These effects are reversed by the TGF-β receptor inhibitor LY2109761.","method":"Co-immunoprecipitation (WDR74-Smad2/3 interaction), gain- and loss-of-function (overexpression/knockdown), Western blot (Smad2/3 phosphorylation), immunofluorescence, mouse DFU model","journal":"Cell biology and toxicology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP confirms direct interaction, phosphorylation assay, pharmacological rescue; single lab, multiple methods","pmids":["35982296"],"is_preprint":false},{"year":2025,"finding":"WDR74 deficiency in mouse embryos (generated by CRISPR-Cas9) leads to impaired 60S ribosome biogenesis with significant reduction in large ribosomal subunit proteins (notably RPL24 and RPL26) but not small subunit proteins, and blocks cell division progression beyond the morula stage.","method":"CRISPR-Cas9 knockout, label-free quantitative proteomics, cell division phenotypic analysis","journal":"Genes to cells : devoted to molecular & cellular mechanisms","confidence":"High","confidence_rationale":"Tier 2 / Moderate — CRISPR KO with quantitative proteomics revealing specific large subunit protein reduction; single lab, rigorous genome editing plus proteomics","pmids":["39840464"],"is_preprint":false},{"year":2021,"finding":"WDR74 decreases phosphorylation of β-catenin and promotes its nuclear accumulation in colorectal cancer cells, activating the Wnt/β-catenin signaling pathway; blocking this pathway with XAV-939 reverses WDR74-mediated effects on proliferation, migration, and invasion.","method":"siRNA knockdown, Western blot (β-catenin phosphorylation and nuclear localization), XAV-939 pharmacological rescue, cell proliferation and invasion assays","journal":"Open life sciences","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — loss-of-function with pathway placement confirmed by pharmacological rescue; single lab, corroborates findings from PMID:31838084","pmids":["34553072"],"is_preprint":false},{"year":2024,"finding":"ATF5 transcriptionally upregulates WDR74, and WDR74 in turn enhances β-catenin nuclear translocation to promote stemness in gastric cancer; METTL14 suppresses this axis by promoting m6A-mediated degradation of ATF5 mRNA. ChIP assays confirmed ATF5 binds the WDR74 promoter.","method":"ChIP assay (ATF5 binding to WDR74 promoter), MeRIP-qPCR (m6A modification of ATF5), Western blot (β-catenin nuclear translocation), rescue/overexpression assays","journal":"Cancer science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP confirms direct transcriptional regulation, MeRIP validates upstream epigenetic mechanism, functional rescue validates pathway; single lab","pmids":["39497511"],"is_preprint":false},{"year":2025,"finding":"CAPG promotes WDR74 transcription, and WDR74 in turn modulates the interaction between p53 and MDM2, resulting in p53 ubiquitination and degradation, thereby inhibiting ferroptosis in hepatocellular carcinoma. This was supported by co-immunoprecipitation and ubiquitination assays.","method":"Co-immunoprecipitation, ubiquitination assays, ChIP sequencing, RNA sequencing, gain- and loss-of-function, xenograft model","journal":"International journal of biological sciences","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP and ubiquitination assays establish mechanism; single lab, corroborates RPL5-MDM2-p53 axis from PMID:32005977","pmids":["40959275"],"is_preprint":false},{"year":2025,"finding":"SNHG1 (a lncRNA encoded adjacent to WDR74) promotes WDR74 transcription in cis by recruiting EWSR1 to the WDR74 promoter region; ChIP-qPCR confirmed EWSR1 binding at the WDR74 promoter, establishing a trans-regulatory mechanism upstream of WDR74 expression in osteosarcoma.","method":"ChIP-qPCR (EWSR1 binding at WDR74 promoter), RNA pulldown, RNA immunoprecipitation, actinomycin D stability assay","journal":"Frontiers in oncology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP-qPCR confirms direct EWSR1 binding at WDR74 promoter; single lab, multiple supporting methods","pmids":["40510136"],"is_preprint":false}],"current_model":"WDR74 (Nsa1 in yeast) is a nucleolar WD40-domain protein that functions primarily as an essential pre-60S ribosome biogenesis factor: it resides in a pre-ribosomal subcomplex (with RPF1, MAK16, and RRP1) that coordinates early ITS1 pre-rRNA cleavage and associates with the MTR4-nuclear exosome complex, from which it is released by the AAA-ATPase NVL2/Rix7 in an ATP hydrolysis-dependent manner to permit downstream 5.8S rRNA maturation; loss of WDR74 reduces 60S ribosome levels, activates Trp53-dependent apoptosis, and blocks mammalian cell division. Beyond ribosome biogenesis, WDR74 also functions as a coactivator of TGF-β/Smad signaling through direct interaction with Smad2/3, and promotes Wnt/β-catenin signaling by preventing β-catenin phosphorylation and driving its nuclear accumulation; additionally, WDR74 controls the RPL5-MDM2-p53 axis to regulate p53 stability and ferroptosis resistance in cancer contexts."},"narrative":{"mechanistic_narrative":"WDR74 (yeast Nsa1) is an essential nucleolar WD40-domain protein that acts as a pre-60S ribosome biogenesis factor coordinating early pre-rRNA processing during large ribosomal subunit assembly [PMID:29107693, PMID:39840464]. It functions within a discrete pre-ribosomal subcomplex—the 'WDR74 module' comprising WDR74, RPF1, MAK16, and RRP1—in which each component is mutually required for association with the MTR4-nuclear exosome complex and for accurate ITS1 cleavage in the 60S pathway [PMID:39706051, PMID:29107693]. WDR74 is released from the MTR4-exosome complex by the AAA-ATPase NVL2 (yeast Rix7) in an ATP-hydrolysis-dependent manner; blocking this release with ATPase-deficient NVL2 retains WDR74 on MTR4, mislocalizes it to the nucleoplasm, and prevents MTR4 from recruiting the adaptor PICT1 required for 3'-end maturation of 5.8S rRNA [PMID:18559667, PMID:26456651, PMID:39706051]. The full-length architecture of the protein comprises an N-terminal WD40 domain and a disordered C-terminus [PMID:29364241]. Loss of WDR74 reduces 60S subunit levels and large-subunit ribosomal proteins, arrests cells beyond the morula stage, and activates Trp53-dependent apoptosis that can be rescued by blocking Trp53 [PMID:21799883, PMID:39840464]. Beyond ribosome biogenesis, WDR74 acts as a coactivator of TGF-β/Smad signaling through direct interaction with Smad2/3, enhancing their phosphorylation and nuclear accumulation [PMID:30594465, PMID:35982296]; promotes Wnt/β-catenin signaling by reducing β-catenin phosphorylation and driving its nuclear accumulation [PMID:31838084, PMID:34553072]; and controls the RPL5-MDM2-p53 axis to stabilize or destabilize p53, influencing apoptosis and ferroptosis in cancer contexts [PMID:32005977, PMID:40959275].","teleology":[{"year":2008,"claim":"Established the regulated dissociation step at the heart of WDR74 function—that an AAA-ATPase actively releases Nsa1 from a late pre-60S particle—defining Nsa1 as a transiently associated assembly factor rather than a structural ribosome component.","evidence":"Genetic epistasis, fluorescence localization, and co-IP in yeast rix7 mutants","pmids":["18559667"],"confidence":"High","gaps":["Did not define the molecular signal triggering release","Yeast ortholog; human relevance not yet tested","No structural basis for the Rix7-Nsa1 interaction"]},{"year":2011,"claim":"Placed WDR74 upstream of Trp53-dependent apoptosis in development by showing its loss arrests embryos and globally reduces Pol I/II/III transcripts, with p53 blockade rescuing the phenotype.","evidence":"RNAi knockdown in mouse preimplantation embryos with Trp53 rescue and RT-qPCR","pmids":["21799883"],"confidence":"High","gaps":["Did not establish the direct molecular link between WDR74 loss and p53 activation","Global transcript reduction could be secondary to general nucleolar stress"]},{"year":2013,"claim":"Mapped Nsa1 into a genetic pathway with Mak5, Nop1, and Nop4, refining where in 60S assembly the factor acts and showing Rix7 has substrates beyond Nsa1.","evidence":"Bypass suppressor and synthetic lethality screens, co-IP, dominant-negative analysis in yeast","pmids":["24312670"],"confidence":"High","gaps":["Genetic interactions do not establish direct physical contacts","Additional Rix7 substrates left unidentified"]},{"year":2015,"claim":"Translated the yeast model to human cells, identifying WDR74 as a component of the MTR4-exosome complex released by NVL2 ATPase activity, with knockdown reducing 60S levels.","evidence":"Mass-spectrometry proteomics, co-IP, ATPase-deficient NVL2 mutant, siRNA with ribosome fractionation","pmids":["26456651"],"confidence":"High","gaps":["Did not resolve the specific rRNA processing step affected","Stoichiometry within the MTR4-exosome complex unknown"]},{"year":2017,"claim":"Pinpointed the molecular defect as failure of early ITS1 pre-rRNA cleavage and linked NVL2-blocked WDR74 release to nucleoplasmic mislocalization and the same processing defect.","evidence":"siRNA with Northern blot pre-rRNA analysis, ATPase-deficient NVL2, proximity ligation assay, fractionation","pmids":["29107693"],"confidence":"High","gaps":["Did not identify the nuclease executing ITS1 cleavage","Whether WDR74 acts catalytically or as a scaffold unresolved"]},{"year":2018,"claim":"Provided the full-length structural model of Nsa1, defining an N-terminal WD40 domain and a disordered C-terminus that together explain its quaternary organization.","evidence":"Hybrid X-ray crystallography (WD40 domain) and SAXS with ab initio/rigid-body modeling on recombinant protein","pmids":["29364241"],"confidence":"High","gaps":["No structure of WDR74 within the pre-60S particle or MTR4 complex","Function of the disordered C-terminus not defined"]},{"year":2018,"claim":"Revealed a moonlighting role outside ribosome biogenesis, identifying WDR74 as a Smad coactivator that directly binds Smad2/3 and enhances their phosphorylation and nuclear accumulation.","evidence":"Co-IP, Western blot for Smad2/3 phosphorylation, nuclear fractionation, TGF-β reporter gain/loss-of-function","pmids":["30594465"],"confidence":"Medium","gaps":["Mechanism by which a nucleolar factor enhances Smad phosphorylation unclear","Direct interaction not validated by reciprocal or structural methods"]},{"year":2019,"claim":"Connected WDR74 to Wnt/β-catenin signaling, showing it drives nuclear β-catenin accumulation and tumor-promoting phenotypes in lung cancer.","evidence":"Overexpression/knockout, Western blot, Wnt reporter assays, xenograft model","pmids":["31838084"],"confidence":"Medium","gaps":["Did not define how WDR74 affects β-catenin stability mechanistically","Direct versus indirect action on the pathway unresolved"]},{"year":2020,"claim":"Defined a ribosomal-protein-mediated route to p53 control, showing WDR74 modulates RPL5 to restrain MDM2 and protect p53 from degradation in melanoma.","evidence":"iTRAQ proteomics, gain/loss-of-function, ubiquitination Western blots, xenograft and metastasis models","pmids":["32005977"],"confidence":"Medium","gaps":["Directionality of p53 regulation appears context-dependent across cancers","Whether RPL5 modulation reflects general ribosome biogenesis disruption unclear"]},{"year":2022,"claim":"Extended the TGF-β/Smad coactivator role to macrophage biology and tissue repair, with pharmacological receptor blockade reversing WDR74-driven effects.","evidence":"Co-IP, gain/loss-of-function, Western blot, immunofluorescence, LY2109761 rescue in a mouse DFU model","pmids":["35982296"],"confidence":"Medium","gaps":["Does not distinguish direct coactivation from broader transcriptional effects","Single-lab confirmation of WDR74-Smad2/3 interaction"]},{"year":2024,"claim":"Defined the 'WDR74 module' (WDR74-RPF1-MAK16-RRP1) as a mutually dependent unit required for MTR4 association and showed NVL2-mediated release enables MTR4 recruitment of PICT1 for 5.8S rRNA 3'-end maturation.","evidence":"Co-IP/MS, siRNA, pre-rRNA processing analysis, interaction mapping","pmids":["39706051"],"confidence":"High","gaps":["Structural organization of the module within pre-60S not resolved","Order of assembly and release events not fully mapped"]},{"year":2024,"claim":"Identified upstream transcriptional control of WDR74 by ATF5, itself regulated by METTL14-mediated m6A, linking WDR74 expression to β-catenin-driven stemness in gastric cancer.","evidence":"ChIP for ATF5 promoter binding, MeRIP-qPCR, Western blot, rescue assays","pmids":["39497511"],"confidence":"Medium","gaps":["Does not address whether WDR74's ribosomal function contributes to stemness","Single-lab regulatory axis"]},{"year":2025,"claim":"Confirmed in a CRISPR-knockout embryo model that WDR74 loss selectively impairs 60S biogenesis, reducing large-subunit proteins (RPL24, RPL26) but not small-subunit proteins, and blocks cell division beyond the morula stage.","evidence":"CRISPR-Cas9 knockout, label-free quantitative proteomics, division phenotyping","pmids":["39840464"],"confidence":"High","gaps":["Does not establish whether division arrest is purely ribosome-dependent or via p53","Mechanism of selective large-subunit protein loss not defined"]},{"year":2025,"claim":"Showed WDR74 modulates the p53-MDM2 interaction to promote p53 ubiquitination and degradation, inhibiting ferroptosis in hepatocellular carcinoma, with CAPG acting as an upstream transcriptional driver.","evidence":"Co-IP, ubiquitination assays, ChIP-seq, RNA-seq, gain/loss-of-function, xenograft","pmids":["40959275"],"confidence":"Medium","gaps":["p53 outcome opposite to the melanoma RPL5-MDM2 model, context dependence unexplained","Direct versus indirect effect on the p53-MDM2 interface not resolved"]},{"year":2025,"claim":"Revealed cis/trans transcriptional regulation of WDR74 by the adjacent lncRNA SNHG1, which recruits EWSR1 to the WDR74 promoter in osteosarcoma.","evidence":"ChIP-qPCR for EWSR1 binding, RNA pulldown, RIP, actinomycin D stability assay","pmids":["40510136"],"confidence":"Medium","gaps":["Does not link this regulation to WDR74's downstream ribosomal or signaling functions","Single-lab regulatory mechanism"]},{"year":null,"claim":"It remains unresolved how WDR74's core nucleolar ribosome biogenesis function mechanistically connects to its reported roles in TGF-β/Smad, Wnt/β-catenin, and p53/MDM2 signaling, and whether the cancer-context signaling roles are direct or downstream consequences of perturbed ribosome assembly.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified mechanism linking nucleolar and signaling functions","Opposite p53 directionality across cancer models unexplained","No structure of WDR74 engaged with signaling partners"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140110","term_label":"transcription regulator activity","supporting_discovery_ids":[5,10]},{"term_id":"GO:0060089","term_label":"molecular transducer activity","supporting_discovery_ids":[6,12]}],"localization":[{"term_id":"GO:0005730","term_label":"nucleolus","supporting_discovery_ids":[0,2,3]},{"term_id":"GO:0005654","term_label":"nucleoplasm","supporting_discovery_ids":[3]}],"pathway":[{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[2,3,9]},{"term_id":"R-HSA-1852241","term_label":"Organelle biogenesis and maintenance","supporting_discovery_ids":[2,11]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[5,6]}],"complexes":["WDR74 module (WDR74-RPF1-MAK16-RRP1)","MTR4-nuclear exosome complex","pre-60S ribosomal particle"],"partners":["MTR4","NVL2","RPF1","MAK16","RRP1","SMAD2","SMAD3","RPL5"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q6RFH5","full_name":"WD repeat-containing protein 74","aliases":["NOP seven-associated protein 1"],"length_aa":385,"mass_kda":42.4,"function":"Regulatory protein of the MTREX-exosome complex involved in the synthesis of the 60S ribosomal subunit (PubMed:26456651). Participates in an early cleavage of the pre-rRNA processing pathway in cooperation with NVL (PubMed:29107693). Required for blastocyst formation, is necessary for RNA transcription, processing and/or stability during preimplantation development (By similarity)","subcellular_location":"Nucleus, nucleolus; Nucleus","url":"https://www.uniprot.org/uniprotkb/Q6RFH5/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/WDR74","classification":"Common Essential","n_dependent_lines":1204,"n_total_lines":1208,"dependency_fraction":0.9966887417218543},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"VPS35","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/WDR74","total_profiled":1310},"omim":[{"mim_id":"617947","title":"WD REPEAT-CONTAINING PROTEIN 74; WDR74","url":"https://www.omim.org/entry/617947"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Nucleoplasm","reliability":"Supported"},{"location":"Nucleoli","reliability":"Supported"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in all","driving_tissues":[{"tissue":"bone marrow","ntpm":119.2}],"url":"https://www.proteinatlas.org/search/WDR74"},"hgnc":{"alias_symbol":["FLJ10439","Nsa1"],"prev_symbol":[]},"alphafold":{"accession":"Q6RFH5","domains":[{"cath_id":"2.130.10.10","chopping":"2-320","consensus_level":"medium","plddt":90.5449,"start":2,"end":320}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q6RFH5","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q6RFH5-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q6RFH5-F1-predicted_aligned_error_v6.png","plddt_mean":83.31},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=WDR74","jax_strain_url":"https://www.jax.org/strain/search?query=WDR74"},"sequence":{"accession":"Q6RFH5","fasta_url":"https://rest.uniprot.org/uniprotkb/Q6RFH5.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q6RFH5/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q6RFH5"}},"corpus_meta":[{"pmid":"35982296","id":"PMC_35982296","title":"WDR74 facilitates TGF-β/Smad pathway activation to promote M2 macrophage polarization and diabetic foot ulcer wound healing in mice.","date":"2022","source":"Cell biology and toxicology","url":"https://pubmed.ncbi.nlm.nih.gov/35982296","citation_count":73,"is_preprint":false},{"pmid":"18559667","id":"PMC_18559667","title":"The AAA ATPase Rix7 powers progression of ribosome biogenesis by stripping Nsa1 from pre-60S particles.","date":"2008","source":"The Journal of cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/18559667","citation_count":73,"is_preprint":false},{"pmid":"31838084","id":"PMC_31838084","title":"WDR74 induces nuclear β-catenin accumulation and activates Wnt-responsive genes to promote lung cancer growth and metastasis.","date":"2019","source":"Cancer letters","url":"https://pubmed.ncbi.nlm.nih.gov/31838084","citation_count":30,"is_preprint":false},{"pmid":"24312670","id":"PMC_24312670","title":"Mak5 and Ebp2 act together on early pre-60S particles and their reduced functionality bypasses the requirement for the essential pre-60S factor Nsa1.","date":"2013","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/24312670","citation_count":30,"is_preprint":false},{"pmid":"32005977","id":"PMC_32005977","title":"WDR74 modulates melanoma tumorigenesis and metastasis through the RPL5-MDM2-p53 pathway.","date":"2020","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/32005977","citation_count":27,"is_preprint":false},{"pmid":"29107693","id":"PMC_29107693","title":"WDR74 participates in an early cleavage of the pre-rRNA processing pathway in cooperation with the nucleolar AAA-ATPase NVL2.","date":"2017","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/29107693","citation_count":26,"is_preprint":false},{"pmid":"21799883","id":"PMC_21799883","title":"Wdr74 is required for blastocyst formation in the mouse.","date":"2011","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/21799883","citation_count":21,"is_preprint":false},{"pmid":"26456651","id":"PMC_26456651","title":"AAA-ATPase NVL2 acts on MTR4-exosome complex to dissociate the nucleolar protein WDR74.","date":"2015","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/26456651","citation_count":20,"is_preprint":false},{"pmid":"39497511","id":"PMC_39497511","title":"METTL14 attenuates cancer stemness by suppressing ATF5/WDR74/β-catenin axis in gastric cancer.","date":"2024","source":"Cancer science","url":"https://pubmed.ncbi.nlm.nih.gov/39497511","citation_count":18,"is_preprint":false},{"pmid":"34553072","id":"PMC_34553072","title":"WDR74 promotes proliferation and metastasis in colorectal cancer cells through regulating the Wnt/β-catenin signaling pathway.","date":"2021","source":"Open life sciences","url":"https://pubmed.ncbi.nlm.nih.gov/34553072","citation_count":10,"is_preprint":false},{"pmid":"30594465","id":"PMC_30594465","title":"WDR74 functions as a novel coactivator in TGF-β signaling.","date":"2018","source":"Journal of genetics and genomics = Yi chuan xue bao","url":"https://pubmed.ncbi.nlm.nih.gov/30594465","citation_count":10,"is_preprint":false},{"pmid":"39935469","id":"PMC_39935469","title":"Identification of WDR74 and TNFRSF12A as biomarkers for early osteoarthritis using machine learning and immunohistochemistry.","date":"2025","source":"Frontiers in immunology","url":"https://pubmed.ncbi.nlm.nih.gov/39935469","citation_count":5,"is_preprint":false},{"pmid":"39706051","id":"PMC_39706051","title":"Pre-ribosomal WDR74 module coordinates the early and late pre-rRNA processing stages for the NVL2-mediated regulation of 60S ribosome biogenesis.","date":"2024","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/39706051","citation_count":3,"is_preprint":false},{"pmid":"40959275","id":"PMC_40959275","title":"Increased CAPG inhibits ferroptosis to drive tumor proliferation and sorafenib resistance in hepatocellular carcinoma via the WDR74-p53-SLC7A11 pathway.","date":"2025","source":"International journal of biological sciences","url":"https://pubmed.ncbi.nlm.nih.gov/40959275","citation_count":2,"is_preprint":false},{"pmid":"37331183","id":"PMC_37331183","title":"WDR74 serves as a novel therapeutic target by its oncogenic role in hepatocellular carcinoma.","date":"2023","source":"Pathology, research and practice","url":"https://pubmed.ncbi.nlm.nih.gov/37331183","citation_count":2,"is_preprint":false},{"pmid":"40510136","id":"PMC_40510136","title":"METTL3-mediated SNHG1 m6A modification promotes proliferation and migration through transcriptional regulation of WDR74 in osteosarcoma.","date":"2025","source":"Frontiers in oncology","url":"https://pubmed.ncbi.nlm.nih.gov/40510136","citation_count":2,"is_preprint":false},{"pmid":"39840464","id":"PMC_39840464","title":"WDR74-Mediated Ribosome Biogenesis and Proteome Dynamics During Mouse Preimplantation Development.","date":"2025","source":"Genes to cells : devoted to molecular & cellular mechanisms","url":"https://pubmed.ncbi.nlm.nih.gov/39840464","citation_count":1,"is_preprint":false},{"pmid":"29364241","id":"PMC_29364241","title":"Combining X-Ray Crystallography with Small Angle X-Ray Scattering to Model Unstructured Regions of Nsa1 from S. Cerevisiae.","date":"2018","source":"Journal of visualized experiments : JoVE","url":"https://pubmed.ncbi.nlm.nih.gov/29364241","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":11050,"output_tokens":4478,"usd":0.05016,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":12480,"output_tokens":4852,"usd":0.09185,"stage2_stop_reason":"end_turn"},"total_usd":0.14201,"stage1_batch_id":"msgbatch_01UT1AahFPCAyn2uVweS7A9z","stage2_batch_id":"msgbatch_01HuKWtKNT4Pqj1bc8eEP5RV","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2008,\n      \"finding\": \"The AAA-ATPase Rix7 (yeast ortholog of NVL2) is required for the energy-dependent release of Nsa1 (yeast ortholog of WDR74) from a discrete late-nucleolar pre-60S particle; in rix7 mutants, Nsa1 cannot dissociate from pre-60S particles and aberrantly accumulates in the cytoplasm associated with aberrant 60S subunits. Rix7 interacts genetically with Nsa1 and is targeted to the Nsa1-defined preribosomal particle.\",\n      \"method\": \"Genetic epistasis, in vivo localization (fluorescence microscopy), co-immunoprecipitation, yeast mutant analysis\",\n      \"journal\": \"The Journal of cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal genetic and biochemical evidence in yeast ortholog, multiple orthogonal methods (genetics, localization, co-IP), independently foundational study\",\n      \"pmids\": [\"18559667\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Nsa1 (WDR74 ortholog) associates with a late-nucleolar pre-60S particle that also contains the DEAD-box RNA helicase Mak5; mutant alleles of MAK5, NOP1, and NOP4 bypass the essential requirement for Nsa1, placing Nsa1 in a pathway involving these factors. Dominant-negative Rix7 retains its growth defect even in the absence of Nsa1, indicating Rix7 has additional nuclear substrates beyond Nsa1.\",\n      \"method\": \"Genetic epistasis (bypass suppressor screen), co-immunoprecipitation, synthetic lethality screens, yeast mutant analysis\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal genetic methods (bypass suppression, synthetic lethality, dominant-negative analysis) in yeast ortholog, replicated with prior work\",\n      \"pmids\": [\"24312670\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"WDR74 is a component of the MTR4-exosome complex in the nucleolus; the AAA-ATPase NVL2 (human ortholog of Rix7) uses its ATPase activity to dissociate WDR74 from this complex. Knockdown of WDR74 decreases 60S ribosome levels in human cells.\",\n      \"method\": \"Proteomic screen (mass spectrometry), co-immunoprecipitation, ATPase-deficient NVL2 mutant analysis, siRNA knockdown with ribosome fractionation\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP with mass spectrometry, ATPase mutant validation, loss-of-function with defined ribosome phenotype; single lab but multiple orthogonal methods\",\n      \"pmids\": [\"26456651\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"WDR74 is required for early pre-rRNA cleavage within ITS1 in the 60S ribosome biogenesis pathway; knockdown of WDR74 causes significant defects in this cleavage step. ATPase-deficient NVL2 prevents dissociation of WDR74 from the MTR4-exosome complex, causing partial migration of WDR74 from the nucleolus to nucleoplasm and an increased interaction between WDR74 and MTR4 in the nucleoplasm, which also produces the same early ITS1 processing defect.\",\n      \"method\": \"siRNA knockdown with pre-rRNA processing analysis (Northern blot), ATPase-deficient NVL2 mutant, in situ proximity ligation assay, subcellular fractionation/localization\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function with specific molecular phenotype (ITS1 cleavage), mutant validation, proximity ligation assay; single lab, multiple orthogonal methods\",\n      \"pmids\": [\"29107693\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"The full-length structure of yeast Nsa1 (WDR74 ortholog) was determined using a hybrid X-ray crystallography / SAXS approach: the N-terminal WD40 domain was solved by X-ray crystallography, and the disordered C-terminus was modeled by SAXS with rigid body and ab initio modeling, revealing the quaternary structure of the entire protein.\",\n      \"method\": \"X-ray crystallography (WD40 domain), SAXS (full-length solution structure), ab initio and rigid body modeling\",\n      \"journal\": \"Journal of visualized experiments : JoVE\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — experimental structure determination using two complementary structural methods on purified recombinant protein; single study but rigorous hybrid approach\",\n      \"pmids\": [\"29364241\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"WDR74 functions as a transcriptional coactivator for Smad proteins in the canonical TGF-β signaling pathway; WDR74 directly interacts with Smad proteins and enhances TGF-β-mediated phosphorylation and nuclear accumulation of Smad2 and Smad3, leading to stronger transcriptional responses.\",\n      \"method\": \"Co-immunoprecipitation (direct interaction with Smad proteins), Western blot (Smad2/3 phosphorylation), nuclear fractionation, gain- and loss-of-function assays with TGF-β reporter\",\n      \"journal\": \"Journal of genetics and genomics = Yi chuan xue bao\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP for direct interaction, phosphorylation assay, nuclear accumulation measurement; single lab with multiple methods\",\n      \"pmids\": [\"30594465\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"WDR74 promotes nuclear β-catenin accumulation and activates downstream Wnt-responsive genes in lung cancer cells; gain- and loss-of-function studies showed WDR74 regulates cell proliferation, cell cycle, chemoresistance, and aggressiveness via the Wnt/β-catenin signaling pathway.\",\n      \"method\": \"Gain- and loss-of-function (overexpression and knockout), Western blot (β-catenin nuclear accumulation), Wnt reporter assays, xenograft mouse model\",\n      \"journal\": \"Cancer letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — functional KO/OE with defined pathway placement and in vivo validation; single lab, multiple methods\",\n      \"pmids\": [\"31838084\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"WDR74 modulates RPL5 protein levels, which in turn regulates MDM2 activity and protects p53 from MDM2-mediated ubiquitination and degradation; WDR74 thus controls the RPL5-MDM2-p53 pathway to promote melanoma cell proliferation, apoptosis resistance, and metastasis.\",\n      \"method\": \"iTRAQ proteomic screening, gain- and loss-of-function approaches, Western blot (RPL5, MDM2, p53 ubiquitination), in vivo xenograft and metastasis models\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — proteomic identification plus functional validation with multiple molecular readouts; single lab\",\n      \"pmids\": [\"32005977\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Wdr74 is essential for blastocyst formation in mouse preimplantation development; Wdr74 knockdown causes embryos to arrest at the morula stage with activated Trp53-dependent apoptosis and global reduction of RNA polymerase I, II, and III transcripts. Blocking Trp53 function rescues blastocyst formation in Wdr74-deficient embryos, placing Wdr74 upstream of Trp53-dependent apoptosis.\",\n      \"method\": \"RNAi knockdown in mouse embryos, RT-qPCR (RNA Pol I/II/III transcripts), genetic epistasis (Trp53 rescue), embryo phenotypic analysis\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — RNAi loss-of-function with specific molecular phenotype plus genetic epistasis rescue establishing pathway position; multiple orthogonal methods\",\n      \"pmids\": [\"21799883\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"WDR74 functions as part of a pre-ribosomal subcomplex termed the 'WDR74 module', consisting of WDR74, RPF1, MAK16, and RRP1; each component of this module is mutually required for interaction of the others with MTR4, and all components are required for accurate pre-rRNA cleavage during 60S biogenesis. Impaired NVL2-mediated release of WDR74 from the MTR4-exosome complex prevents MTR4 from recruiting PICT1, an MTR4 adaptor required for 3'-end maturation of 5.8S rRNA.\",\n      \"method\": \"Co-immunoprecipitation combined with mass spectrometry, siRNA knockdown, pre-rRNA processing analysis, interaction mapping\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP/MS identification of the subcomplex, combined with loss-of-function showing mutual dependency and specific rRNA processing defects; single lab, multiple orthogonal methods\",\n      \"pmids\": [\"39706051\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"WDR74 interacts with Smad2/3 in macrophages (co-immunoprecipitation) and promotes TGF-β/Smad pathway activation; WDR74 overexpression increases Smad2/3 phosphorylation and promotes M2 macrophage polarization and ECM production in a diabetic foot ulcer mouse model. These effects are reversed by the TGF-β receptor inhibitor LY2109761.\",\n      \"method\": \"Co-immunoprecipitation (WDR74-Smad2/3 interaction), gain- and loss-of-function (overexpression/knockdown), Western blot (Smad2/3 phosphorylation), immunofluorescence, mouse DFU model\",\n      \"journal\": \"Cell biology and toxicology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP confirms direct interaction, phosphorylation assay, pharmacological rescue; single lab, multiple methods\",\n      \"pmids\": [\"35982296\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"WDR74 deficiency in mouse embryos (generated by CRISPR-Cas9) leads to impaired 60S ribosome biogenesis with significant reduction in large ribosomal subunit proteins (notably RPL24 and RPL26) but not small subunit proteins, and blocks cell division progression beyond the morula stage.\",\n      \"method\": \"CRISPR-Cas9 knockout, label-free quantitative proteomics, cell division phenotypic analysis\",\n      \"journal\": \"Genes to cells : devoted to molecular & cellular mechanisms\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — CRISPR KO with quantitative proteomics revealing specific large subunit protein reduction; single lab, rigorous genome editing plus proteomics\",\n      \"pmids\": [\"39840464\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"WDR74 decreases phosphorylation of β-catenin and promotes its nuclear accumulation in colorectal cancer cells, activating the Wnt/β-catenin signaling pathway; blocking this pathway with XAV-939 reverses WDR74-mediated effects on proliferation, migration, and invasion.\",\n      \"method\": \"siRNA knockdown, Western blot (β-catenin phosphorylation and nuclear localization), XAV-939 pharmacological rescue, cell proliferation and invasion assays\",\n      \"journal\": \"Open life sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — loss-of-function with pathway placement confirmed by pharmacological rescue; single lab, corroborates findings from PMID:31838084\",\n      \"pmids\": [\"34553072\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"ATF5 transcriptionally upregulates WDR74, and WDR74 in turn enhances β-catenin nuclear translocation to promote stemness in gastric cancer; METTL14 suppresses this axis by promoting m6A-mediated degradation of ATF5 mRNA. ChIP assays confirmed ATF5 binds the WDR74 promoter.\",\n      \"method\": \"ChIP assay (ATF5 binding to WDR74 promoter), MeRIP-qPCR (m6A modification of ATF5), Western blot (β-catenin nuclear translocation), rescue/overexpression assays\",\n      \"journal\": \"Cancer science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP confirms direct transcriptional regulation, MeRIP validates upstream epigenetic mechanism, functional rescue validates pathway; single lab\",\n      \"pmids\": [\"39497511\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"CAPG promotes WDR74 transcription, and WDR74 in turn modulates the interaction between p53 and MDM2, resulting in p53 ubiquitination and degradation, thereby inhibiting ferroptosis in hepatocellular carcinoma. This was supported by co-immunoprecipitation and ubiquitination assays.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assays, ChIP sequencing, RNA sequencing, gain- and loss-of-function, xenograft model\",\n      \"journal\": \"International journal of biological sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP and ubiquitination assays establish mechanism; single lab, corroborates RPL5-MDM2-p53 axis from PMID:32005977\",\n      \"pmids\": [\"40959275\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"SNHG1 (a lncRNA encoded adjacent to WDR74) promotes WDR74 transcription in cis by recruiting EWSR1 to the WDR74 promoter region; ChIP-qPCR confirmed EWSR1 binding at the WDR74 promoter, establishing a trans-regulatory mechanism upstream of WDR74 expression in osteosarcoma.\",\n      \"method\": \"ChIP-qPCR (EWSR1 binding at WDR74 promoter), RNA pulldown, RNA immunoprecipitation, actinomycin D stability assay\",\n      \"journal\": \"Frontiers in oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP-qPCR confirms direct EWSR1 binding at WDR74 promoter; single lab, multiple supporting methods\",\n      \"pmids\": [\"40510136\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"WDR74 (Nsa1 in yeast) is a nucleolar WD40-domain protein that functions primarily as an essential pre-60S ribosome biogenesis factor: it resides in a pre-ribosomal subcomplex (with RPF1, MAK16, and RRP1) that coordinates early ITS1 pre-rRNA cleavage and associates with the MTR4-nuclear exosome complex, from which it is released by the AAA-ATPase NVL2/Rix7 in an ATP hydrolysis-dependent manner to permit downstream 5.8S rRNA maturation; loss of WDR74 reduces 60S ribosome levels, activates Trp53-dependent apoptosis, and blocks mammalian cell division. Beyond ribosome biogenesis, WDR74 also functions as a coactivator of TGF-β/Smad signaling through direct interaction with Smad2/3, and promotes Wnt/β-catenin signaling by preventing β-catenin phosphorylation and driving its nuclear accumulation; additionally, WDR74 controls the RPL5-MDM2-p53 axis to regulate p53 stability and ferroptosis resistance in cancer contexts.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"WDR74 (yeast Nsa1) is an essential nucleolar WD40-domain protein that acts as a pre-60S ribosome biogenesis factor coordinating early pre-rRNA processing during large ribosomal subunit assembly [#3, #11]. It functions within a discrete pre-ribosomal subcomplex—the 'WDR74 module' comprising WDR74, RPF1, MAK16, and RRP1—in which each component is mutually required for association with the MTR4-nuclear exosome complex and for accurate ITS1 cleavage in the 60S pathway [#9, #3]. WDR74 is released from the MTR4-exosome complex by the AAA-ATPase NVL2 (yeast Rix7) in an ATP-hydrolysis-dependent manner; blocking this release with ATPase-deficient NVL2 retains WDR74 on MTR4, mislocalizes it to the nucleoplasm, and prevents MTR4 from recruiting the adaptor PICT1 required for 3'-end maturation of 5.8S rRNA [#0, #2, #9]. The full-length architecture of the protein comprises an N-terminal WD40 domain and a disordered C-terminus [#4]. Loss of WDR74 reduces 60S subunit levels and large-subunit ribosomal proteins, arrests cells beyond the morula stage, and activates Trp53-dependent apoptosis that can be rescued by blocking Trp53 [#8, #11]. Beyond ribosome biogenesis, WDR74 acts as a coactivator of TGF-β/Smad signaling through direct interaction with Smad2/3, enhancing their phosphorylation and nuclear accumulation [#5, #10]; promotes Wnt/β-catenin signaling by reducing β-catenin phosphorylation and driving its nuclear accumulation [#6, #12]; and controls the RPL5-MDM2-p53 axis to stabilize or destabilize p53, influencing apoptosis and ferroptosis in cancer contexts [#7, #14].\",\n  \"teleology\": [\n    {\n      \"year\": 2008,\n      \"claim\": \"Established the regulated dissociation step at the heart of WDR74 function—that an AAA-ATPase actively releases Nsa1 from a late pre-60S particle—defining Nsa1 as a transiently associated assembly factor rather than a structural ribosome component.\",\n      \"evidence\": \"Genetic epistasis, fluorescence localization, and co-IP in yeast rix7 mutants\",\n      \"pmids\": [\"18559667\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define the molecular signal triggering release\", \"Yeast ortholog; human relevance not yet tested\", \"No structural basis for the Rix7-Nsa1 interaction\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Placed WDR74 upstream of Trp53-dependent apoptosis in development by showing its loss arrests embryos and globally reduces Pol I/II/III transcripts, with p53 blockade rescuing the phenotype.\",\n      \"evidence\": \"RNAi knockdown in mouse preimplantation embryos with Trp53 rescue and RT-qPCR\",\n      \"pmids\": [\"21799883\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not establish the direct molecular link between WDR74 loss and p53 activation\", \"Global transcript reduction could be secondary to general nucleolar stress\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Mapped Nsa1 into a genetic pathway with Mak5, Nop1, and Nop4, refining where in 60S assembly the factor acts and showing Rix7 has substrates beyond Nsa1.\",\n      \"evidence\": \"Bypass suppressor and synthetic lethality screens, co-IP, dominant-negative analysis in yeast\",\n      \"pmids\": [\"24312670\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Genetic interactions do not establish direct physical contacts\", \"Additional Rix7 substrates left unidentified\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Translated the yeast model to human cells, identifying WDR74 as a component of the MTR4-exosome complex released by NVL2 ATPase activity, with knockdown reducing 60S levels.\",\n      \"evidence\": \"Mass-spectrometry proteomics, co-IP, ATPase-deficient NVL2 mutant, siRNA with ribosome fractionation\",\n      \"pmids\": [\"26456651\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not resolve the specific rRNA processing step affected\", \"Stoichiometry within the MTR4-exosome complex unknown\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Pinpointed the molecular defect as failure of early ITS1 pre-rRNA cleavage and linked NVL2-blocked WDR74 release to nucleoplasmic mislocalization and the same processing defect.\",\n      \"evidence\": \"siRNA with Northern blot pre-rRNA analysis, ATPase-deficient NVL2, proximity ligation assay, fractionation\",\n      \"pmids\": [\"29107693\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not identify the nuclease executing ITS1 cleavage\", \"Whether WDR74 acts catalytically or as a scaffold unresolved\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Provided the full-length structural model of Nsa1, defining an N-terminal WD40 domain and a disordered C-terminus that together explain its quaternary organization.\",\n      \"evidence\": \"Hybrid X-ray crystallography (WD40 domain) and SAXS with ab initio/rigid-body modeling on recombinant protein\",\n      \"pmids\": [\"29364241\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No structure of WDR74 within the pre-60S particle or MTR4 complex\", \"Function of the disordered C-terminus not defined\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Revealed a moonlighting role outside ribosome biogenesis, identifying WDR74 as a Smad coactivator that directly binds Smad2/3 and enhances their phosphorylation and nuclear accumulation.\",\n      \"evidence\": \"Co-IP, Western blot for Smad2/3 phosphorylation, nuclear fractionation, TGF-β reporter gain/loss-of-function\",\n      \"pmids\": [\"30594465\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism by which a nucleolar factor enhances Smad phosphorylation unclear\", \"Direct interaction not validated by reciprocal or structural methods\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Connected WDR74 to Wnt/β-catenin signaling, showing it drives nuclear β-catenin accumulation and tumor-promoting phenotypes in lung cancer.\",\n      \"evidence\": \"Overexpression/knockout, Western blot, Wnt reporter assays, xenograft model\",\n      \"pmids\": [\"31838084\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Did not define how WDR74 affects β-catenin stability mechanistically\", \"Direct versus indirect action on the pathway unresolved\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Defined a ribosomal-protein-mediated route to p53 control, showing WDR74 modulates RPL5 to restrain MDM2 and protect p53 from degradation in melanoma.\",\n      \"evidence\": \"iTRAQ proteomics, gain/loss-of-function, ubiquitination Western blots, xenograft and metastasis models\",\n      \"pmids\": [\"32005977\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Directionality of p53 regulation appears context-dependent across cancers\", \"Whether RPL5 modulation reflects general ribosome biogenesis disruption unclear\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Extended the TGF-β/Smad coactivator role to macrophage biology and tissue repair, with pharmacological receptor blockade reversing WDR74-driven effects.\",\n      \"evidence\": \"Co-IP, gain/loss-of-function, Western blot, immunofluorescence, LY2109761 rescue in a mouse DFU model\",\n      \"pmids\": [\"35982296\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Does not distinguish direct coactivation from broader transcriptional effects\", \"Single-lab confirmation of WDR74-Smad2/3 interaction\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Defined the 'WDR74 module' (WDR74-RPF1-MAK16-RRP1) as a mutually dependent unit required for MTR4 association and showed NVL2-mediated release enables MTR4 recruitment of PICT1 for 5.8S rRNA 3'-end maturation.\",\n      \"evidence\": \"Co-IP/MS, siRNA, pre-rRNA processing analysis, interaction mapping\",\n      \"pmids\": [\"39706051\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural organization of the module within pre-60S not resolved\", \"Order of assembly and release events not fully mapped\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Identified upstream transcriptional control of WDR74 by ATF5, itself regulated by METTL14-mediated m6A, linking WDR74 expression to β-catenin-driven stemness in gastric cancer.\",\n      \"evidence\": \"ChIP for ATF5 promoter binding, MeRIP-qPCR, Western blot, rescue assays\",\n      \"pmids\": [\"39497511\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Does not address whether WDR74's ribosomal function contributes to stemness\", \"Single-lab regulatory axis\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Confirmed in a CRISPR-knockout embryo model that WDR74 loss selectively impairs 60S biogenesis, reducing large-subunit proteins (RPL24, RPL26) but not small-subunit proteins, and blocks cell division beyond the morula stage.\",\n      \"evidence\": \"CRISPR-Cas9 knockout, label-free quantitative proteomics, division phenotyping\",\n      \"pmids\": [\"39840464\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Does not establish whether division arrest is purely ribosome-dependent or via p53\", \"Mechanism of selective large-subunit protein loss not defined\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Showed WDR74 modulates the p53-MDM2 interaction to promote p53 ubiquitination and degradation, inhibiting ferroptosis in hepatocellular carcinoma, with CAPG acting as an upstream transcriptional driver.\",\n      \"evidence\": \"Co-IP, ubiquitination assays, ChIP-seq, RNA-seq, gain/loss-of-function, xenograft\",\n      \"pmids\": [\"40959275\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"p53 outcome opposite to the melanoma RPL5-MDM2 model, context dependence unexplained\", \"Direct versus indirect effect on the p53-MDM2 interface not resolved\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Revealed cis/trans transcriptional regulation of WDR74 by the adjacent lncRNA SNHG1, which recruits EWSR1 to the WDR74 promoter in osteosarcoma.\",\n      \"evidence\": \"ChIP-qPCR for EWSR1 binding, RNA pulldown, RIP, actinomycin D stability assay\",\n      \"pmids\": [\"40510136\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Does not link this regulation to WDR74's downstream ribosomal or signaling functions\", \"Single-lab regulatory mechanism\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"It remains unresolved how WDR74's core nucleolar ribosome biogenesis function mechanistically connects to its reported roles in TGF-β/Smad, Wnt/β-catenin, and p53/MDM2 signaling, and whether the cancer-context signaling roles are direct or downstream consequences of perturbed ribosome assembly.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified mechanism linking nucleolar and signaling functions\", \"Opposite p53 directionality across cancer models unexplained\", \"No structure of WDR74 engaged with signaling partners\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140110\", \"supporting_discovery_ids\": [5, 10]},\n      {\"term_id\": \"GO:0060089\", \"supporting_discovery_ids\": [6, 12]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005730\", \"supporting_discovery_ids\": [0, 2, 3]},\n      {\"term_id\": \"GO:0005654\", \"supporting_discovery_ids\": [3]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [2, 3, 9]},\n      {\"term_id\": \"R-HSA-1852241\", \"supporting_discovery_ids\": [2, 11]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [5, 6]}\n    ],\n    \"complexes\": [\n      \"WDR74 module (WDR74-RPF1-MAK16-RRP1)\",\n      \"MTR4-nuclear exosome complex\",\n      \"pre-60S ribosomal particle\"\n    ],\n    \"partners\": [\n      \"MTR4\",\n      \"NVL2\",\n      \"RPF1\",\n      \"MAK16\",\n      \"RRP1\",\n      \"SMAD2\",\n      \"SMAD3\",\n      \"RPL5\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}