{"gene":"EMG1","run_date":"2026-06-09T23:54:43","timeline":{"discoveries":[{"year":2001,"finding":"EMG1 (Emg1) is required for biogenesis of the 40S ribosomal subunit in yeast; depletion causes selectively reduced levels of 20S pre-rRNA and mature 18S rRNA. Nuclear localization of Emg1 depends on physical interaction with the nucleolar protein Nop14.","method":"Genetic depletion, temperature-sensitive allele, rRNA Northern blotting, nuclear localization assay","journal":"Molecular biology of the cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (genetic depletion, ts allele, rRNA analysis, localization assay), replicated in mammalian orthologs in the same study","pmids":["11694595"],"is_preprint":false},{"year":2007,"finding":"Crystal structure of yeast Emg1 at 2 Å in complex with SAM identifies it as a novel member of the SPOUT-class (alpha/beta knot fold) methyltransferase superfamily. A point mutation in a basic patch abolished RNA binding in vitro; mutations disrupting SAM binding caused >100-fold reduction in SAM binding but did not prevent growth or ribosome biogenesis, indicating catalytic activity is not essential for ribosome biogenesis.","method":"X-ray crystallography, in vitro SAM binding assays, RNA binding assay, yeast complementation","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure plus mutagenesis plus functional complementation in a single rigorous study","pmids":["18063569"],"is_preprint":false},{"year":2008,"finding":"Crystal structure of archaeal Nep1 (Methanocaldococcus jannaschii) in free, SAH-bound, and sinefungin-bound forms reveals an extended SPOUT-class methyltransferase fold with a preformed, pre-organized SAM-binding site topologically equivalent to other SPOUT methyltransferases.","method":"X-ray crystallography (2.15–2.25 Å resolution), ligand co-crystallization","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1 / Strong — multiple crystal structures with bound cofactor analogs in a single rigorous structural study","pmids":["18208838"],"is_preprint":false},{"year":2010,"finding":"Nep1 (Emg1) is a pseudouridine-N1-specific methyltransferase: it selectively methylates RNAs containing a pseudouridine at a position corresponding to the hypermodified m1acp3-Ψ in eukaryotic 18S rRNA, transferring the methyl group to the N1 of pseudouridine. This enzymatic activity is conserved in human NEP1.","method":"Fluorescence spectroscopy, NMR spectroscopy, MALDI-MS, HPLC, yeast three-hybrid screening for RNA-binding specificity, in vitro methyltransferase assay","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro enzymatic assay with multiple orthogonal analytical methods (MS, HPLC, NMR) confirming N1-specific pseudouridine methylation","pmids":["20047967"],"is_preprint":false},{"year":2010,"finding":"In yeast, Nep1 catalyzes in vivo methylation of pseudouridine at position 1191 (Ψ1191) within loop 35 of 18S rRNA, the site of the hypermodification m1acp3Ψ. Nep1 is not required for the acp-modification but provides the Ψ1191 methylation. Nep1 has a dual function: as Ψ1191-methyltransferase and as an essential ribosome assembly factor, since restored growth of nep1-1(ts) by SAM addition was observed even when Ψ1191 methylation was prevented. The Bowen-Conradi D86G mutation does not affect methyltransferase activity but enhances dimerization and RNA-target affinity, and prevents nucleolar accumulation of Nep1.","method":"14C-methionine labeling of 18S rRNA in yeast mutants, ESI-MS analysis of acp-modified Ψ-nucleosides in Δnep1 mutants, genetic suppressor analysis, SAM rescue experiments","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1 / Strong — direct in vivo isotopic labeling, mass spectrometry, genetic suppressor experiments in multiple yeast mutant backgrounds, multiple orthogonal methods","pmids":["20972225"],"is_preprint":false},{"year":2009,"finding":"A D86G mutation in human EMG1 causes Bowen-Conradi syndrome. In BCS patient fibroblasts, EMG1 protein is dramatically reduced without change in mRNA levels. Overexpression of EMG1-D86G in mammalian cells decreases soluble EMG1, and yeast two-hybrid analysis shows D86G increases interaction between EMG1 subunits, suggesting the mutation causes aggregation.","method":"Patient sequencing, Western blotting, yeast two-hybrid, mammalian overexpression, structural modeling","journal":"American journal of human genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple methods (Western blot, yeast two-hybrid, mammalian overexpression) in a single lab; structural modeling is computational","pmids":["19463982"],"is_preprint":false},{"year":2010,"finding":"Crystal structures of S. cerevisiae Nep1 dimer and its RNA complexes show Nep1 recognizes its RNA site via base-specific interactions, stabilizes a stem-loop in bound RNA, and the bound RNA structure contradicts predicted rRNA secondary structures, suggesting Nep1 remodels rRNA upon binding. A uridine base is positioned in the active site for methyltransfer at C5, supporting N1-specific pseudouridine methyltransferase activity. Mutations reducing methyl-donor binding are not lethal.","method":"X-ray crystallography of protein-RNA complexes, mutagenesis","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structures of multiple complexes with RNA and SAM analogs, plus mutagenesis in a single rigorous study","pmids":["21087996"],"is_preprint":false},{"year":2010,"finding":"EMG1 knockout in mice arrests embryonic development prior to the blastocyst stage with defects in early cell lineage-specification and nucleologenesis. Loss of p53 failed to rescue Emg1-/- pre-implantation lethality, unlike some other ribosome biogenesis defects.","method":"Mouse gene targeting (knockout), embryo phenotyping, genetic epistasis with p53 null","journal":"BMC developmental biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — clean knockout with defined developmental phenotype, genetic epistasis with p53 tested, replicates essential role seen in yeast","pmids":["20858271"],"is_preprint":false},{"year":2012,"finding":"Genetic interaction screen in yeast identified that NEP1 (EMG1) interacts genetically with ribosome biogenesis genes RPS18A/B, RRP8, EFG1, UTP30, ribosome quality control genes UBP3, BRE5, UBP6, and no-go decay factor DOM34. UTP30 deletion enforces all nep1-1(ts) phenotypes; UTP30 overexpression partially restores nep1-1(ts) growth; genetic and biochemical data suggest Utp30 and Nep1 act together during pre-ribosomal complex formation and provide the surface for Rps19 assembly to the 90S pre-ribosome.","method":"Genome-wide synthetic genetic array (SGA), biochemical co-analysis, yeast complementation","journal":"Yeast (Chichester, England)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genome-wide genetic screen with specific follow-up, single lab","pmids":["22588997"],"is_preprint":false},{"year":2007,"finding":"Tma23 and Nop6 mutations act as recessive suppressors of nep1(ts) and nep1 deletion in yeast. GFP fusions localized both proteins to the nucleolus, supporting their role in ribosome biogenesis. Both proteins have high lysine content; Nop6 contains an RNA-binding motif, suggesting RNA-binding functions.","method":"Suppressor screen, GFP localization, genetic complementation","journal":"FEMS yeast research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic suppressor identification with localization confirmation, single lab","pmids":["17425675"],"is_preprint":false},{"year":2016,"finding":"EMG1 is imported into the nucleus by importins Impα/β or Impβ/7. Binding of the Impβ/7 heterodimer prevents unspecific aggregation of both EMG1 and EMG1-D86G on RNAs in vitro, indicating importins act as chaperones for EMG1 by binding to its basic regions. The BCS D86G mutation leads to reduced nucleolar localization, accumulation of EMG1-D86G in nuclear foci, and proteasome-dependent degradation. Pre-ribosomal factors NOP14, NOC4L, and UTP14A form a nucleolar subcomplex containing EMG1 required for its nucleolar recruitment.","method":"Nuclear import assays, Co-immunoprecipitation, in vitro aggregation assay, inhibitor treatments (proteasome inhibitor), fluorescence microscopy","journal":"Human molecular genetics","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (import assay, CoIP, in vitro aggregation, localization microscopy, pharmacological inhibition) in a single study","pmids":["27798105"],"is_preprint":false},{"year":2014,"finding":"In BCS patient lymphoblasts (D86G mutation), cells accumulate in G2/M resulting in reduced proliferation, and 18S rRNA processing is consistently delayed, though levels of 40S ribosomes and protein synthesis rates are not different from controls. The D86G substitution does not interfere with EMG1 nucleolar localization in mammalian cells.","method":"Cell cycle analysis (flow cytometry), proliferation assays, pulse-chase rRNA processing analysis, Western blotting, immunofluorescence","journal":"BBA clinical","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple methods in patient cells, single lab, functional ribosome biogenesis readout","pmids":["26676230"],"is_preprint":false},{"year":2020,"finding":"EMG1 interacts with NOP14 as shown by GST pulldown and co-immunoprecipitation. Together, EMG1 and NOP14 regulate cell growth, apoptosis, migration and invasion in melanoma cells, and simultaneous overexpression decreases WNT3a, β-catenin, phospho-GSK-3β, and c-Myc levels, implicating the Wnt/β-catenin pathway.","method":"GST pulldown, co-immunoprecipitation, overexpression, Western blotting","journal":"Translational cancer research","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — reciprocal pulldown plus CoIP confirms interaction; pathway placement by protein level changes, single lab","pmids":["35117729"],"is_preprint":false},{"year":2025,"finding":"EMG1 forms a protein complex with the transcription factor GRHL3 and is required for correct nuclear localization of GRHL3 and activation of the canonical Wnt/β-catenin signaling pathway. Conditional knockout of Emg1 in the GRHL3-positive surface ectoderm causes spina bifida (neural tube defects). Compound mutant phenotypes of Emg1 and Grhl3 indicate genetic interaction in neurulation and palate development.","method":"Conditional knockout mouse model, protein complex identification, immunofluorescence, genetic epistasis analysis","journal":"Development (Cambridge, England)","confidence":"High","confidence_rationale":"Tier 2 / Strong — conditional knockout with specific developmental phenotype, protein complex identification, genetic epistasis, multiple orthogonal methods","pmids":["40761126"],"is_preprint":false},{"year":2025,"finding":"EMG1 methyltransferase activity is required for KSHV-induced specialized ribosomes to scan through KSHV upstream open reading frames (uORFs) in late lytic genes. During KSHV lytic replication, EMG1 has enhanced association with precursor-40S ribosome complexes. Depletion of EMG1 reduces viral protein expression and infectious virus production.","method":"EMG1 depletion (knockdown), ribosome profiling/occupancy assays, viral replication assays, precursor-40S complex immunoprecipitation","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (depletion, ribosome occupancy, viral output), specific methyltransferase-activity requirement demonstrated, peer-reviewed","pmids":["40215162"],"is_preprint":false},{"year":2023,"finding":"Molecular dynamics simulations and quantum-chemical calculations, combined with kinetic and mutational experiments, elucidate the catalytic mechanism of Nep1: a conserved aspartate (D101 in S. cerevisiae) is essential for catalysis, acting as a base for deprotonation of pseudouridine N1. A conserved hydroxyl-containing residue (S233 in S. cerevisiae) and active-site water molecules can mediate proton transfer. The water-mediated proton transfer pathway is the most energetically favorable. Mutation of D101 abolishes activity; mutation of S233 does not.","method":"Molecular dynamics simulation, DFT quantum-chemical calculations, site-directed mutagenesis, kinetic assays","journal":"Computational and structural biotechnology journal","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — in vitro mutagenesis and kinetics plus computational modeling; single lab, novel mechanistic detail not independently replicated","pmids":["37649713"],"is_preprint":false},{"year":2025,"finding":"Crystal structures of Pyrococcus horikoshii Nep1 (PhNep1) in apo, adenosine-bound, and 5-methylthioadenosine-bound forms reveal an α/β SPOUT fold with a trefoil knot, two novel extensions (a globular loop and a β-α-β extension), a preformed cofactor-binding pocket, and a homodimer stabilized by inter-subunit hydrogen bonds and hydrophobic interactions. PhNep1 specifically methylates a pseudouridine at position 926 of helix 35 in 16S rRNA; conserved arginine residues at the dimeric interface assist RNA complex stability.","method":"X-ray crystallography (apo and holo forms), in vitro methyltransferase assay with consensus RNA substrates","journal":"The FEBS journal","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structures of multiple forms plus in vitro enzymatic activity confirmed with specific RNA substrate in single rigorous study","pmids":["39918246"],"is_preprint":false},{"year":2025,"finding":"FTO-mediated m6A demethylation of EMG1 mRNA decreases EMG1 protein expression and reduces ribosome biosynthesis, suppressing bladder cancer cell proliferation, migration, and invasion in vitro and in vivo.","method":"FTO overexpression/knockdown, m6A modification analysis, in vitro and in vivo tumor models, Western blotting","journal":"Biochimica et biophysica acta. Molecular cell research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — gain- and loss-of-function with mechanistic link to m6A modification of EMG1 mRNA; single lab, multiple methods","pmids":["41962773"],"is_preprint":false}],"current_model":"EMG1 (Nep1) is an essential nucleolar SPOUT-class pseudouridine-N1-methyltransferase that methylates a specific pseudouridine in the small ribosomal subunit rRNA (Ψ1191 in yeast 18S rRNA), functions as a ribosome assembly factor for 40S subunit biogenesis (with a separable essential role beyond its catalytic activity), forms a nucleolar subcomplex with NOP14/NOC4L/UTP14A required for its nucleolar recruitment, is imported into the nucleus via importins Impα/β or Impβ/7 (which also chaperone it against RNA aggregation), and in development cooperates with the transcription factor GRHL3 in β-catenin-mediated surface ectoderm differentiation; its methyltransferase activity is additionally required for KSHV-induced specialized ribosome function, and the disease-causing D86G (Bowen-Conradi syndrome) mutation enhances dimerization, promotes protein aggregation and proteasomal degradation, and impairs nucleolar accumulation without abolishing catalytic activity."},"narrative":{"mechanistic_narrative":"EMG1 (Nep1) is an essential nucleolar SPOUT-class methyltransferase that functions in biogenesis of the small (40S) ribosomal subunit, where its depletion selectively reduces 20S pre-rRNA and mature 18S rRNA [PMID:11694595, PMID:18063569]. Enzymatically it is a pseudouridine-N1-specific methyltransferase that transfers a methyl group to the N1 of pseudouridine, generating the m1acp3-Ψ hypermodification at Ψ1191 of 18S rRNA in yeast (Ψ926 in archaeal 16S rRNA), with catalysis depending on a conserved active-site aspartate acting as a base [PMID:20047967, PMID:20972225, PMID:39918246, PMID:37649713]. Structures show EMG1 adopts an alpha/beta knot fold with a preformed SAM-binding pocket, acts as a homodimer, and recognizes its target via base-specific contacts that remodel the bound rRNA stem-loop [PMID:18063569, PMID:18208838, PMID:21087996]. EMG1 has a catalytically separable, essential role in ribosome assembly, since mutations that abolish SAM binding or block Ψ1191 methylation do not prevent ribosome biogenesis or growth [PMID:18063569, PMID:20972225]. It is imported into the nucleus by Impα/β or Impβ/7, which also chaperone its basic regions against nonspecific RNA aggregation, and is recruited to the nucleolus within a subcomplex containing NOP14, NOC4L, and UTP14A [PMID:27798105]. Beyond ribosome assembly, EMG1 is required for early embryonic development and nucleologenesis [PMID:20858271], cooperates with the transcription factor GRHL3 to drive nuclear GRHL3 localization and Wnt/β-catenin–mediated surface ectoderm differentiation [PMID:40761126], and its methyltransferase activity supports KSHV-induced specialized ribosome scanning of viral uORFs [PMID:40215162]. The D86G missense mutation causes Bowen-Conradi syndrome; it does not abolish catalytic activity but enhances dimerization and RNA affinity, promotes aggregation and proteasomal degradation, and impairs nucleolar accumulation [PMID:19463982, PMID:20972225, PMID:27798105].","teleology":[{"year":2001,"claim":"Established EMG1 as a 40S ribosomal subunit biogenesis factor whose nuclear function depends on a defined protein partner, framing it as a nucleolar assembly component rather than a free-standing enzyme.","evidence":"Genetic depletion, ts allele and rRNA Northern blotting in yeast with Nop14 interaction and localization assays","pmids":["11694595"],"confidence":"High","gaps":["Molecular activity unknown at this stage","Did not define which pre-rRNA processing step requires EMG1"]},{"year":2007,"claim":"Defined EMG1 as a SPOUT-class methyltransferase by structure and, critically, showed that catalytic activity is dispensable for ribosome biogenesis, revealing a separable non-catalytic essential role.","evidence":"X-ray crystallography of yeast Emg1 with SAM, SAM- and RNA-binding mutagenesis, yeast complementation; archaeal Nep1 structures with cofactor analogs","pmids":["18063569","18208838"],"confidence":"High","gaps":["RNA substrate and methylated position not yet identified","Nature of the catalysis-independent essential function unresolved"]},{"year":2010,"claim":"Identified the precise enzymatic activity and target: N1-methylation of pseudouridine at Ψ1191 of 18S rRNA, and confirmed a dual catalytic / assembly-factor function.","evidence":"In vitro methyltransferase assays with MS/HPLC/NMR, in vivo 14C-methionine labeling, ESI-MS, and genetic suppressor/SAM rescue in yeast; protein-RNA co-crystal structures","pmids":["20047967","20972225","21087996"],"confidence":"High","gaps":["Mechanism by which assembly function is separable from methylation not defined","rRNA remodeling role inferred from static structures"]},{"year":2010,"claim":"Demonstrated organismal essentiality, showing EMG1 loss arrests early mouse development with nucleologenesis defects in a p53-independent manner.","evidence":"Mouse knockout, embryo phenotyping, genetic epistasis with p53 null","pmids":["20858271"],"confidence":"High","gaps":["Did not resolve whether lethality reflects catalytic or assembly role","Cell-type-specific requirements not addressed"]},{"year":2009,"claim":"Linked EMG1 to human disease by identifying D86G as the cause of Bowen-Conradi syndrome and indicating a protein-stability/aggregation defect.","evidence":"Patient sequencing, Western blotting, yeast two-hybrid, mammalian overexpression, structural modeling","pmids":["19463982"],"confidence":"Medium","gaps":["Aggregation inferred indirectly from solubility and two-hybrid data","Structural modeling is computational, not experimental"]},{"year":2012,"claim":"Placed EMG1 within a genetic network for early pre-ribosome assembly, linking it functionally to Utp30 and the surface for Rps19 loading onto the 90S pre-ribosome.","evidence":"Genome-wide synthetic genetic array with biochemical follow-up and complementation in yeast","pmids":["22588997"],"confidence":"Medium","gaps":["Genetic interactions do not establish direct physical contacts","Single-lab screen without orthogonal validation of all hits"]},{"year":2016,"claim":"Resolved how EMG1 reaches and is protected within the nucleolus, defining importin-mediated import/chaperoning and a NOP14/NOC4L/UTP14A subcomplex required for nucleolar recruitment, and recast D86G pathology as a trafficking/degradation defect.","evidence":"Nuclear import and in vitro aggregation assays, Co-IP, proteasome inhibition, fluorescence microscopy","pmids":["27798105"],"confidence":"High","gaps":["Stoichiometry and assembly order of the subcomplex not defined","Conflicts with patient-cell data on D86G nucleolar localization"]},{"year":2014,"claim":"Characterized cellular consequences of D86G in patient cells, showing delayed 18S processing and G2/M accumulation despite normal steady-state 40S and translation, indicating a partial/kinetic defect.","evidence":"Flow cytometry, proliferation and pulse-chase rRNA processing assays, immunofluorescence in BCS lymphoblasts","pmids":["26676230"],"confidence":"Medium","gaps":["Reports normal D86G nucleolar localization, conflicting with overexpression studies","Mechanism connecting processing delay to G2/M arrest unresolved"]},{"year":2023,"claim":"Defined the chemical catalytic mechanism, identifying a conserved aspartate as the catalytic base for pseudouridine N1 deprotonation and a water-mediated proton-transfer pathway.","evidence":"Molecular dynamics, DFT calculations, site-directed mutagenesis and kinetic assays","pmids":["37649713"],"confidence":"Medium","gaps":["Mechanism is largely computational and not independently replicated","Catalytic detail does not address the non-catalytic assembly role"]},{"year":2025,"claim":"Extended EMG1 function beyond ribosome assembly into development and viral biology, showing a GRHL3 partnership driving Wnt/β-catenin–dependent ectoderm/neural tube morphogenesis and a methyltransferase-dependent role in KSHV specialized ribosomes; also added a fresh archaeal structure with defined RNA target.","evidence":"Conditional knockout mice with protein complex identification and epistasis (GRHL3); EMG1 depletion with ribosome occupancy and viral output assays (KSHV); P. horikoshii crystal structures with in vitro methyltransferase assays","pmids":["40761126","40215162","39918246"],"confidence":"High","gaps":["How a ribosome assembly factor controls GRHL3 nuclear localization mechanistically is unclear","Whether developmental roles require catalysis vs assembly function not separated","Generality of specialized-ribosome role beyond KSHV unknown"]},{"year":2025,"claim":"Tied EMG1 abundance to a cancer-relevant regulatory axis, showing FTO-mediated m6A demethylation of EMG1 mRNA reduces EMG1, ribosome biosynthesis, and bladder tumor cell aggressiveness.","evidence":"FTO gain/loss-of-function, m6A analysis, in vitro and in vivo bladder tumor models","pmids":["41962773"],"confidence":"Medium","gaps":["Direct m6A site on EMG1 mRNA and reader involvement not fully defined","Causality between EMG1 level and tumor phenotype is correlative in part"]},{"year":null,"claim":"The molecular basis of EMG1's catalysis-independent essential ribosome assembly function, and how this is mechanistically distinct from its non-ribosomal roles in GRHL3-mediated development and specialized translation, remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No structure of EMG1 within an intact pre-40S particle","No separation-of-function alleles distinguishing assembly vs developmental roles","Mechanism linking nucleolar assembly factor to nuclear transcription-factor function unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140098","term_label":"catalytic activity, acting on RNA","supporting_discovery_ids":[3,4,6,16]},{"term_id":"GO:0016740","term_label":"transferase activity","supporting_discovery_ids":[1,3,15]},{"term_id":"GO:0003723","term_label":"RNA binding","supporting_discovery_ids":[1,6,16]}],"localization":[{"term_id":"GO:0005730","term_label":"nucleolus","supporting_discovery_ids":[0,9,10]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[0,10]}],"pathway":[{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[0,4]},{"term_id":"R-HSA-1266738","term_label":"Developmental Biology","supporting_discovery_ids":[7,13]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[12,13]}],"complexes":["EMG1/NOP14/NOC4L/UTP14A nucleolar subcomplex","EMG1-GRHL3 complex"],"partners":["NOP14","NOC4L","UTP14A","GRHL3","UTP30","RPS19"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q92979","full_name":"Ribosomal RNA small subunit methyltransferase NEP1","aliases":["18S rRNA (pseudouridine(1248)-N1)-methyltransferase","18S rRNA Psi1248 methyltransferase","Nucleolar protein EMG1 homolog","Protein C2f","Ribosome biogenesis protein NEP1"],"length_aa":244,"mass_kda":26.7,"function":"S-adenosyl-L-methionine-dependent pseudouridine N(1)-methyltransferase that methylates pseudouridine at position 1248 (Psi1248) in 18S rRNA. Involved the biosynthesis of the hypermodified N1-methyl-N3-(3-amino-3-carboxypropyl) pseudouridine (m1acp3-Psi) conserved in eukaryotic 18S rRNA. Is not able to methylate uridine at this position (PubMed:20047967). Also has an essential role in 40S ribosomal subunit biogenesis independent on its methyltransferase activity, facilitating the incorporation of ribosomal protein S19 during the formation of pre-ribosomes (By similarity). Part of the small subunit (SSU) processome, first precursor of the small eukaryotic ribosomal subunit. During the assembly of the SSU processome in the nucleolus, many ribosome biogenesis factors, an RNA chaperone and ribosomal proteins associate with the nascent pre-rRNA and work in concert to generate RNA folding, modifications, rearrangements and cleavage as well as targeted degradation of pre-ribosomal RNA by the RNA exosome (PubMed:34516797)","subcellular_location":"Nucleus, nucleolus","url":"https://www.uniprot.org/uniprotkb/Q92979/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/EMG1","classification":"Not Classified","n_dependent_lines":36,"n_total_lines":74,"dependency_fraction":0.4864864864864865},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000126749","cell_line_id":"CID001092","localizations":[{"compartment":"nucleolus_gc","grade":3},{"compartment":"nucleoplasm","grade":1}],"interactors":[{"gene":"NOP14","stoichiometry":4.0},{"gene":"NOC4L","stoichiometry":4.0},{"gene":"IPO5","stoichiometry":0.2},{"gene":"IPO4","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/target/CID001092","total_profiled":1310},"omim":[{"mim_id":"611531","title":"EMG1 N1-SPECIFIC PSEUDOURIDINE METHYLTRANSFERASE; EMG1","url":"https://www.omim.org/entry/611531"},{"mim_id":"611526","title":"NOP14 NUCLEOLAR PROTEIN; NOP14","url":"https://www.omim.org/entry/611526"},{"mim_id":"611159","title":"KERATIN 78, TYPE II; KRT78","url":"https://www.omim.org/entry/611159"},{"mim_id":"211180","title":"BOWEN-CONRADI SYNDROME; BWCNS","url":"https://www.omim.org/entry/211180"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Nucleoli rim","reliability":"Supported"},{"location":"Nucleoplasm","reliability":"Additional"},{"location":"Mitotic chromosome","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/EMG1"},"hgnc":{"alias_symbol":["C2F","NEP1","Grcc2f"],"prev_symbol":[]},"alphafold":{"accession":"Q92979","domains":[{"cath_id":"3.40.1280.10","chopping":"43-53_85-129_150-242","consensus_level":"high","plddt":96.3872,"start":43,"end":242}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q92979","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q92979-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q92979-F1-predicted_aligned_error_v6.png","plddt_mean":89.0},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=EMG1","jax_strain_url":"https://www.jax.org/strain/search?query=EMG1"},"sequence":{"accession":"Q92979","fasta_url":"https://rest.uniprot.org/uniprotkb/Q92979.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q92979/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q92979"}},"corpus_meta":[{"pmid":"17194768","id":"PMC_17194768","title":"Phytotoxicity and innate immune responses induced by Nep1-like proteins.","date":"2006","source":"The Plant cell","url":"https://pubmed.ncbi.nlm.nih.gov/17194768","citation_count":232,"is_preprint":false},{"pmid":"16430931","id":"PMC_16430931","title":"Nep1-like proteins from plant pathogens: recruitment and diversification of the NPP1 domain across taxa.","date":"2006","source":"Phytochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/16430931","citation_count":208,"is_preprint":false},{"pmid":"23051172","id":"PMC_23051172","title":"Evidence for functional diversification within a fungal NEP1-like protein family.","date":"2013","source":"Molecular plant-microbe interactions : MPMI","url":"https://pubmed.ncbi.nlm.nih.gov/23051172","citation_count":125,"is_preprint":false},{"pmid":"20972225","id":"PMC_20972225","title":"The Bowen-Conradi syndrome protein Nep1 (Emg1) has a dual role in eukaryotic ribosome biogenesis, as an essential assembly factor and in the methylation of Ψ1191 in yeast 18S 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Nuclear localization of Emg1 depends on physical interaction with the nucleolar protein Nop14.\",\n      \"method\": \"Genetic depletion, temperature-sensitive allele, rRNA Northern blotting, nuclear localization assay\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (genetic depletion, ts allele, rRNA analysis, localization assay), replicated in mammalian orthologs in the same study\",\n      \"pmids\": [\"11694595\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Crystal structure of yeast Emg1 at 2 Å in complex with SAM identifies it as a novel member of the SPOUT-class (alpha/beta knot fold) methyltransferase superfamily. A point mutation in a basic patch abolished RNA binding in vitro; mutations disrupting SAM binding caused >100-fold reduction in SAM binding but did not prevent growth or ribosome biogenesis, indicating catalytic activity is not essential for ribosome biogenesis.\",\n      \"method\": \"X-ray crystallography, in vitro SAM binding assays, RNA binding assay, yeast complementation\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure plus mutagenesis plus functional complementation in a single rigorous study\",\n      \"pmids\": [\"18063569\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Crystal structure of archaeal Nep1 (Methanocaldococcus jannaschii) in free, SAH-bound, and sinefungin-bound forms reveals an extended SPOUT-class methyltransferase fold with a preformed, pre-organized SAM-binding site topologically equivalent to other SPOUT methyltransferases.\",\n      \"method\": \"X-ray crystallography (2.15–2.25 Å resolution), ligand co-crystallization\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — multiple crystal structures with bound cofactor analogs in a single rigorous structural study\",\n      \"pmids\": [\"18208838\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Nep1 (Emg1) is a pseudouridine-N1-specific methyltransferase: it selectively methylates RNAs containing a pseudouridine at a position corresponding to the hypermodified m1acp3-Ψ in eukaryotic 18S rRNA, transferring the methyl group to the N1 of pseudouridine. This enzymatic activity is conserved in human NEP1.\",\n      \"method\": \"Fluorescence spectroscopy, NMR spectroscopy, MALDI-MS, HPLC, yeast three-hybrid screening for RNA-binding specificity, in vitro methyltransferase assay\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro enzymatic assay with multiple orthogonal analytical methods (MS, HPLC, NMR) confirming N1-specific pseudouridine methylation\",\n      \"pmids\": [\"20047967\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"In yeast, Nep1 catalyzes in vivo methylation of pseudouridine at position 1191 (Ψ1191) within loop 35 of 18S rRNA, the site of the hypermodification m1acp3Ψ. Nep1 is not required for the acp-modification but provides the Ψ1191 methylation. Nep1 has a dual function: as Ψ1191-methyltransferase and as an essential ribosome assembly factor, since restored growth of nep1-1(ts) by SAM addition was observed even when Ψ1191 methylation was prevented. The Bowen-Conradi D86G mutation does not affect methyltransferase activity but enhances dimerization and RNA-target affinity, and prevents nucleolar accumulation of Nep1.\",\n      \"method\": \"14C-methionine labeling of 18S rRNA in yeast mutants, ESI-MS analysis of acp-modified Ψ-nucleosides in Δnep1 mutants, genetic suppressor analysis, SAM rescue experiments\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — direct in vivo isotopic labeling, mass spectrometry, genetic suppressor experiments in multiple yeast mutant backgrounds, multiple orthogonal methods\",\n      \"pmids\": [\"20972225\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"A D86G mutation in human EMG1 causes Bowen-Conradi syndrome. In BCS patient fibroblasts, EMG1 protein is dramatically reduced without change in mRNA levels. Overexpression of EMG1-D86G in mammalian cells decreases soluble EMG1, and yeast two-hybrid analysis shows D86G increases interaction between EMG1 subunits, suggesting the mutation causes aggregation.\",\n      \"method\": \"Patient sequencing, Western blotting, yeast two-hybrid, mammalian overexpression, structural modeling\",\n      \"journal\": \"American journal of human genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple methods (Western blot, yeast two-hybrid, mammalian overexpression) in a single lab; structural modeling is computational\",\n      \"pmids\": [\"19463982\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Crystal structures of S. cerevisiae Nep1 dimer and its RNA complexes show Nep1 recognizes its RNA site via base-specific interactions, stabilizes a stem-loop in bound RNA, and the bound RNA structure contradicts predicted rRNA secondary structures, suggesting Nep1 remodels rRNA upon binding. A uridine base is positioned in the active site for methyltransfer at C5, supporting N1-specific pseudouridine methyltransferase activity. Mutations reducing methyl-donor binding are not lethal.\",\n      \"method\": \"X-ray crystallography of protein-RNA complexes, mutagenesis\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structures of multiple complexes with RNA and SAM analogs, plus mutagenesis in a single rigorous study\",\n      \"pmids\": [\"21087996\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"EMG1 knockout in mice arrests embryonic development prior to the blastocyst stage with defects in early cell lineage-specification and nucleologenesis. Loss of p53 failed to rescue Emg1-/- pre-implantation lethality, unlike some other ribosome biogenesis defects.\",\n      \"method\": \"Mouse gene targeting (knockout), embryo phenotyping, genetic epistasis with p53 null\",\n      \"journal\": \"BMC developmental biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — clean knockout with defined developmental phenotype, genetic epistasis with p53 tested, replicates essential role seen in yeast\",\n      \"pmids\": [\"20858271\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Genetic interaction screen in yeast identified that NEP1 (EMG1) interacts genetically with ribosome biogenesis genes RPS18A/B, RRP8, EFG1, UTP30, ribosome quality control genes UBP3, BRE5, UBP6, and no-go decay factor DOM34. UTP30 deletion enforces all nep1-1(ts) phenotypes; UTP30 overexpression partially restores nep1-1(ts) growth; genetic and biochemical data suggest Utp30 and Nep1 act together during pre-ribosomal complex formation and provide the surface for Rps19 assembly to the 90S pre-ribosome.\",\n      \"method\": \"Genome-wide synthetic genetic array (SGA), biochemical co-analysis, yeast complementation\",\n      \"journal\": \"Yeast (Chichester, England)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genome-wide genetic screen with specific follow-up, single lab\",\n      \"pmids\": [\"22588997\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Tma23 and Nop6 mutations act as recessive suppressors of nep1(ts) and nep1 deletion in yeast. GFP fusions localized both proteins to the nucleolus, supporting their role in ribosome biogenesis. Both proteins have high lysine content; Nop6 contains an RNA-binding motif, suggesting RNA-binding functions.\",\n      \"method\": \"Suppressor screen, GFP localization, genetic complementation\",\n      \"journal\": \"FEMS yeast research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic suppressor identification with localization confirmation, single lab\",\n      \"pmids\": [\"17425675\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"EMG1 is imported into the nucleus by importins Impα/β or Impβ/7. Binding of the Impβ/7 heterodimer prevents unspecific aggregation of both EMG1 and EMG1-D86G on RNAs in vitro, indicating importins act as chaperones for EMG1 by binding to its basic regions. The BCS D86G mutation leads to reduced nucleolar localization, accumulation of EMG1-D86G in nuclear foci, and proteasome-dependent degradation. Pre-ribosomal factors NOP14, NOC4L, and UTP14A form a nucleolar subcomplex containing EMG1 required for its nucleolar recruitment.\",\n      \"method\": \"Nuclear import assays, Co-immunoprecipitation, in vitro aggregation assay, inhibitor treatments (proteasome inhibitor), fluorescence microscopy\",\n      \"journal\": \"Human molecular genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (import assay, CoIP, in vitro aggregation, localization microscopy, pharmacological inhibition) in a single study\",\n      \"pmids\": [\"27798105\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"In BCS patient lymphoblasts (D86G mutation), cells accumulate in G2/M resulting in reduced proliferation, and 18S rRNA processing is consistently delayed, though levels of 40S ribosomes and protein synthesis rates are not different from controls. The D86G substitution does not interfere with EMG1 nucleolar localization in mammalian cells.\",\n      \"method\": \"Cell cycle analysis (flow cytometry), proliferation assays, pulse-chase rRNA processing analysis, Western blotting, immunofluorescence\",\n      \"journal\": \"BBA clinical\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple methods in patient cells, single lab, functional ribosome biogenesis readout\",\n      \"pmids\": [\"26676230\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"EMG1 interacts with NOP14 as shown by GST pulldown and co-immunoprecipitation. Together, EMG1 and NOP14 regulate cell growth, apoptosis, migration and invasion in melanoma cells, and simultaneous overexpression decreases WNT3a, β-catenin, phospho-GSK-3β, and c-Myc levels, implicating the Wnt/β-catenin pathway.\",\n      \"method\": \"GST pulldown, co-immunoprecipitation, overexpression, Western blotting\",\n      \"journal\": \"Translational cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — reciprocal pulldown plus CoIP confirms interaction; pathway placement by protein level changes, single lab\",\n      \"pmids\": [\"35117729\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"EMG1 forms a protein complex with the transcription factor GRHL3 and is required for correct nuclear localization of GRHL3 and activation of the canonical Wnt/β-catenin signaling pathway. Conditional knockout of Emg1 in the GRHL3-positive surface ectoderm causes spina bifida (neural tube defects). Compound mutant phenotypes of Emg1 and Grhl3 indicate genetic interaction in neurulation and palate development.\",\n      \"method\": \"Conditional knockout mouse model, protein complex identification, immunofluorescence, genetic epistasis analysis\",\n      \"journal\": \"Development (Cambridge, England)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — conditional knockout with specific developmental phenotype, protein complex identification, genetic epistasis, multiple orthogonal methods\",\n      \"pmids\": [\"40761126\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"EMG1 methyltransferase activity is required for KSHV-induced specialized ribosomes to scan through KSHV upstream open reading frames (uORFs) in late lytic genes. During KSHV lytic replication, EMG1 has enhanced association with precursor-40S ribosome complexes. Depletion of EMG1 reduces viral protein expression and infectious virus production.\",\n      \"method\": \"EMG1 depletion (knockdown), ribosome profiling/occupancy assays, viral replication assays, precursor-40S complex immunoprecipitation\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (depletion, ribosome occupancy, viral output), specific methyltransferase-activity requirement demonstrated, peer-reviewed\",\n      \"pmids\": [\"40215162\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Molecular dynamics simulations and quantum-chemical calculations, combined with kinetic and mutational experiments, elucidate the catalytic mechanism of Nep1: a conserved aspartate (D101 in S. cerevisiae) is essential for catalysis, acting as a base for deprotonation of pseudouridine N1. A conserved hydroxyl-containing residue (S233 in S. cerevisiae) and active-site water molecules can mediate proton transfer. The water-mediated proton transfer pathway is the most energetically favorable. Mutation of D101 abolishes activity; mutation of S233 does not.\",\n      \"method\": \"Molecular dynamics simulation, DFT quantum-chemical calculations, site-directed mutagenesis, kinetic assays\",\n      \"journal\": \"Computational and structural biotechnology journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro mutagenesis and kinetics plus computational modeling; single lab, novel mechanistic detail not independently replicated\",\n      \"pmids\": [\"37649713\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Crystal structures of Pyrococcus horikoshii Nep1 (PhNep1) in apo, adenosine-bound, and 5-methylthioadenosine-bound forms reveal an α/β SPOUT fold with a trefoil knot, two novel extensions (a globular loop and a β-α-β extension), a preformed cofactor-binding pocket, and a homodimer stabilized by inter-subunit hydrogen bonds and hydrophobic interactions. PhNep1 specifically methylates a pseudouridine at position 926 of helix 35 in 16S rRNA; conserved arginine residues at the dimeric interface assist RNA complex stability.\",\n      \"method\": \"X-ray crystallography (apo and holo forms), in vitro methyltransferase assay with consensus RNA substrates\",\n      \"journal\": \"The FEBS journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structures of multiple forms plus in vitro enzymatic activity confirmed with specific RNA substrate in single rigorous study\",\n      \"pmids\": [\"39918246\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"FTO-mediated m6A demethylation of EMG1 mRNA decreases EMG1 protein expression and reduces ribosome biosynthesis, suppressing bladder cancer cell proliferation, migration, and invasion in vitro and in vivo.\",\n      \"method\": \"FTO overexpression/knockdown, m6A modification analysis, in vitro and in vivo tumor models, Western blotting\",\n      \"journal\": \"Biochimica et biophysica acta. Molecular cell research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — gain- and loss-of-function with mechanistic link to m6A modification of EMG1 mRNA; single lab, multiple methods\",\n      \"pmids\": [\"41962773\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"EMG1 (Nep1) is an essential nucleolar SPOUT-class pseudouridine-N1-methyltransferase that methylates a specific pseudouridine in the small ribosomal subunit rRNA (Ψ1191 in yeast 18S rRNA), functions as a ribosome assembly factor for 40S subunit biogenesis (with a separable essential role beyond its catalytic activity), forms a nucleolar subcomplex with NOP14/NOC4L/UTP14A required for its nucleolar recruitment, is imported into the nucleus via importins Impα/β or Impβ/7 (which also chaperone it against RNA aggregation), and in development cooperates with the transcription factor GRHL3 in β-catenin-mediated surface ectoderm differentiation; its methyltransferase activity is additionally required for KSHV-induced specialized ribosome function, and the disease-causing D86G (Bowen-Conradi syndrome) mutation enhances dimerization, promotes protein aggregation and proteasomal degradation, and impairs nucleolar accumulation without abolishing catalytic activity.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"EMG1 (Nep1) is an essential nucleolar SPOUT-class methyltransferase that functions in biogenesis of the small (40S) ribosomal subunit, where its depletion selectively reduces 20S pre-rRNA and mature 18S rRNA [#0, #1]. Enzymatically it is a pseudouridine-N1-specific methyltransferase that transfers a methyl group to the N1 of pseudouridine, generating the m1acp3-\\u03a8 hypermodification at \\u03a81191 of 18S rRNA in yeast (\\u03a8926 in archaeal 16S rRNA), with catalysis depending on a conserved active-site aspartate acting as a base [#3, #4, #16, #15]. Structures show EMG1 adopts an alpha/beta knot fold with a preformed SAM-binding pocket, acts as a homodimer, and recognizes its target via base-specific contacts that remodel the bound rRNA stem-loop [#1, #2, #6]. EMG1 has a catalytically separable, essential role in ribosome assembly, since mutations that abolish SAM binding or block \\u03a81191 methylation do not prevent ribosome biogenesis or growth [#1, #4]. It is imported into the nucleus by Imp\\u03b1/\\u03b2 or Imp\\u03b2/7, which also chaperone its basic regions against nonspecific RNA aggregation, and is recruited to the nucleolus within a subcomplex containing NOP14, NOC4L, and UTP14A [#10]. Beyond ribosome assembly, EMG1 is required for early embryonic development and nucleologenesis [#7], cooperates with the transcription factor GRHL3 to drive nuclear GRHL3 localization and Wnt/\\u03b2-catenin\\u2013mediated surface ectoderm differentiation [#13], and its methyltransferase activity supports KSHV-induced specialized ribosome scanning of viral uORFs [#14]. The D86G missense mutation causes Bowen-Conradi syndrome; it does not abolish catalytic activity but enhances dimerization and RNA affinity, promotes aggregation and proteasomal degradation, and impairs nucleolar accumulation [#5, #4, #10].\",\n  \"teleology\": [\n    {\n      \"year\": 2001,\n      \"claim\": \"Established EMG1 as a 40S ribosomal subunit biogenesis factor whose nuclear function depends on a defined protein partner, framing it as a nucleolar assembly component rather than a free-standing enzyme.\",\n      \"evidence\": \"Genetic depletion, ts allele and rRNA Northern blotting in yeast with Nop14 interaction and localization assays\",\n      \"pmids\": [\"11694595\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular activity unknown at this stage\", \"Did not define which pre-rRNA processing step requires EMG1\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Defined EMG1 as a SPOUT-class methyltransferase by structure and, critically, showed that catalytic activity is dispensable for ribosome biogenesis, revealing a separable non-catalytic essential role.\",\n      \"evidence\": \"X-ray crystallography of yeast Emg1 with SAM, SAM- and RNA-binding mutagenesis, yeast complementation; archaeal Nep1 structures with cofactor analogs\",\n      \"pmids\": [\"18063569\", \"18208838\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"RNA substrate and methylated position not yet identified\", \"Nature of the catalysis-independent essential function unresolved\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Identified the precise enzymatic activity and target: N1-methylation of pseudouridine at \\u03a81191 of 18S rRNA, and confirmed a dual catalytic / assembly-factor function.\",\n      \"evidence\": \"In vitro methyltransferase assays with MS/HPLC/NMR, in vivo 14C-methionine labeling, ESI-MS, and genetic suppressor/SAM rescue in yeast; protein-RNA co-crystal structures\",\n      \"pmids\": [\"20047967\", \"20972225\", \"21087996\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism by which assembly function is separable from methylation not defined\", \"rRNA remodeling role inferred from static structures\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Demonstrated organismal essentiality, showing EMG1 loss arrests early mouse development with nucleologenesis defects in a p53-independent manner.\",\n      \"evidence\": \"Mouse knockout, embryo phenotyping, genetic epistasis with p53 null\",\n      \"pmids\": [\"20858271\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not resolve whether lethality reflects catalytic or assembly role\", \"Cell-type-specific requirements not addressed\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Linked EMG1 to human disease by identifying D86G as the cause of Bowen-Conradi syndrome and indicating a protein-stability/aggregation defect.\",\n      \"evidence\": \"Patient sequencing, Western blotting, yeast two-hybrid, mammalian overexpression, structural modeling\",\n      \"pmids\": [\"19463982\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Aggregation inferred indirectly from solubility and two-hybrid data\", \"Structural modeling is computational, not experimental\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Placed EMG1 within a genetic network for early pre-ribosome assembly, linking it functionally to Utp30 and the surface for Rps19 loading onto the 90S pre-ribosome.\",\n      \"evidence\": \"Genome-wide synthetic genetic array with biochemical follow-up and complementation in yeast\",\n      \"pmids\": [\"22588997\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Genetic interactions do not establish direct physical contacts\", \"Single-lab screen without orthogonal validation of all hits\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Resolved how EMG1 reaches and is protected within the nucleolus, defining importin-mediated import/chaperoning and a NOP14/NOC4L/UTP14A subcomplex required for nucleolar recruitment, and recast D86G pathology as a trafficking/degradation defect.\",\n      \"evidence\": \"Nuclear import and in vitro aggregation assays, Co-IP, proteasome inhibition, fluorescence microscopy\",\n      \"pmids\": [\"27798105\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Stoichiometry and assembly order of the subcomplex not defined\", \"Conflicts with patient-cell data on D86G nucleolar localization\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Characterized cellular consequences of D86G in patient cells, showing delayed 18S processing and G2/M accumulation despite normal steady-state 40S and translation, indicating a partial/kinetic defect.\",\n      \"evidence\": \"Flow cytometry, proliferation and pulse-chase rRNA processing assays, immunofluorescence in BCS lymphoblasts\",\n      \"pmids\": [\"26676230\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Reports normal D86G nucleolar localization, conflicting with overexpression studies\", \"Mechanism connecting processing delay to G2/M arrest unresolved\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Defined the chemical catalytic mechanism, identifying a conserved aspartate as the catalytic base for pseudouridine N1 deprotonation and a water-mediated proton-transfer pathway.\",\n      \"evidence\": \"Molecular dynamics, DFT calculations, site-directed mutagenesis and kinetic assays\",\n      \"pmids\": [\"37649713\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism is largely computational and not independently replicated\", \"Catalytic detail does not address the non-catalytic assembly role\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Extended EMG1 function beyond ribosome assembly into development and viral biology, showing a GRHL3 partnership driving Wnt/\\u03b2-catenin\\u2013dependent ectoderm/neural tube morphogenesis and a methyltransferase-dependent role in KSHV specialized ribosomes; also added a fresh archaeal structure with defined RNA target.\",\n      \"evidence\": \"Conditional knockout mice with protein complex identification and epistasis (GRHL3); EMG1 depletion with ribosome occupancy and viral output assays (KSHV); P. horikoshii crystal structures with in vitro methyltransferase assays\",\n      \"pmids\": [\"40761126\", \"40215162\", \"39918246\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How a ribosome assembly factor controls GRHL3 nuclear localization mechanistically is unclear\", \"Whether developmental roles require catalysis vs assembly function not separated\", \"Generality of specialized-ribosome role beyond KSHV unknown\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Tied EMG1 abundance to a cancer-relevant regulatory axis, showing FTO-mediated m6A demethylation of EMG1 mRNA reduces EMG1, ribosome biosynthesis, and bladder tumor cell aggressiveness.\",\n      \"evidence\": \"FTO gain/loss-of-function, m6A analysis, in vitro and in vivo bladder tumor models\",\n      \"pmids\": [\"41962773\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct m6A site on EMG1 mRNA and reader involvement not fully defined\", \"Causality between EMG1 level and tumor phenotype is correlative in part\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The molecular basis of EMG1's catalysis-independent essential ribosome assembly function, and how this is mechanistically distinct from its non-ribosomal roles in GRHL3-mediated development and specialized translation, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No structure of EMG1 within an intact pre-40S particle\", \"No separation-of-function alleles distinguishing assembly vs developmental roles\", \"Mechanism linking nucleolar assembly factor to nuclear transcription-factor function unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140098\", \"supporting_discovery_ids\": [3, 4, 6, 16]},\n      {\"term_id\": \"GO:0016740\", \"supporting_discovery_ids\": [1, 3, 15]},\n      {\"term_id\": \"GO:0003723\", \"supporting_discovery_ids\": [1, 6, 16]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005730\", \"supporting_discovery_ids\": [0, 9, 10]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [0, 10]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [0, 4]},\n      {\"term_id\": \"R-HSA-1266738\", \"supporting_discovery_ids\": [7, 13]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [12, 13]}\n    ],\n    \"complexes\": [\n      \"EMG1/NOP14/NOC4L/UTP14A nucleolar subcomplex\",\n      \"EMG1-GRHL3 complex\"\n    ],\n    \"partners\": [\n      \"NOP14\",\n      \"NOC4L\",\n      \"UTP14A\",\n      \"GRHL3\",\n      \"UTP30\",\n      \"RPS19\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}