{"gene":"EDEM2","run_date":"2026-06-09T23:54:42","timeline":{"discoveries":[{"year":2004,"finding":"EDEM2 (C20orf31) localizes to the ER lumen in HEK293 cells, associates with misfolded alpha1-antitrypsin, and its overexpression accelerates degradation of misfolded alpha1-antitrypsin, establishing its role in ERAD. Recombinant EDEM2 showed no detectable alpha-1,2 mannosidase activity in vitro.","method":"Recombinant protein expression, subcellular fractionation/ER localization, co-immunoprecipitation with misfolded alpha1-antitrypsin, pulse-chase degradation assay, in vitro mannosidase activity assay","journal":"Glycobiology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP, localization, and functional overexpression assay in single lab; in vitro mannosidase assay provided negative result for enzymatic activity","pmids":["15537790"],"is_preprint":false},{"year":2014,"finding":"EDEM2 catalyzes the first mannose trimming step in mammalian gpERAD, converting Man9GlcNAc2 to Man8GlcNAc2, as determined by TALEN-mediated gene knockout in human cells. EDEM2 knockout cells accumulate Man9GlcNAc2, establishing EDEM2 as an alpha-1,2-mannosidase with a distinct upstream role from EDEM1 and EDEM3.","method":"TALEN-mediated gene knockout in human cell line, glycan structure analysis, genetic epistasis with EDEM1/2/3 single and combined knockouts","journal":"The Journal of cell biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — TALEN KO with defined glycan phenotype, multiple gene knockouts compared, replicated by independent lab (George et al. 2020)","pmids":["25092655"],"is_preprint":false},{"year":2014,"finding":"EDEM2 is required for ERAD of both glycosylated and non-glycosylated sonic hedgehog (SHH), indicating it can recognize misfolded polypeptide backbone independent of glycan trimming. EDEM2 interacts with non-glycosylated SHH (N278A mutant), as well as with calnexin and SEL1L.","method":"siRNA knockdown of EDEM1/2/3, degradation assays, co-immunoprecipitation with non-glycosylated SHH variant","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP and functional knockdown with specific substrate, single lab, two orthogonal methods","pmids":["24910992"],"is_preprint":false},{"year":2014,"finding":"EDEM2 promotes retrotranslocation of ricin A-chain (RTA) out of the ER to the cytosol, independently of ER translocon accessibility, and shows greater interaction with ricin than EDEM1 does. The interaction with RTA depends on RTA's structural conformation.","method":"Co-immunoprecipitation, pull-down assay, ricin cytotoxicity/retrotranslocation assay with EDEM2 knockdown","journal":"The Biochemical journal","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — co-IP/pulldown plus functional retrotranslocation assay, single lab, two methods","pmids":["24200403"],"is_preprint":false},{"year":2015,"finding":"Substrate recognition by EDEM2 depends on the hydrophobicity of protein determinants: mutations increasing or decreasing hydrophobicity of ERAD substrates (ricin, BACE457) correspondingly increase or decrease EDEM2 binding, and EDEM2 can bind hydrophobic transmembrane regions of misfolded substrates. Binding does not require substrate glycosylation.","method":"Hydrophobicity-modulating mutagenesis of ERAD substrates, co-immunoprecipitation in human cells","journal":"BMC cell biology","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — mutagenesis combined with co-IP in human cells, single lab, two orthogonal approaches","pmids":["25655076"],"is_preprint":false},{"year":2018,"finding":"EDEM2 possesses mannosidase activity in vitro that is dependent on the unfolded/denatured state of glycoprotein substrates; activity is modest on free oligosaccharides or folded glycoproteins but significantly higher on denatured glycoproteins. EDEM2 associates with the oxidoreductase TXNDC11, which enhances its mannosidase activity on glycoproteins but not on free N-glycans.","method":"In vitro mannosidase activity assay with free N-glycans, folded, and denatured glycoprotein substrates; co-immunoprecipitation with TXNDC11 and other oxidoreductases","journal":"Communications biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — direct in vitro enzymatic assay with substrate unfolding controls plus co-IP for partner identification, single lab but multiple orthogonal methods","pmids":["30374462"],"is_preprint":false},{"year":2020,"finding":"EDEM2 forms a stable disulfide bond with TXNDC11 via C558 on EDEM2 (outside the mannosidase homology domain) linked to C692 in Trx5 domain of TXNDC11 (which uniquely carries the CXXC motif). This covalent EDEM2-TXNDC11 complex is essential for mannose trimming (Man9GlcNAc2 → Man8GlcNAc2 isomer B) and subsequent gpERAD in HCT116 cells. Purified EDEM2-TXNDC11 complex converts Man9GlcNAc2 to Man8GlcNAc2(isomerB) in vitro, providing the first clear demonstration of in vitro mannosidase activity of an EDEM family protein.","method":"CRISPR/KO, site-directed mutagenesis of EDEM2 (C558) and TXNDC11 (C692), disulfide bond mapping, purification of EDEM2-TXNDC11 complex from transfected cells, in vitro mannosidase assay with Man9GlcNAc2 substrate, glycan structure analysis by MS","journal":"eLife","confidence":"High","confidence_rationale":"Tier 1 / Strong — reconstituted in vitro mannosidase activity, mutagenesis identifying specific disulfide-bonded residues, KO validation in cells, multiple orthogonal methods in single rigorous study","pmids":["32065582"],"is_preprint":false},{"year":2021,"finding":"Affinity proteomics identified seven novel EDEM2-associated proteins, all involved in ER quality control and ERAD. Novel endogenous EDEM2-dependent ERAD substrate candidates were identified, including integrin alpha-1 and protocadherin 2, whose levels were negatively correlated with EDEM2 expression in melanoma cells.","method":"Affinity proteomics (co-IP/MS), sucrose density fractionation proteomics, deglycoproteomics (ConA enrichment + endoglycosidase digestion + nanoLC-MS/MS), SILAC-based pulse proteomics, biochemical validation","journal":"Molecular & cellular proteomics : MCP","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — MS-based affinity proteomics with multiple orthogonal methods (SDF, SILAC, deglycoproteomics), single lab","pmids":["34332121"],"is_preprint":false},{"year":2025,"finding":"EDEM2 is transcriptionally regulated by XBP1s (spliced XBP1) in cardiomyocytes. EDEM2 promotes SEC23A-mediated intracellular translocation of ATGL (adipose triglyceride lipase), thereby inhibiting ATGL degradation and preventing excessive lipid droplet accumulation in cardiomyocytes. EDEM2 knockdown leads to lipid droplet accumulation and elevated triglycerides/diglycerides, effects that are ATGL-dependent.","method":"AAV9-mediated loss- and gain-of-function in mice, siRNA knockdown in rat cardiomyocytes and hiPSC-derived cardiomyocytes, luciferase reporter assay for XBP1s-EDEM2 transcriptional regulation, mass spectrometry proteomics, lipidomic analysis, transmission electron microscopy","journal":"Circulation","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo KO/OE with defined lipid phenotype, luciferase reporter for transcriptional regulation, multiple orthogonal methods, single lab","pmids":["40130322"],"is_preprint":false}],"current_model":"EDEM2 is an ER-resident alpha-1,2-mannosidase that initiates glycoprotein ERAD by catalyzing the first mannose trimming step (Man9GlcNAc2 → Man8GlcNAc2 isomer B); this activity requires stable disulfide bonding of EDEM2 C558 to TXNDC11 C692 and is enhanced by substrate unfolding, while EDEM2 also recognizes misfolded substrates through hydrophobic polypeptide determinants independent of glycosylation, interacts with calnexin and SEL1L to channel substrates toward ERAD, and—outside its canonical ERAD role—is transcriptionally induced by XBP1s in cardiomyocytes where it promotes SEC23A-mediated ATGL translocation to regulate lipid homeostasis."},"narrative":{"mechanistic_narrative":"EDEM2 is an ER-luminal factor that initiates glycoprotein ER-associated degradation (ERAD) by catalyzing the first mannose-trimming step, converting Man9GlcNAc2 to the Man8GlcNAc2 isomer B that commits misfolded glycoproteins to disposal [PMID:25092655]. Its alpha-1,2-mannosidase activity is conditional: it acts efficiently on denatured glycoproteins rather than free oligosaccharides or folded substrates, coupling catalysis to the unfolded state of its targets [PMID:30374462]. This activity is contingent on a stable disulfide bond between EDEM2 C558 and C692 in the Trx5 (CXXC) domain of the oxidoreductase TXNDC11; the covalent EDEM2–TXNDC11 complex reconstitutes Man9-to-Man8 trimming in vitro and is required for downstream gpERAD in cells [PMID:30374462, PMID:32065582]. Beyond glycan recognition, EDEM2 engages substrates through hydrophobic polypeptide determinants independent of glycosylation, binding misfolded and non-glycosylated substrates and exposed hydrophobic regions [PMID:24910992, PMID:25655076], and it interfaces with the ER quality-control machinery including calnexin and SEL1L to channel substrates toward degradation [PMID:24910992]. Outside its canonical ERAD role, EDEM2 is transcriptionally induced by XBP1s in cardiomyocytes, where it promotes SEC23A-mediated translocation of the lipase ATGL to restrain lipid droplet accumulation and regulate cardiac lipid homeostasis [PMID:40130322].","teleology":[{"year":2004,"claim":"Established EDEM2 as an ER-resident ERAD factor before any enzymatic role was known, answering whether the orphan ORF participated in misfolded-glycoprotein disposal.","evidence":"ER localization, co-IP with misfolded alpha1-antitrypsin, and overexpression degradation assay in HEK293, with a negative in vitro mannosidase result","pmids":["15537790"],"confidence":"Medium","gaps":["No enzymatic activity detected, leaving its mechanism of action undefined","Single substrate (alpha1-antitrypsin) and single lab","Did not distinguish direct catalysis from adaptor function"]},{"year":2014,"claim":"Defined EDEM2's specific catalytic position in the ERAD mannose-trimming cascade, resolving how it differs from EDEM1 and EDEM3.","evidence":"TALEN knockout in human cells with glycan structure analysis and epistasis across EDEM1/2/3 knockouts","pmids":["25092655"],"confidence":"High","gaps":["Did not demonstrate direct enzymatic activity in vitro at this stage","Cofactor requirement for the activity unknown","Did not address non-glycan substrate recognition"]},{"year":2014,"claim":"Showed EDEM2 recognizes substrates beyond their glycans and physically links to core ER quality-control machinery, broadening its role from pure mannosidase to substrate-channeling factor.","evidence":"siRNA knockdown plus co-IP with non-glycosylated SHH, calnexin, and SEL1L; ricin A-chain retrotranslocation assays","pmids":["24910992","24200403"],"confidence":"Medium","gaps":["Molecular basis of polypeptide recognition not yet defined","Direct vs. indirect interaction with calnexin/SEL1L not resolved","Single lab per substrate"]},{"year":2015,"claim":"Identified hydrophobicity of substrate determinants as the basis for glycosylation-independent recognition, explaining how EDEM2 engages misfolded polypeptide regions.","evidence":"Hydrophobicity-modulating mutagenesis of ricin and BACE457 with co-IP in human cells","pmids":["25655076"],"confidence":"Medium","gaps":["No structural map of the polypeptide-binding surface","Relationship between hydrophobic binding and mannosidase catalysis unclear","Single lab"]},{"year":2018,"claim":"Demonstrated that EDEM2's mannosidase activity is gated by substrate unfolding and enhanced by an oxidoreductase partner, reconciling earlier negative in vitro activity results.","evidence":"In vitro mannosidase assays comparing free N-glycans, folded and denatured glycoproteins, plus co-IP identifying TXNDC11","pmids":["30374462"],"confidence":"High","gaps":["Did not define the chemical nature of the EDEM2-TXNDC11 link","Mechanism by which unfolding enhances activity not resolved"]},{"year":2020,"claim":"Pinpointed the covalent EDEM2-TXNDC11 disulfide as essential for catalysis and reconstituted the first clear in vitro EDEM-family mannosidase activity, settling how EDEM2 becomes enzymatically competent.","evidence":"CRISPR KO, site-directed mutagenesis of EDEM2 C558 and TXNDC11 C692, disulfide mapping, complex purification, and in vitro Man9-to-Man8 trimming with MS glycan analysis in HCT116","pmids":["32065582"],"confidence":"High","gaps":["No high-resolution structure of the complex","How TXNDC11 redox state modulates activity not defined","Connection between disulfide-driven catalysis and hydrophobic substrate recognition unaddressed"]},{"year":2021,"claim":"Mapped the EDEM2 interactome and endogenous substrate landscape, extending its role beyond model substrates to ER quality-control networks and disease-relevant targets.","evidence":"Affinity proteomics, sucrose density fractionation, deglycoproteomics, and SILAC pulse proteomics in melanoma cells with biochemical validation","pmids":["34332121"],"confidence":"Medium","gaps":["Substrate candidates (integrin alpha-1, protocadherin 2) inferred from correlation, not direct degradation kinetics","Functional roles of novel interactors not characterized","Single cell-type context"]},{"year":2025,"claim":"Revealed a non-canonical EDEM2 function in cardiac lipid homeostasis under XBP1s control, distinguishing its ERAD role from a lipid-regulatory role.","evidence":"AAV9 loss/gain-of-function in mice, siRNA in rat and hiPSC cardiomyocytes, luciferase reporter for XBP1s regulation, proteomics, lipidomics, and EM","pmids":["40130322"],"confidence":"Medium","gaps":["Mechanistic link between EDEM2 and SEC23A-mediated ATGL translocation not resolved at molecular level","Whether mannosidase activity is required for the lipid role is untested","Cardiomyocyte-specific; generality unknown"]},{"year":null,"claim":"How EDEM2's two recognition modes — glycan trimming and hydrophobicity-based polypeptide binding — are structurally integrated, and whether the lipid-regulatory function depends on its catalytic activity, remain open.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No crystal/cryo-EM structure of the EDEM2-TXNDC11 complex with substrate","Mechanistic coupling of catalysis to ERAD channeling unresolved","Catalytic dependence of the ATGL/lipid pathway untested"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140098","term_label":"catalytic activity, acting on RNA","supporting_discovery_ids":[1,5,6]},{"term_id":"GO:0016787","term_label":"hydrolase activity","supporting_discovery_ids":[1,6]}],"localization":[{"term_id":"GO:0005783","term_label":"endoplasmic reticulum","supporting_discovery_ids":[0]}],"pathway":[{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[0,1,2,6]},{"term_id":"R-HSA-8953897","term_label":"Cellular responses to stimuli","supporting_discovery_ids":[8]}],"complexes":["EDEM2-TXNDC11 disulfide-linked complex"],"partners":["TXNDC11","CANX","SEL1L","SEC23A"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9BV94","full_name":"ER degradation-enhancing alpha-mannosidase-like protein 2","aliases":[],"length_aa":578,"mass_kda":64.8,"function":"Involved in the endoplasmic reticulum-associated degradation (ERAD) pathway that targets misfolded glycoproteins for degradation in an N-glycan-dependent manner (PubMed:15537790, PubMed:25092655). May initiate ERAD by promoting the first mannose trimming step of ERAD substrates, from Man9GlcNAc2 to Man8GlcNAc2 (PubMed:25092655). Seems to recognize and bind to exposed hydrophobic regions in target proteins (By similarity)","subcellular_location":"Endoplasmic reticulum lumen","url":"https://www.uniprot.org/uniprotkb/Q9BV94/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/EDEM2","classification":"Not Classified","n_dependent_lines":10,"n_total_lines":1208,"dependency_fraction":0.008278145695364239},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/EDEM2","total_profiled":1310},"omim":[{"mim_id":"610302","title":"ENDOPLASMIC RETICULUM DEGRADATION-ENHANCING ALPHA-MANNOSIDASE-LIKE PROTEIN 2; EDEM2","url":"https://www.omim.org/entry/610302"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/EDEM2"},"hgnc":{"alias_symbol":["FLJ10783","bA4204.1"],"prev_symbol":["C20orf49","C20orf31"]},"alphafold":{"accession":"Q9BV94","domains":[{"cath_id":"1.50.10.10","chopping":"31-69_329-449","consensus_level":"medium","plddt":97.1114,"start":31,"end":449},{"cath_id":"1.50.10.10","chopping":"72-310_460-465_479-504","consensus_level":"medium","plddt":94.9207,"start":72,"end":504}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9BV94","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9BV94-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9BV94-F1-predicted_aligned_error_v6.png","plddt_mean":85.38},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=EDEM2","jax_strain_url":"https://www.jax.org/strain/search?query=EDEM2"},"sequence":{"accession":"Q9BV94","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9BV94.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9BV94/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9BV94"}},"corpus_meta":[{"pmid":"15537790","id":"PMC_15537790","title":"Human EDEM2, a novel homolog of family 47 glycosidases, is involved in ER-associated degradation of glycoproteins.","date":"2004","source":"Glycobiology","url":"https://pubmed.ncbi.nlm.nih.gov/15537790","citation_count":122,"is_preprint":false},{"pmid":"25092655","id":"PMC_25092655","title":"EDEM2 initiates mammalian glycoprotein ERAD by catalyzing the first mannose trimming step.","date":"2014","source":"The Journal of cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/25092655","citation_count":121,"is_preprint":false},{"pmid":"17499246","id":"PMC_17499246","title":"Glycoprotein folding and the role of EDEM1, EDEM2 and EDEM3 in degradation of folding-defective glycoproteins.","date":"2007","source":"FEBS letters","url":"https://pubmed.ncbi.nlm.nih.gov/17499246","citation_count":112,"is_preprint":false},{"pmid":"30374462","id":"PMC_30374462","title":"Mannosidase activity of EDEM1 and EDEM2 depends on an unfolded state of their glycoprotein substrates.","date":"2018","source":"Communications biology","url":"https://pubmed.ncbi.nlm.nih.gov/30374462","citation_count":46,"is_preprint":false},{"pmid":"32065582","id":"PMC_32065582","title":"EDEM2 stably disulfide-bonded to TXNDC11 catalyzes the first mannose trimming step in mammalian glycoprotein ERAD.","date":"2020","source":"eLife","url":"https://pubmed.ncbi.nlm.nih.gov/32065582","citation_count":38,"is_preprint":false},{"pmid":"24910992","id":"PMC_24910992","title":"EDEM2 and OS-9 are required for ER-associated degradation of non-glycosylated sonic hedgehog.","date":"2014","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/24910992","citation_count":23,"is_preprint":false},{"pmid":"25655076","id":"PMC_25655076","title":"Hydrophobicity of protein determinants influences the recognition of substrates by EDEM1 and EDEM2 in human cells.","date":"2015","source":"BMC cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/25655076","citation_count":19,"is_preprint":false},{"pmid":"40130322","id":"PMC_40130322","title":"XBP1s-EDEM2 Prevents the Onset and Development of HFpEF by Ameliorating Cardiac Lipotoxicity.","date":"2025","source":"Circulation","url":"https://pubmed.ncbi.nlm.nih.gov/40130322","citation_count":16,"is_preprint":false},{"pmid":"34332121","id":"PMC_34332121","title":"Affinity Proteomics and Deglycoproteomics Uncover Novel EDEM2 Endogenous Substrates and an Integrative ERAD Network.","date":"2021","source":"Molecular & cellular proteomics : MCP","url":"https://pubmed.ncbi.nlm.nih.gov/34332121","citation_count":16,"is_preprint":false},{"pmid":"24200403","id":"PMC_24200403","title":"The role of EDEM2 compared with EDEM1 in ricin transport from the endoplasmic reticulum to the cytosol.","date":"2014","source":"The Biochemical journal","url":"https://pubmed.ncbi.nlm.nih.gov/24200403","citation_count":13,"is_preprint":false},{"pmid":"32556999","id":"PMC_32556999","title":"A novel gene in early childhood diabetes: EDEM2 silencing decreases SLC2A2 and PXD1 expression, leading to impaired insulin secretion.","date":"2020","source":"Molecular genetics and genomics : MGG","url":"https://pubmed.ncbi.nlm.nih.gov/32556999","citation_count":9,"is_preprint":false},{"pmid":"34712749","id":"PMC_34712749","title":"Dataset of human EDEM2 melanoma cells proteomics, affinity proteomics and deglycoproteomics.","date":"2021","source":"Data in brief","url":"https://pubmed.ncbi.nlm.nih.gov/34712749","citation_count":7,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":8540,"output_tokens":2740,"usd":0.03336,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":9879,"output_tokens":3133,"usd":0.06386,"stage2_stop_reason":"end_turn"},"total_usd":0.09722,"stage1_batch_id":"msgbatch_01AsR7g8dGTEiEw9U83bJeWn","stage2_batch_id":"msgbatch_01YV5ZyxtFNUZEjxRcpd9jzb","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2004,\n      \"finding\": \"EDEM2 (C20orf31) localizes to the ER lumen in HEK293 cells, associates with misfolded alpha1-antitrypsin, and its overexpression accelerates degradation of misfolded alpha1-antitrypsin, establishing its role in ERAD. Recombinant EDEM2 showed no detectable alpha-1,2 mannosidase activity in vitro.\",\n      \"method\": \"Recombinant protein expression, subcellular fractionation/ER localization, co-immunoprecipitation with misfolded alpha1-antitrypsin, pulse-chase degradation assay, in vitro mannosidase activity assay\",\n      \"journal\": \"Glycobiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP, localization, and functional overexpression assay in single lab; in vitro mannosidase assay provided negative result for enzymatic activity\",\n      \"pmids\": [\"15537790\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"EDEM2 catalyzes the first mannose trimming step in mammalian gpERAD, converting Man9GlcNAc2 to Man8GlcNAc2, as determined by TALEN-mediated gene knockout in human cells. EDEM2 knockout cells accumulate Man9GlcNAc2, establishing EDEM2 as an alpha-1,2-mannosidase with a distinct upstream role from EDEM1 and EDEM3.\",\n      \"method\": \"TALEN-mediated gene knockout in human cell line, glycan structure analysis, genetic epistasis with EDEM1/2/3 single and combined knockouts\",\n      \"journal\": \"The Journal of cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — TALEN KO with defined glycan phenotype, multiple gene knockouts compared, replicated by independent lab (George et al. 2020)\",\n      \"pmids\": [\"25092655\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"EDEM2 is required for ERAD of both glycosylated and non-glycosylated sonic hedgehog (SHH), indicating it can recognize misfolded polypeptide backbone independent of glycan trimming. EDEM2 interacts with non-glycosylated SHH (N278A mutant), as well as with calnexin and SEL1L.\",\n      \"method\": \"siRNA knockdown of EDEM1/2/3, degradation assays, co-immunoprecipitation with non-glycosylated SHH variant\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP and functional knockdown with specific substrate, single lab, two orthogonal methods\",\n      \"pmids\": [\"24910992\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"EDEM2 promotes retrotranslocation of ricin A-chain (RTA) out of the ER to the cytosol, independently of ER translocon accessibility, and shows greater interaction with ricin than EDEM1 does. The interaction with RTA depends on RTA's structural conformation.\",\n      \"method\": \"Co-immunoprecipitation, pull-down assay, ricin cytotoxicity/retrotranslocation assay with EDEM2 knockdown\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — co-IP/pulldown plus functional retrotranslocation assay, single lab, two methods\",\n      \"pmids\": [\"24200403\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Substrate recognition by EDEM2 depends on the hydrophobicity of protein determinants: mutations increasing or decreasing hydrophobicity of ERAD substrates (ricin, BACE457) correspondingly increase or decrease EDEM2 binding, and EDEM2 can bind hydrophobic transmembrane regions of misfolded substrates. Binding does not require substrate glycosylation.\",\n      \"method\": \"Hydrophobicity-modulating mutagenesis of ERAD substrates, co-immunoprecipitation in human cells\",\n      \"journal\": \"BMC cell biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — mutagenesis combined with co-IP in human cells, single lab, two orthogonal approaches\",\n      \"pmids\": [\"25655076\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"EDEM2 possesses mannosidase activity in vitro that is dependent on the unfolded/denatured state of glycoprotein substrates; activity is modest on free oligosaccharides or folded glycoproteins but significantly higher on denatured glycoproteins. EDEM2 associates with the oxidoreductase TXNDC11, which enhances its mannosidase activity on glycoproteins but not on free N-glycans.\",\n      \"method\": \"In vitro mannosidase activity assay with free N-glycans, folded, and denatured glycoprotein substrates; co-immunoprecipitation with TXNDC11 and other oxidoreductases\",\n      \"journal\": \"Communications biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — direct in vitro enzymatic assay with substrate unfolding controls plus co-IP for partner identification, single lab but multiple orthogonal methods\",\n      \"pmids\": [\"30374462\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"EDEM2 forms a stable disulfide bond with TXNDC11 via C558 on EDEM2 (outside the mannosidase homology domain) linked to C692 in Trx5 domain of TXNDC11 (which uniquely carries the CXXC motif). This covalent EDEM2-TXNDC11 complex is essential for mannose trimming (Man9GlcNAc2 → Man8GlcNAc2 isomer B) and subsequent gpERAD in HCT116 cells. Purified EDEM2-TXNDC11 complex converts Man9GlcNAc2 to Man8GlcNAc2(isomerB) in vitro, providing the first clear demonstration of in vitro mannosidase activity of an EDEM family protein.\",\n      \"method\": \"CRISPR/KO, site-directed mutagenesis of EDEM2 (C558) and TXNDC11 (C692), disulfide bond mapping, purification of EDEM2-TXNDC11 complex from transfected cells, in vitro mannosidase assay with Man9GlcNAc2 substrate, glycan structure analysis by MS\",\n      \"journal\": \"eLife\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — reconstituted in vitro mannosidase activity, mutagenesis identifying specific disulfide-bonded residues, KO validation in cells, multiple orthogonal methods in single rigorous study\",\n      \"pmids\": [\"32065582\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Affinity proteomics identified seven novel EDEM2-associated proteins, all involved in ER quality control and ERAD. Novel endogenous EDEM2-dependent ERAD substrate candidates were identified, including integrin alpha-1 and protocadherin 2, whose levels were negatively correlated with EDEM2 expression in melanoma cells.\",\n      \"method\": \"Affinity proteomics (co-IP/MS), sucrose density fractionation proteomics, deglycoproteomics (ConA enrichment + endoglycosidase digestion + nanoLC-MS/MS), SILAC-based pulse proteomics, biochemical validation\",\n      \"journal\": \"Molecular & cellular proteomics : MCP\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — MS-based affinity proteomics with multiple orthogonal methods (SDF, SILAC, deglycoproteomics), single lab\",\n      \"pmids\": [\"34332121\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"EDEM2 is transcriptionally regulated by XBP1s (spliced XBP1) in cardiomyocytes. EDEM2 promotes SEC23A-mediated intracellular translocation of ATGL (adipose triglyceride lipase), thereby inhibiting ATGL degradation and preventing excessive lipid droplet accumulation in cardiomyocytes. EDEM2 knockdown leads to lipid droplet accumulation and elevated triglycerides/diglycerides, effects that are ATGL-dependent.\",\n      \"method\": \"AAV9-mediated loss- and gain-of-function in mice, siRNA knockdown in rat cardiomyocytes and hiPSC-derived cardiomyocytes, luciferase reporter assay for XBP1s-EDEM2 transcriptional regulation, mass spectrometry proteomics, lipidomic analysis, transmission electron microscopy\",\n      \"journal\": \"Circulation\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo KO/OE with defined lipid phenotype, luciferase reporter for transcriptional regulation, multiple orthogonal methods, single lab\",\n      \"pmids\": [\"40130322\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"EDEM2 is an ER-resident alpha-1,2-mannosidase that initiates glycoprotein ERAD by catalyzing the first mannose trimming step (Man9GlcNAc2 → Man8GlcNAc2 isomer B); this activity requires stable disulfide bonding of EDEM2 C558 to TXNDC11 C692 and is enhanced by substrate unfolding, while EDEM2 also recognizes misfolded substrates through hydrophobic polypeptide determinants independent of glycosylation, interacts with calnexin and SEL1L to channel substrates toward ERAD, and—outside its canonical ERAD role—is transcriptionally induced by XBP1s in cardiomyocytes where it promotes SEC23A-mediated ATGL translocation to regulate lipid homeostasis.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"EDEM2 is an ER-luminal factor that initiates glycoprotein ER-associated degradation (ERAD) by catalyzing the first mannose-trimming step, converting Man9GlcNAc2 to the Man8GlcNAc2 isomer B that commits misfolded glycoproteins to disposal [#1]. Its alpha-1,2-mannosidase activity is conditional: it acts efficiently on denatured glycoproteins rather than free oligosaccharides or folded substrates, coupling catalysis to the unfolded state of its targets [#5]. This activity is contingent on a stable disulfide bond between EDEM2 C558 and C692 in the Trx5 (CXXC) domain of the oxidoreductase TXNDC11; the covalent EDEM2–TXNDC11 complex reconstitutes Man9-to-Man8 trimming in vitro and is required for downstream gpERAD in cells [#5, #6]. Beyond glycan recognition, EDEM2 engages substrates through hydrophobic polypeptide determinants independent of glycosylation, binding misfolded and non-glycosylated substrates and exposed hydrophobic regions [#2, #4], and it interfaces with the ER quality-control machinery including calnexin and SEL1L to channel substrates toward degradation [#2]. Outside its canonical ERAD role, EDEM2 is transcriptionally induced by XBP1s in cardiomyocytes, where it promotes SEC23A-mediated translocation of the lipase ATGL to restrain lipid droplet accumulation and regulate cardiac lipid homeostasis [#8].\",\n  \"teleology\": [\n    {\n      \"year\": 2004,\n      \"claim\": \"Established EDEM2 as an ER-resident ERAD factor before any enzymatic role was known, answering whether the orphan ORF participated in misfolded-glycoprotein disposal.\",\n      \"evidence\": \"ER localization, co-IP with misfolded alpha1-antitrypsin, and overexpression degradation assay in HEK293, with a negative in vitro mannosidase result\",\n      \"pmids\": [\"15537790\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No enzymatic activity detected, leaving its mechanism of action undefined\", \"Single substrate (alpha1-antitrypsin) and single lab\", \"Did not distinguish direct catalysis from adaptor function\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Defined EDEM2's specific catalytic position in the ERAD mannose-trimming cascade, resolving how it differs from EDEM1 and EDEM3.\",\n      \"evidence\": \"TALEN knockout in human cells with glycan structure analysis and epistasis across EDEM1/2/3 knockouts\",\n      \"pmids\": [\"25092655\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not demonstrate direct enzymatic activity in vitro at this stage\", \"Cofactor requirement for the activity unknown\", \"Did not address non-glycan substrate recognition\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Showed EDEM2 recognizes substrates beyond their glycans and physically links to core ER quality-control machinery, broadening its role from pure mannosidase to substrate-channeling factor.\",\n      \"evidence\": \"siRNA knockdown plus co-IP with non-glycosylated SHH, calnexin, and SEL1L; ricin A-chain retrotranslocation assays\",\n      \"pmids\": [\"24910992\", \"24200403\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular basis of polypeptide recognition not yet defined\", \"Direct vs. indirect interaction with calnexin/SEL1L not resolved\", \"Single lab per substrate\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Identified hydrophobicity of substrate determinants as the basis for glycosylation-independent recognition, explaining how EDEM2 engages misfolded polypeptide regions.\",\n      \"evidence\": \"Hydrophobicity-modulating mutagenesis of ricin and BACE457 with co-IP in human cells\",\n      \"pmids\": [\"25655076\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No structural map of the polypeptide-binding surface\", \"Relationship between hydrophobic binding and mannosidase catalysis unclear\", \"Single lab\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Demonstrated that EDEM2's mannosidase activity is gated by substrate unfolding and enhanced by an oxidoreductase partner, reconciling earlier negative in vitro activity results.\",\n      \"evidence\": \"In vitro mannosidase assays comparing free N-glycans, folded and denatured glycoproteins, plus co-IP identifying TXNDC11\",\n      \"pmids\": [\"30374462\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define the chemical nature of the EDEM2-TXNDC11 link\", \"Mechanism by which unfolding enhances activity not resolved\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Pinpointed the covalent EDEM2-TXNDC11 disulfide as essential for catalysis and reconstituted the first clear in vitro EDEM-family mannosidase activity, settling how EDEM2 becomes enzymatically competent.\",\n      \"evidence\": \"CRISPR KO, site-directed mutagenesis of EDEM2 C558 and TXNDC11 C692, disulfide mapping, complex purification, and in vitro Man9-to-Man8 trimming with MS glycan analysis in HCT116\",\n      \"pmids\": [\"32065582\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No high-resolution structure of the complex\", \"How TXNDC11 redox state modulates activity not defined\", \"Connection between disulfide-driven catalysis and hydrophobic substrate recognition unaddressed\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Mapped the EDEM2 interactome and endogenous substrate landscape, extending its role beyond model substrates to ER quality-control networks and disease-relevant targets.\",\n      \"evidence\": \"Affinity proteomics, sucrose density fractionation, deglycoproteomics, and SILAC pulse proteomics in melanoma cells with biochemical validation\",\n      \"pmids\": [\"34332121\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Substrate candidates (integrin alpha-1, protocadherin 2) inferred from correlation, not direct degradation kinetics\", \"Functional roles of novel interactors not characterized\", \"Single cell-type context\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Revealed a non-canonical EDEM2 function in cardiac lipid homeostasis under XBP1s control, distinguishing its ERAD role from a lipid-regulatory role.\",\n      \"evidence\": \"AAV9 loss/gain-of-function in mice, siRNA in rat and hiPSC cardiomyocytes, luciferase reporter for XBP1s regulation, proteomics, lipidomics, and EM\",\n      \"pmids\": [\"40130322\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanistic link between EDEM2 and SEC23A-mediated ATGL translocation not resolved at molecular level\", \"Whether mannosidase activity is required for the lipid role is untested\", \"Cardiomyocyte-specific; generality unknown\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How EDEM2's two recognition modes — glycan trimming and hydrophobicity-based polypeptide binding — are structurally integrated, and whether the lipid-regulatory function depends on its catalytic activity, remain open.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No crystal/cryo-EM structure of the EDEM2-TXNDC11 complex with substrate\", \"Mechanistic coupling of catalysis to ERAD channeling unresolved\", \"Catalytic dependence of the ATGL/lipid pathway untested\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140098\", \"supporting_discovery_ids\": [1, 5, 6]},\n      {\"term_id\": \"GO:0016787\", \"supporting_discovery_ids\": [1, 6]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005783\", \"supporting_discovery_ids\": [0]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [0, 1, 2, 6]},\n      {\"term_id\": \"R-HSA-8953897\", \"supporting_discovery_ids\": [8]}\n    ],\n    \"complexes\": [\n      \"EDEM2-TXNDC11 disulfide-linked complex\"\n    ],\n    \"partners\": [\n      \"TXNDC11\",\n      \"CANX\",\n      \"SEL1L\",\n      \"SEC23A\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":5,"faith_pct":100.0}}