{"gene":"AP5M1","run_date":"2026-06-09T22:02:43","timeline":{"discoveries":[{"year":2008,"finding":"MUDENG (AP5M1) encodes a 490 amino acid protein containing an adaptin domain homologous to the mu2 subunit of adaptor protein complexes related to clathrin-mediated endocytosis; ectopic overexpression of MUDENG induced cell death in Jurkat T cells and HeLa cells, establishing it as a pro-death protein.","method":"Cloning of full-length ORF, ectopic overexpression in cell lines (Jurkat, HeLa), cell viability assays","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — single lab, cell-based overexpression with defined phenotype, two cell lines tested, but no mechanistic pathway placement beyond domain identification","pmids":["18395520"],"is_preprint":false},{"year":2013,"finding":"MUDENG (AP5M1) is cleaved by caspase-3 at residues D276 and D290 within its adaptin domain during TRAIL-induced apoptosis; in vitro cleavage assay with recombinant active caspase-3 confirmed these cleavage sites, and cleaved MUDENG showed reduced cell-killing activity, indicating that intact adaptin domain integrity is required for MUDENG's pro-death function.","method":"In vitro caspase cleavage assay using recombinant active caspase proteins, site identification by mutagenesis of cleavage sites, cell viability assays in Jurkat and BJAB cells","journal":"Biochemical and biophysical research communications","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstituted caspase cleavage assay with mutagenesis-validated cleavage sites, confirmed in two cell lines; single lab but multiple orthogonal methods","pmids":["23665015"],"is_preprint":false},{"year":2016,"finding":"MUDENG (AP5M1/MuD) functions as an anti-apoptotic protein in human astroglioma cells: MuD protein levels decrease ~33% following TRAIL stimulation (via caspase-3-mediated cleavage), stable overexpression of MuD enhanced cell survival upon TRAIL treatment (77% vs 46%), and MuD depletion increased susceptibility to TRAIL by enhancing cleavage of caspase-3, caspase-9, and Bid. MuD depletion also caused Bcl-2 conversion to a truncated pro-apoptotic 25-kDa fragment, and the TRAIL-sensitizing effect of MuD depletion was abrogated by Bid co-depletion, placing MuD function at the Bid/Bcl-2 junction. MuD localizes predominantly in the endoplasmic reticulum and partly in mitochondria.","method":"Stable transfection (overexpression and shRNA knockdown), TRAIL stimulation, caspase activation assays, Bid and Bcl-2 immunoblotting, subcellular fractionation/localization, genetic epistasis via Bid co-knockdown","journal":"Oncogenesis","confidence":"High","confidence_rationale":"Tier 2 / Moderate — clean KD/KO and OE with defined cellular phenotype, pathway placement via epistasis (Bid co-depletion rescue), subcellular localization with functional context; single lab with multiple orthogonal methods","pmids":["27136675"],"is_preprint":false},{"year":2019,"finding":"AP5M1 (MUDENG) induces apoptosis in cervical cancer cells in a BAX-dependent manner: AP5M1 overexpression upregulated BAX protein levels, and AP5M1 completely lost apoptotic activity in BAX-knockout or BAX-knockdown cervical cancer cells, demonstrating functional dependence on BAX for the mitochondrial apoptotic pathway.","method":"Overexpression in cervical cancer cell lines, BAX knockout and knockdown (siRNA), cell viability/apoptosis assays, mRNA correlation analysis in patient tissues","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — clean BAX-KO and KD with defined cellular rescue phenotype establishing pathway placement; single lab","pmids":["31427081"],"is_preprint":false},{"year":2013,"finding":"A monoclonal antibody (M3H9) against residues 244–326 in the middle domain of human MUDENG (AP5M1) was generated, confirming protein expression in astroglioma cell lines, primary astrocytes, and formalin-fixed mouse ovary and uterus tissues, and validating the middle domain as an antigenic region.","method":"Monoclonal antibody generation, ELISA, immunoblot, immunohistochemistry on formalin-fixed paraffin-embedded tissues","journal":"Monoclonal antibodies in immunodiagnosis and immunotherapy","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, antibody characterization study; confirms protein expression and domain antigenicity but establishes no new mechanism","pmids":["23909422"],"is_preprint":false}],"current_model":"AP5M1 (MUDENG/MuD) is a conserved adaptin-domain-containing protein that localizes predominantly to the endoplasmic reticulum and partly to mitochondria, where it exerts an anti-apoptotic function by acting at the Bid/Bcl-2 junction downstream of TRAIL-induced caspase activation; caspase-3 cleaves AP5M1 at D276 and D290 within its adaptin domain to generate a fragment with reduced pro-death activity, and AP5M1's ability to induce apoptosis in cancer cells is entirely dependent on BAX upregulation."},"narrative":{"mechanistic_narrative":"AP5M1 (MUDENG/MuD) is a 490-amino-acid adaptin-domain-containing protein that regulates the intrinsic apoptotic program, with context-dependent pro-death and anti-apoptotic activities documented across multiple cancer cell types [PMID:18395520, PMID:27136675]. Ectopic overexpression drives cell death in Jurkat and HeLa cells, and in cervical cancer cells this killing operates strictly through the mitochondrial pathway: AP5M1 upregulates BAX and loses all apoptotic activity in BAX-knockout or BAX-knockdown cells [PMID:18395520, PMID:31427081]. During TRAIL-induced apoptosis, AP5M1 is itself a caspase-3 substrate, cleaved at D276 and D290 within its adaptin domain to generate a fragment with reduced cell-killing activity, indicating that an intact adaptin domain is required for its pro-death function [PMID:23665015]. In astroglioma cells AP5M1 acts as an anti-apoptotic factor at the Bid/Bcl-2 junction downstream of TRAIL: its depletion enhances cleavage of caspase-3, caspase-9, and Bid, drives conversion of Bcl-2 to a truncated 25-kDa pro-apoptotic fragment, and this TRAIL-sensitizing effect is abrogated by co-depletion of Bid [PMID:27136675]. AP5M1 localizes predominantly to the endoplasmic reticulum and partly to mitochondria [PMID:27136675]. Beyond its role in apoptotic signaling, no clathrin- or adaptor-complex trafficking function has been characterized for AP5M1 in the available corpus.","teleology":[{"year":2008,"claim":"Established AP5M1/MUDENG as a discrete protein and first linked it to cell death, defining the founding question of how an adaptin-domain protein influences viability.","evidence":"Full-length ORF cloning and ectopic overexpression in Jurkat and HeLa cells with viability assays","pmids":["18395520"],"confidence":"Medium","gaps":["Overexpression phenotype only; no endogenous loss-of-function","No pathway placement beyond domain homology","Trafficking role of the adaptin domain untested"]},{"year":2013,"claim":"Identified AP5M1 as a caspase-3 substrate cleaved at D276/D290, showing that adaptin-domain integrity is required for its pro-death activity and placing the protein downstream of caspase activation.","evidence":"In vitro recombinant caspase-3 cleavage assay with mutagenesis-validated sites, confirmed in Jurkat and BJAB cells under TRAIL","pmids":["23665015"],"confidence":"High","gaps":["Functional consequence of the cleavage fragment in vivo not resolved","Does not explain how intact protein engages the death machinery"]},{"year":2013,"claim":"Provided reagent and expression validation across cell lines and tissues, confirming endogenous protein presence and the middle domain as antigenic.","evidence":"Monoclonal antibody (M3H9) generation, ELISA, immunoblot, and IHC on astroglioma, astrocytes, and mouse ovary/uterus","pmids":["23909422"],"confidence":"Low","gaps":["Antibody characterization only; establishes no mechanism","Physiological role in the reproductive tissues not addressed"]},{"year":2016,"claim":"Reframed AP5M1 as anti-apoptotic in astroglioma and placed its function at the Bid/Bcl-2 junction, resolving where in the TRAIL pathway it acts.","evidence":"Stable overexpression and shRNA knockdown with TRAIL stimulation, caspase/Bid/Bcl-2 immunoblotting, Bid co-depletion epistasis, and subcellular fractionation","pmids":["27136675"],"confidence":"High","gaps":["Direct binding partners at the Bid/Bcl-2 node not identified","Mechanism reconciling pro-death overexpression and anti-apoptotic depletion phenotypes unresolved","Molecular basis of ER/mitochondrial localization unknown"]},{"year":2019,"claim":"Demonstrated that AP5M1's apoptotic activity in cervical cancer is entirely BAX-dependent, anchoring it to the mitochondrial apoptotic pathway.","evidence":"Overexpression with BAX knockout and siRNA knockdown, apoptosis assays, and patient-tissue mRNA correlation","pmids":["31427081"],"confidence":"Medium","gaps":["Whether AP5M1 acts directly or indirectly on BAX not determined","Reconciliation with anti-apoptotic role in astroglioma not addressed"]},{"year":null,"claim":"Whether AP5M1 retains any clathrin- or adaptor-complex trafficking function predicted by its mu2-homologous adaptin domain, and how this relates to its apoptotic role, remains unresolved.","evidence":"No trafficking or vesicular function assayed in the available corpus","pmids":[],"confidence":"Low","gaps":["No identified membrane cargo or coat partners","No structural model of the adaptin domain in context","Direct molecular partners at apoptotic nodes unidentified"]}],"mechanism_profile":{"molecular_activity":[],"localization":[{"term_id":"GO:0005783","term_label":"endoplasmic reticulum","supporting_discovery_ids":[2]},{"term_id":"GO:0005739","term_label":"mitochondrion","supporting_discovery_ids":[2]}],"pathway":[{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[1,2,3]}],"complexes":[],"partners":[],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9H0R1","full_name":"AP-5 complex subunit mu-1","aliases":["Adaptor-related protein complex 5 subunit mu-1","Mu5","Mu-2-related death-inducing protein","MuD","Putative HIV-1 infection-related protein"],"length_aa":490,"mass_kda":54.8,"function":"As part of AP-5, a probable fifth adaptor protein complex it may be involved in endosomal transport. According to PubMed:18395520, it may play a role in cell death","subcellular_location":"Cytoplasm, cytosol; Late endosome membrane; Lysosome membrane","url":"https://www.uniprot.org/uniprotkb/Q9H0R1/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/AP5M1","classification":"Not Classified","n_dependent_lines":1,"n_total_lines":1208,"dependency_fraction":0.0008278145695364238},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/AP5M1","total_profiled":1310},"omim":[{"mim_id":"614368","title":"ADAPTOR-RELATED PROTEIN COMPLEX 5, MU-1 SUBUNIT; AP5M1","url":"https://www.omim.org/entry/614368"},{"mim_id":"613653","title":"ADAPTOR-RELATED PROTEIN COMPLEX 5, ZETA-1 SUBUNIT; AP5Z1","url":"https://www.omim.org/entry/613653"},{"mim_id":"613647","title":"SPASTIC PARAPLEGIA 48, AUTOSOMAL RECESSIVE; SPG48","url":"https://www.omim.org/entry/613647"}],"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/AP5M1"},"hgnc":{"alias_symbol":["FLJ10813","MuD","mu5"],"prev_symbol":["C14orf108","MUDENG"]},"alphafold":{"accession":"Q9H0R1","domains":[{"cath_id":"3.30.450.60","chopping":"2-181","consensus_level":"high","plddt":87.4464,"start":2,"end":181},{"cath_id":"2.60.40.1170","chopping":"207-318_441-472","consensus_level":"medium","plddt":87.3742,"start":207,"end":472},{"cath_id":"2.60.40.1170","chopping":"322-437","consensus_level":"medium","plddt":92.1948,"start":322,"end":437}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9H0R1","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9H0R1-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9H0R1-F1-predicted_aligned_error_v6.png","plddt_mean":86.94},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=AP5M1","jax_strain_url":"https://www.jax.org/strain/search?query=AP5M1"},"sequence":{"accession":"Q9H0R1","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9H0R1.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9H0R1/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9H0R1"}},"corpus_meta":[{"pmid":"16648843","id":"PMC_16648843","title":"The 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\"Cloning of full-length ORF, ectopic overexpression in cell lines (Jurkat, HeLa), cell viability assays\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — single lab, cell-based overexpression with defined phenotype, two cell lines tested, but no mechanistic pathway placement beyond domain identification\",\n      \"pmids\": [\"18395520\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"MUDENG (AP5M1) is cleaved by caspase-3 at residues D276 and D290 within its adaptin domain during TRAIL-induced apoptosis; in vitro cleavage assay with recombinant active caspase-3 confirmed these cleavage sites, and cleaved MUDENG showed reduced cell-killing activity, indicating that intact adaptin domain integrity is required for MUDENG's pro-death function.\",\n      \"method\": \"In vitro caspase cleavage assay using recombinant active caspase proteins, site identification by mutagenesis of cleavage sites, cell viability assays in Jurkat and BJAB cells\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstituted caspase cleavage assay with mutagenesis-validated cleavage sites, confirmed in two cell lines; single lab but multiple orthogonal methods\",\n      \"pmids\": [\"23665015\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"MUDENG (AP5M1/MuD) functions as an anti-apoptotic protein in human astroglioma cells: MuD protein levels decrease ~33% following TRAIL stimulation (via caspase-3-mediated cleavage), stable overexpression of MuD enhanced cell survival upon TRAIL treatment (77% vs 46%), and MuD depletion increased susceptibility to TRAIL by enhancing cleavage of caspase-3, caspase-9, and Bid. MuD depletion also caused Bcl-2 conversion to a truncated pro-apoptotic 25-kDa fragment, and the TRAIL-sensitizing effect of MuD depletion was abrogated by Bid co-depletion, placing MuD function at the Bid/Bcl-2 junction. MuD localizes predominantly in the endoplasmic reticulum and partly in mitochondria.\",\n      \"method\": \"Stable transfection (overexpression and shRNA knockdown), TRAIL stimulation, caspase activation assays, Bid and Bcl-2 immunoblotting, subcellular fractionation/localization, genetic epistasis via Bid co-knockdown\",\n      \"journal\": \"Oncogenesis\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — clean KD/KO and OE with defined cellular phenotype, pathway placement via epistasis (Bid co-depletion rescue), subcellular localization with functional context; single lab with multiple orthogonal methods\",\n      \"pmids\": [\"27136675\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"AP5M1 (MUDENG) induces apoptosis in cervical cancer cells in a BAX-dependent manner: AP5M1 overexpression upregulated BAX protein levels, and AP5M1 completely lost apoptotic activity in BAX-knockout or BAX-knockdown cervical cancer cells, demonstrating functional dependence on BAX for the mitochondrial apoptotic pathway.\",\n      \"method\": \"Overexpression in cervical cancer cell lines, BAX knockout and knockdown (siRNA), cell viability/apoptosis assays, mRNA correlation analysis in patient tissues\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — clean BAX-KO and KD with defined cellular rescue phenotype establishing pathway placement; single lab\",\n      \"pmids\": [\"31427081\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"A monoclonal antibody (M3H9) against residues 244–326 in the middle domain of human MUDENG (AP5M1) was generated, confirming protein expression in astroglioma cell lines, primary astrocytes, and formalin-fixed mouse ovary and uterus tissues, and validating the middle domain as an antigenic region.\",\n      \"method\": \"Monoclonal antibody generation, ELISA, immunoblot, immunohistochemistry on formalin-fixed paraffin-embedded tissues\",\n      \"journal\": \"Monoclonal antibodies in immunodiagnosis and immunotherapy\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, antibody characterization study; confirms protein expression and domain antigenicity but establishes no new mechanism\",\n      \"pmids\": [\"23909422\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"AP5M1 (MUDENG/MuD) is a conserved adaptin-domain-containing protein that localizes predominantly to the endoplasmic reticulum and partly to mitochondria, where it exerts an anti-apoptotic function by acting at the Bid/Bcl-2 junction downstream of TRAIL-induced caspase activation; caspase-3 cleaves AP5M1 at D276 and D290 within its adaptin domain to generate a fragment with reduced pro-death activity, and AP5M1's ability to induce apoptosis in cancer cells is entirely dependent on BAX upregulation.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"AP5M1 (MUDENG/MuD) is a 490-amino-acid adaptin-domain-containing protein that regulates the intrinsic apoptotic program, with context-dependent pro-death and anti-apoptotic activities documented across multiple cancer cell types [#0, #2]. Ectopic overexpression drives cell death in Jurkat and HeLa cells, and in cervical cancer cells this killing operates strictly through the mitochondrial pathway: AP5M1 upregulates BAX and loses all apoptotic activity in BAX-knockout or BAX-knockdown cells [#0, #3]. During TRAIL-induced apoptosis, AP5M1 is itself a caspase-3 substrate, cleaved at D276 and D290 within its adaptin domain to generate a fragment with reduced cell-killing activity, indicating that an intact adaptin domain is required for its pro-death function [#1]. In astroglioma cells AP5M1 acts as an anti-apoptotic factor at the Bid/Bcl-2 junction downstream of TRAIL: its depletion enhances cleavage of caspase-3, caspase-9, and Bid, drives conversion of Bcl-2 to a truncated 25-kDa pro-apoptotic fragment, and this TRAIL-sensitizing effect is abrogated by co-depletion of Bid [#2]. AP5M1 localizes predominantly to the endoplasmic reticulum and partly to mitochondria [#2]. Beyond its role in apoptotic signaling, no clathrin- or adaptor-complex trafficking function has been characterized for AP5M1 in the available corpus.\",\n  \"teleology\": [\n    {\n      \"year\": 2008,\n      \"claim\": \"Established AP5M1/MUDENG as a discrete protein and first linked it to cell death, defining the founding question of how an adaptin-domain protein influences viability.\",\n      \"evidence\": \"Full-length ORF cloning and ectopic overexpression in Jurkat and HeLa cells with viability assays\",\n      \"pmids\": [\"18395520\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Overexpression phenotype only; no endogenous loss-of-function\", \"No pathway placement beyond domain homology\", \"Trafficking role of the adaptin domain untested\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Identified AP5M1 as a caspase-3 substrate cleaved at D276/D290, showing that adaptin-domain integrity is required for its pro-death activity and placing the protein downstream of caspase activation.\",\n      \"evidence\": \"In vitro recombinant caspase-3 cleavage assay with mutagenesis-validated sites, confirmed in Jurkat and BJAB cells under TRAIL\",\n      \"pmids\": [\"23665015\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Functional consequence of the cleavage fragment in vivo not resolved\", \"Does not explain how intact protein engages the death machinery\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Provided reagent and expression validation across cell lines and tissues, confirming endogenous protein presence and the middle domain as antigenic.\",\n      \"evidence\": \"Monoclonal antibody (M3H9) generation, ELISA, immunoblot, and IHC on astroglioma, astrocytes, and mouse ovary/uterus\",\n      \"pmids\": [\"23909422\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Antibody characterization only; establishes no mechanism\", \"Physiological role in the reproductive tissues not addressed\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Reframed AP5M1 as anti-apoptotic in astroglioma and placed its function at the Bid/Bcl-2 junction, resolving where in the TRAIL pathway it acts.\",\n      \"evidence\": \"Stable overexpression and shRNA knockdown with TRAIL stimulation, caspase/Bid/Bcl-2 immunoblotting, Bid co-depletion epistasis, and subcellular fractionation\",\n      \"pmids\": [\"27136675\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct binding partners at the Bid/Bcl-2 node not identified\", \"Mechanism reconciling pro-death overexpression and anti-apoptotic depletion phenotypes unresolved\", \"Molecular basis of ER/mitochondrial localization unknown\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Demonstrated that AP5M1's apoptotic activity in cervical cancer is entirely BAX-dependent, anchoring it to the mitochondrial apoptotic pathway.\",\n      \"evidence\": \"Overexpression with BAX knockout and siRNA knockdown, apoptosis assays, and patient-tissue mRNA correlation\",\n      \"pmids\": [\"31427081\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether AP5M1 acts directly or indirectly on BAX not determined\", \"Reconciliation with anti-apoptotic role in astroglioma not addressed\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"Whether AP5M1 retains any clathrin- or adaptor-complex trafficking function predicted by its mu2-homologous adaptin domain, and how this relates to its apoptotic role, remains unresolved.\",\n      \"evidence\": \"No trafficking or vesicular function assayed in the available corpus\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No identified membrane cargo or coat partners\", \"No structural model of the adaptin domain in context\", \"Direct molecular partners at apoptotic nodes unidentified\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [],\n    \"localization\": [\n      {\"term_id\": \"GO:0005783\", \"supporting_discovery_ids\": [2]},\n      {\"term_id\": \"GO:0005739\", \"supporting_discovery_ids\": [2]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [1, 2, 3]}\n    ],\n    \"complexes\": [],\n    \"partners\": [],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":5,"faith_pct":100.0}}