{"gene":"NECAP2","run_date":"2026-06-10T05:19:52","timeline":{"discoveries":[{"year":2016,"finding":"NECAP2 regulates fast endocytic recycling of EGFR and transferrin receptor by controlling clathrin coat (AP-1 adapter) recruitment to early endosomes; knockdown of NECAP2 causes enlarged early endosomes and loss of AP-1 from the organelle. Structure-function analysis defined the protein-binding interfaces in NECAP2 required for AP-1 recruitment. NECAP2 acts specifically in the fast recycling pathway (direct early endosome-to-plasma membrane route) and does not regulate clathrin-mediated endocytosis, EGFR degradation, or slow Rab11-dependent recycling.","method":"siRNA knockdown, recycling assays (EGFR and transferrin receptor), immunofluorescence/endosome morphology analysis, structure-function mutagenesis of NECAP2 binding interfaces","journal":"Journal of cell science","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal pathway dissection with multiple cargo assays, structure-function mutagenesis, and defined organelle phenotype in a single focused study","pmids":["27206861"],"is_preprint":false},{"year":2004,"finding":"NECAP2 (along with NECAP1) was identified as a component of clathrin-coated vesicles (CCVs) isolated from adult brain, and characterization revealed a new consensus motif in NECAP proteins that mediates interactions with the clathrin adaptor protein AP-2.","method":"Subcellular fractionation of brain CCVs, mass spectrometry proteomics, identification of AP-2 interaction motif","journal":"Biochemical Society transactions","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — subcellular proteomics plus motif characterization, but abstract-level detail limits full assessment of biochemical validation","pmids":["15494011"],"is_preprint":false},{"year":2013,"finding":"In an siRNA screen of 172 endocytosis and actin-regulatory genes, NECAP2 knockdown was identified as impairing EGFR endocytosis in human HBL100 breast cancer cells, placing NECAP2 among genes required for EGFR endocytic trafficking.","method":"High-content fluorescence microscopy-based siRNA screen for EGFR endocytosis","journal":"Oncogene","confidence":"Low","confidence_rationale":"Tier 3 / Weak — identified in a large screen with no mechanistic follow-up specific to NECAP2; single method, no pathway placement beyond the screen hit","pmids":["23792445"],"is_preprint":false}],"current_model":"NECAP2 is a component of clathrin-coated vesicles that recruits the AP-1 clathrin adaptor to early endosomes via defined protein-binding interfaces, specifically driving the fast (direct early endosome-to-plasma membrane) recycling pathway for cargos such as EGFR and transferrin receptor, without affecting clathrin-mediated endocytosis or slow Rab11-dependent recycling; it also harbors a consensus motif mediating interaction with the AP-2 clathrin adaptor at the plasma membrane."},"narrative":{"mechanistic_narrative":"NECAP2 is a component of clathrin-coated vesicles that controls clathrin adaptor recruitment to drive fast endocytic recycling [PMID:27206861, PMID:15494011]. It was originally identified by proteomics of brain-derived clathrin-coated vesicles, where a consensus motif in NECAP proteins was found to mediate interaction with the AP-2 clathrin adaptor [PMID:15494011]. Functionally, NECAP2 governs the fast, direct early endosome-to-plasma membrane recycling route for cargos including EGFR and transferrin receptor by recruiting the AP-1 clathrin adaptor to early endosomes; its loss removes AP-1 from the organelle and produces enlarged early endosomes, with defined protein-binding interfaces in NECAP2 required for this recruitment [PMID:27206861]. This activity is pathway-selective: NECAP2 does not regulate clathrin-mediated endocytosis, EGFR degradation, or slow Rab11-dependent recycling [PMID:27206861]. Beyond these clathrin-adaptor interactions and the recycling phenotype, no further mechanistic detail has been characterized in the available corpus.","teleology":[{"year":2004,"claim":"Establishing that NECAP2 is a bona fide constituent of clathrin-coated vesicles and engages the AP-2 adaptor placed it within the clathrin trafficking machinery and gave a molecular handle on its mechanism.","evidence":"Subcellular fractionation of brain clathrin-coated vesicles with mass spectrometry and identification of an AP-2 interaction motif","pmids":["15494011"],"confidence":"Medium","gaps":["Functional consequence of the AP-2 interaction was not assayed","No cargo or pathway role was defined at this stage","Abstract-level biochemical validation only"]},{"year":2013,"claim":"An unbiased functional screen tied NECAP2 to receptor endocytic trafficking, implicating it in EGFR handling before any dedicated mechanism was known.","evidence":"High-content microscopy-based siRNA screen of 172 endocytosis/actin genes scoring EGFR endocytosis in HBL100 cells","pmids":["23792445"],"confidence":"Low","gaps":["Single screen hit with no NECAP2-specific mechanistic follow-up","Did not distinguish endocytosis from recycling steps","No interaction partners or interfaces tested"]},{"year":2016,"claim":"Defining NECAP2 as a selective driver of fast endosome-to-plasma membrane recycling via AP-1 recruitment resolved which trafficking step it controls and which adaptor interface it uses.","evidence":"siRNA knockdown with EGFR and transferrin receptor recycling assays, endosome morphology imaging, and structure-function mutagenesis of NECAP2 binding interfaces","pmids":["27206861"],"confidence":"High","gaps":["Structural basis of the NECAP2–AP-1 interface not solved atomically","How NECAP2 is itself targeted to early endosomes is unresolved","Relationship between the AP-2 motif role and AP-1-dependent recycling not reconciled"]},{"year":null,"claim":"How NECAP2 is recruited to early endosomes and the structural mechanism by which it engages AP-1 versus AP-2 remains unresolved.","evidence":"","pmids":[],"confidence":"High","gaps":["No structure of NECAP2 in complex with AP-1 or AP-2","Upstream regulators controlling NECAP2 endosomal targeting unknown","Physiological consequences in tissue or organismal context uncharacterized"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[0,1]}],"localization":[{"term_id":"GO:0031410","term_label":"cytoplasmic vesicle","supporting_discovery_ids":[1]},{"term_id":"GO:0005768","term_label":"endosome","supporting_discovery_ids":[0]}],"pathway":[{"term_id":"R-HSA-5653656","term_label":"Vesicle-mediated transport","supporting_discovery_ids":[0,1]}],"complexes":["clathrin-coated vesicle"],"partners":["AP1","AP2"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9NVZ3","full_name":"Adaptin ear-binding coat-associated protein 2","aliases":["NECAP endocytosis-associated protein 2","NECAP-2"],"length_aa":263,"mass_kda":28.3,"function":"Involved in endocytosis","subcellular_location":"Cytoplasmic vesicle, clathrin-coated vesicle membrane; Cell membrane","url":"https://www.uniprot.org/uniprotkb/Q9NVZ3/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/NECAP2","classification":"Not Classified","n_dependent_lines":3,"n_total_lines":1208,"dependency_fraction":0.0024834437086092716},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000157191","cell_line_id":"CID000539","localizations":[{"compartment":"cytoplasmic","grade":3},{"compartment":"nucleoplasm","grade":3},{"compartment":"membrane","grade":2},{"compartment":"cell_contact","grade":1}],"interactors":[{"gene":"AP2M1","stoichiometry":4.0},{"gene":"AP2S1","stoichiometry":4.0},{"gene":"AP2A2","stoichiometry":0.2},{"gene":"AP2A1","stoichiometry":0.2},{"gene":"EPS15L1","stoichiometry":0.2},{"gene":"SMG1","stoichiometry":0.2},{"gene":"EPS15","stoichiometry":0.2},{"gene":"GLOD4","stoichiometry":0.2},{"gene":"BMP2K","stoichiometry":0.2},{"gene":"AP1B1","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/target/CID000539","total_profiled":1310},"omim":[{"mim_id":"611624","title":"NECAP ENDOCYTOSIS-ASSOCIATED PROTEIN 2; NECAP2","url":"https://www.omim.org/entry/611624"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Endoplasmic reticulum","reliability":"Approved"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/NECAP2"},"hgnc":{"alias_symbol":["FLJ10420"],"prev_symbol":[]},"alphafold":{"accession":"Q9NVZ3","domains":[{"cath_id":"2.30.29.30","chopping":"8-141","consensus_level":"high","plddt":88.115,"start":8,"end":141}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9NVZ3","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9NVZ3-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9NVZ3-F1-predicted_aligned_error_v6.png","plddt_mean":69.31},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=NECAP2","jax_strain_url":"https://www.jax.org/strain/search?query=NECAP2"},"sequence":{"accession":"Q9NVZ3","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9NVZ3.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9NVZ3/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9NVZ3"}},"corpus_meta":[{"pmid":"23792445","id":"PMC_23792445","title":"Annexin A2 depletion delays EGFR endocytic trafficking via cofilin activation and enhances EGFR signaling and metastasis formation.","date":"2013","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/23792445","citation_count":49,"is_preprint":false},{"pmid":"36639822","id":"PMC_36639822","title":"CDCA8 induced by NF-YA promotes hepatocellular carcinoma progression by regulating the MEK/ERK pathway.","date":"2023","source":"Experimental hematology & oncology","url":"https://pubmed.ncbi.nlm.nih.gov/36639822","citation_count":30,"is_preprint":false},{"pmid":"27206861","id":"PMC_27206861","title":"NECAP2 controls clathrin coat recruitment to early endosomes for fast endocytic recycling.","date":"2016","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/27206861","citation_count":22,"is_preprint":false},{"pmid":"15494011","id":"PMC_15494011","title":"Molecular mechanisms in clathrin-mediated membrane budding revealed through subcellular proteomics.","date":"2004","source":"Biochemical Society transactions","url":"https://pubmed.ncbi.nlm.nih.gov/15494011","citation_count":19,"is_preprint":false},{"pmid":"30523707","id":"PMC_30523707","title":"Integrated comparison of the miRNAome and mRNAome in muscles of dermatomyositis and polymyositis reveals common and specific miRNA-mRNAs.","date":"2018","source":"Epigenomics","url":"https://pubmed.ncbi.nlm.nih.gov/30523707","citation_count":13,"is_preprint":false},{"pmid":"30537176","id":"PMC_30537176","title":"Functional analysis of DNA methylation of the PACSIN1 promoter in pig peripheral blood mononuclear cells.","date":"2018","source":"Journal of cellular biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/30537176","citation_count":1,"is_preprint":false},{"pmid":"42188696","id":"PMC_42188696","title":"YAP1 Upregulates Cytoskeleton Regulator ARHGEF1 and Tissue Regeneration Factor NEDD9 in a Multiplex Proteomic Study.","date":"2026","source":"Neurology international","url":"https://pubmed.ncbi.nlm.nih.gov/42188696","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":5867,"output_tokens":885,"usd":0.015438,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":7348,"output_tokens":1701,"usd":0.039633,"stage2_stop_reason":"end_turn"},"total_usd":0.055071,"stage1_batch_id":"msgbatch_013a8chtv3AWyTqv4z4sEF4C","stage2_batch_id":"msgbatch_01AHsuW63vinsvuTgzmQ3n62","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2016,\n      \"finding\": \"NECAP2 regulates fast endocytic recycling of EGFR and transferrin receptor by controlling clathrin coat (AP-1 adapter) recruitment to early endosomes; knockdown of NECAP2 causes enlarged early endosomes and loss of AP-1 from the organelle. Structure-function analysis defined the protein-binding interfaces in NECAP2 required for AP-1 recruitment. NECAP2 acts specifically in the fast recycling pathway (direct early endosome-to-plasma membrane route) and does not regulate clathrin-mediated endocytosis, EGFR degradation, or slow Rab11-dependent recycling.\",\n      \"method\": \"siRNA knockdown, recycling assays (EGFR and transferrin receptor), immunofluorescence/endosome morphology analysis, structure-function mutagenesis of NECAP2 binding interfaces\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal pathway dissection with multiple cargo assays, structure-function mutagenesis, and defined organelle phenotype in a single focused study\",\n      \"pmids\": [\"27206861\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"NECAP2 (along with NECAP1) was identified as a component of clathrin-coated vesicles (CCVs) isolated from adult brain, and characterization revealed a new consensus motif in NECAP proteins that mediates interactions with the clathrin adaptor protein AP-2.\",\n      \"method\": \"Subcellular fractionation of brain CCVs, mass spectrometry proteomics, identification of AP-2 interaction motif\",\n      \"journal\": \"Biochemical Society transactions\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — subcellular proteomics plus motif characterization, but abstract-level detail limits full assessment of biochemical validation\",\n      \"pmids\": [\"15494011\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"In an siRNA screen of 172 endocytosis and actin-regulatory genes, NECAP2 knockdown was identified as impairing EGFR endocytosis in human HBL100 breast cancer cells, placing NECAP2 among genes required for EGFR endocytic trafficking.\",\n      \"method\": \"High-content fluorescence microscopy-based siRNA screen for EGFR endocytosis\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — identified in a large screen with no mechanistic follow-up specific to NECAP2; single method, no pathway placement beyond the screen hit\",\n      \"pmids\": [\"23792445\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"NECAP2 is a component of clathrin-coated vesicles that recruits the AP-1 clathrin adaptor to early endosomes via defined protein-binding interfaces, specifically driving the fast (direct early endosome-to-plasma membrane) recycling pathway for cargos such as EGFR and transferrin receptor, without affecting clathrin-mediated endocytosis or slow Rab11-dependent recycling; it also harbors a consensus motif mediating interaction with the AP-2 clathrin adaptor at the plasma membrane.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"NECAP2 is a component of clathrin-coated vesicles that controls clathrin adaptor recruitment to drive fast endocytic recycling [#0, #1]. It was originally identified by proteomics of brain-derived clathrin-coated vesicles, where a consensus motif in NECAP proteins was found to mediate interaction with the AP-2 clathrin adaptor [#1]. Functionally, NECAP2 governs the fast, direct early endosome-to-plasma membrane recycling route for cargos including EGFR and transferrin receptor by recruiting the AP-1 clathrin adaptor to early endosomes; its loss removes AP-1 from the organelle and produces enlarged early endosomes, with defined protein-binding interfaces in NECAP2 required for this recruitment [#0]. This activity is pathway-selective: NECAP2 does not regulate clathrin-mediated endocytosis, EGFR degradation, or slow Rab11-dependent recycling [#0]. Beyond these clathrin-adaptor interactions and the recycling phenotype, no further mechanistic detail has been characterized in the available corpus.\",\n  \"teleology\": [\n    {\n      \"year\": 2004,\n      \"claim\": \"Establishing that NECAP2 is a bona fide constituent of clathrin-coated vesicles and engages the AP-2 adaptor placed it within the clathrin trafficking machinery and gave a molecular handle on its mechanism.\",\n      \"evidence\": \"Subcellular fractionation of brain clathrin-coated vesicles with mass spectrometry and identification of an AP-2 interaction motif\",\n      \"pmids\": [\n        \"15494011\"\n      ],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Functional consequence of the AP-2 interaction was not assayed\",\n        \"No cargo or pathway role was defined at this stage\",\n        \"Abstract-level biochemical validation only\"\n      ]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"An unbiased functional screen tied NECAP2 to receptor endocytic trafficking, implicating it in EGFR handling before any dedicated mechanism was known.\",\n      \"evidence\": \"High-content microscopy-based siRNA screen of 172 endocytosis/actin genes scoring EGFR endocytosis in HBL100 cells\",\n      \"pmids\": [\n        \"23792445\"\n      ],\n      \"confidence\": \"Low\",\n      \"gaps\": [\n        \"Single screen hit with no NECAP2-specific mechanistic follow-up\",\n        \"Did not distinguish endocytosis from recycling steps\",\n        \"No interaction partners or interfaces tested\"\n      ]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Defining NECAP2 as a selective driver of fast endosome-to-plasma membrane recycling via AP-1 recruitment resolved which trafficking step it controls and which adaptor interface it uses.\",\n      \"evidence\": \"siRNA knockdown with EGFR and transferrin receptor recycling assays, endosome morphology imaging, and structure-function mutagenesis of NECAP2 binding interfaces\",\n      \"pmids\": [\n        \"27206861\"\n      ],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Structural basis of the NECAP2–AP-1 interface not solved atomically\",\n        \"How NECAP2 is itself targeted to early endosomes is unresolved\",\n        \"Relationship between the AP-2 motif role and AP-1-dependent recycling not reconciled\"\n      ]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How NECAP2 is recruited to early endosomes and the structural mechanism by which it engages AP-1 versus AP-2 remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"No structure of NECAP2 in complex with AP-1 or AP-2\",\n        \"Upstream regulators controlling NECAP2 endosomal targeting unknown\",\n        \"Physiological consequences in tissue or organismal context uncharacterized\"\n      ]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\n        \"term_id\": \"GO:0060090\",\n        \"supporting_discovery_ids\": [\n          0,\n          1\n        ]\n      }\n    ],\n    \"localization\": [\n      {\n        \"term_id\": \"GO:0031410\",\n        \"supporting_discovery_ids\": [\n          1\n        ]\n      },\n      {\n        \"term_id\": \"GO:0005768\",\n        \"supporting_discovery_ids\": [\n          0\n        ]\n      }\n    ],\n    \"pathway\": [\n      {\n        \"term_id\": \"R-HSA-5653656\",\n        \"supporting_discovery_ids\": [\n          0,\n          1\n        ]\n      }\n    ],\n    \"complexes\": [\n      \"clathrin-coated vesicle\"\n    ],\n    \"partners\": [\n      \"AP1\",\n      \"AP2\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"faith_supported":4,"faith_total":4,"faith_pct":100.0}}