{"gene":"ACAP2","run_date":"2026-04-28T17:12:37","timeline":{"discoveries":[{"year":2012,"finding":"ACAP2 (centaurin-β2) functions as a Rab35 effector and as an Arf6-GTPase-activating protein (GAP) during neurite outgrowth. Rab35 recruits ACAP2 to Arf6-positive endosomes in response to NGF stimulation, and ACAP2's Arf6-GAP activity at these endosomes is required for neurite outgrowth in PC12 cells.","method":"Knockdown and rescue experiments, live-cell imaging, co-localization, GTPase activity assays","journal":"Journal of cell science","confidence":"High","confidence_rationale":"Tier 2 — reciprocal functional assays, knockdown/rescue, replicated across multiple cellular contexts","pmids":["22344257"],"is_preprint":false},{"year":2011,"finding":"Rab35 recruits ACAP2 (an ARF6-GAP) to phagocytic cups during FcγR-mediated phagocytosis in macrophages; GTP-Rab35-dependent recruitment of ACAP2 regulates actin-dependent phagosome formation, and overexpression of ACAP2 together with GTP-locked Rab35 synergistically inhibits phagocytosis.","method":"Live-cell imaging, RNAi knockdown, expression of GDP/GTP-locked Rab35 mutants, co-expression assays","journal":"Journal of cell science","confidence":"High","confidence_rationale":"Tier 2 — clean KO/KD with defined cellular phenotype, multiple mutant constructs, functional epistasis","pmids":["22045739"],"is_preprint":false},{"year":2014,"finding":"Rab35 and its effector ACAP2 negatively regulate oligodendrocyte morphological differentiation and myelination by inactivating (switching off) Arf6; knockdown of Rab35 or ACAP2 promotes differentiation and myelination, while Arf6 knockdown inhibits it, placing Rab35/ACAP2 upstream of Arf6 in this pathway.","method":"siRNA knockdown, oligodendrocyte-neuronal co-culture, GTPase activity assays, pharmacological inhibition of cytohesin-2","journal":"Molecular biology of the cell","confidence":"High","confidence_rationale":"Tier 2 — genetic epistasis with multiple knockdowns and pharmacological validation","pmids":["24600047"],"is_preprint":false},{"year":2015,"finding":"Two threonine residues (Thr-76 and Thr-81) in the switch II region of Rab35 are required for binding ACAP2, and two asparagine residues (Asn-610 and Asn-691) in ACAP2 are key for specific Rab35 recognition; binding-deficient mutants of either protein fail to support neurite outgrowth in PC12 cells.","method":"Deletion and point mutation analyses, co-immunoprecipitation, knockdown-rescue experiments","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1-2 — mutagenesis with functional validation and rescue experiments","pmids":["25694427"],"is_preprint":false},{"year":2015,"finding":"Human ACAP2 is a functional homolog of C. elegans CNT-1; ACAP2 has a pro-apoptotic function and shares an identical phosphoinositide-binding pattern with truncated CNT-1 (tCNT-1). Knockdown of ACAP2 blocks apoptosis in cancer cells in response to 5-fluorouracil.","method":"siRNA knockdown, apoptosis assay, phosphoinositide-binding assay","journal":"Cell cycle","confidence":"Medium","confidence_rationale":"Tier 2-3 — single lab, knockdown with defined phenotype plus binding assay","pmids":["25853217"],"is_preprint":false},{"year":2006,"finding":"Vaccinia virus K1L protein binds ACAP2 (a GAP for ARF6); however, mutational analysis showed that residues required for VV replication in human or rabbit cells are distinct from the ACAP2-binding site on K1L, indicating K1L's host-range function is independent of its ACAP2 interaction.","method":"Mutagenesis, co-immunoprecipitation/binding assays, viral replication assays","journal":"Virology","confidence":"Medium","confidence_rationale":"Tier 2 — mutagenesis plus binding assay in single study; identifies ACAP2 as a K1L-binding partner","pmids":["16806385"],"is_preprint":false},{"year":2025,"finding":"RNF126 (a ubiquitin E3 ligase) ubiquitinates and promotes degradation of ACAP2 protein in ovarian cancer cells; RNF126-mediated ACAP2 degradation reprograms lipid metabolism to promote ovarian cancer proliferation, migration, and metastasis.","method":"Co-immunoprecipitation, cycloheximide chase assay, siRNA knockdown, in vivo xenograft model","journal":"Biochemical genetics","confidence":"Medium","confidence_rationale":"Tier 2 — Co-IP plus protein stability assay in single study identifying RNF126 as ACAP2 ubiquitin ligase","pmids":["40251363"],"is_preprint":false}],"current_model":"ACAP2 (centaurin-β2) is an Arf6 GTPase-activating protein (GAP) that is recruited to specific membrane compartments (Arf6-positive endosomes, phagocytic cups) by acting as an effector of GTP-bound Rab35 — via specific residues in Rab35's switch II region and ACAP2's C-terminal domain — to inactivate Arf6 and thereby regulate neurite outgrowth, phagocytosis, and oligodendrocyte differentiation/myelination; additionally, ACAP2 has a pro-apoptotic phosphoinositide-binding function and is subject to ubiquitin-mediated proteasomal degradation by the E3 ligase RNF126."},"narrative":{"teleology":[{"year":2006,"claim":"ACAP2 was identified as a physical interactor of vaccinia virus K1L protein, establishing it as a GAP for ARF6 accessible to viral manipulation, though K1L's host-range function proved independent of this binding.","evidence":"Mutagenesis and co-immunoprecipitation with vaccinia K1L in cell culture","pmids":["16806385"],"confidence":"Medium","gaps":["Only a single study; no independent confirmation of the K1L–ACAP2 interaction","Biological significance of the K1L–ACAP2 interaction remains undefined","No endogenous cellular phenotype linked to this interaction"]},{"year":2011,"claim":"The upstream activator Rab35 was shown to recruit ACAP2 to phagocytic cups during Fcγ receptor–mediated phagocytosis, establishing the Rab35→ACAP2→Arf6 axis as a regulatory module controlling actin-dependent phagosome formation in macrophages.","evidence":"Live-cell imaging, RNAi knockdown, and expression of GDP/GTP-locked Rab35 mutants in macrophages","pmids":["22045739"],"confidence":"High","gaps":["The precise step at which Arf6 inactivation blocks phagosome closure was not resolved","Whether other Rab35 effectors contribute in parallel was not tested"]},{"year":2012,"claim":"The same Rab35→ACAP2→Arf6 cascade was extended to neuronal differentiation, showing that NGF-stimulated Rab35 recruits ACAP2 to Arf6-positive endosomes and that ACAP2 GAP activity is required for neurite outgrowth.","evidence":"Knockdown-rescue, live-cell imaging, and GTPase activity assays in PC12 cells","pmids":["22344257"],"confidence":"High","gaps":["Downstream effectors of Arf6 inactivation that drive neurite extension were not identified","Whether ACAP2 acts at recycling endosomes versus the plasma membrane was not fully distinguished"]},{"year":2014,"claim":"The Rab35/ACAP2 module was shown to negatively regulate oligodendrocyte differentiation and myelination by inactivating Arf6, broadening the pathway's physiological scope to CNS glia.","evidence":"siRNA knockdown epistasis and oligodendrocyte–neuronal co-culture with pharmacological validation","pmids":["24600047"],"confidence":"High","gaps":["In vivo myelination phenotypes in knockout animals were not examined","The Arf6 effector(s) controlling morphological differentiation downstream were not identified"]},{"year":2015,"claim":"The molecular basis of Rab35–ACAP2 specificity was resolved: Thr-76/Thr-81 in Rab35 switch II and Asn-610/Asn-691 in ACAP2's C-terminal domain are essential for binding, and binding-deficient mutants fail to support neurite outgrowth.","evidence":"Point mutagenesis, co-immunoprecipitation, and knockdown-rescue in PC12 cells","pmids":["25694427"],"confidence":"High","gaps":["No structural model of the Rab35–ACAP2 complex exists","Whether post-translational modifications modulate this interface is unknown"]},{"year":2015,"claim":"A GAP-independent pro-apoptotic role for ACAP2 was revealed: ACAP2 shares phosphoinositide-binding properties with its C. elegans ortholog CNT-1, and its knockdown blocks 5-FU–induced apoptosis, linking ACAP2 to programmed cell death.","evidence":"siRNA knockdown and phosphoinositide-binding assays in human cancer cell lines","pmids":["25853217"],"confidence":"Medium","gaps":["Single-lab finding; independent replication in other cancer cell types is lacking","Whether the pro-apoptotic function requires GAP activity or only phosphoinositide binding is unresolved","Mechanism connecting phosphoinositide binding to apoptosis execution is unknown"]},{"year":2025,"claim":"ACAP2 protein stability was shown to be controlled by RNF126-mediated ubiquitination and proteasomal degradation, and this degradation reprograms lipid metabolism to promote ovarian cancer progression.","evidence":"Co-immunoprecipitation, cycloheximide chase, siRNA knockdown, and in vivo xenograft in ovarian cancer cells","pmids":["40251363"],"confidence":"Medium","gaps":["The specific ubiquitination sites on ACAP2 were not mapped","Mechanism linking ACAP2 loss to lipid metabolic reprogramming is not delineated","Whether RNF126-ACAP2 regulation occurs in normal physiology or is cancer-specific is unknown"]},{"year":null,"claim":"A high-resolution structural understanding of ACAP2's GAP domain in complex with Arf6 and of the Rab35–ACAP2 interface is lacking, and the in vivo physiological consequences of ACAP2 loss in mammalian development and myelination have not been characterized genetically.","evidence":"","pmids":[],"confidence":"High","gaps":["No crystal or cryo-EM structure of ACAP2 alone or in complex","No conditional or constitutive ACAP2 knockout mouse phenotype reported","Relationship between GAP-dependent and GAP-independent (apoptotic) functions is unresolved"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[0,1,2]},{"term_id":"GO:0008289","term_label":"lipid binding","supporting_discovery_ids":[4]}],"localization":[{"term_id":"GO:0005768","term_label":"endosome","supporting_discovery_ids":[0,1]},{"term_id":"GO:0031410","term_label":"cytoplasmic vesicle","supporting_discovery_ids":[0,1]}],"pathway":[{"term_id":"R-HSA-5653656","term_label":"Vesicle-mediated transport","supporting_discovery_ids":[0,1,2]},{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[4]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[1]}],"complexes":[],"partners":["RAB35","ARF6","RNF126"],"other_free_text":[]},"mechanistic_narrative":"ACAP2 (centaurin-β2) is an Arf6 GTPase-activating protein that functions as an effector of GTP-bound Rab35, coupling Rab35-dependent membrane recruitment to local Arf6 inactivation at endosomes and phagocytic cups to regulate neurite outgrowth, Fcγ receptor–mediated phagocytosis, and oligodendrocyte differentiation/myelination [PMID:22344257, PMID:22045739, PMID:24600047]. Specificity of the Rab35–ACAP2 interaction is determined by two threonine residues in Rab35 switch II (Thr-76, Thr-81) and two asparagine residues in the ACAP2 C-terminal domain (Asn-610, Asn-691), and disruption of this interface abolishes neurite outgrowth [PMID:25694427]. ACAP2 also possesses a phosphoinositide-binding–dependent pro-apoptotic function, as its knockdown blocks 5-fluorouracil–induced apoptosis in cancer cells [PMID:25853217]. ACAP2 protein levels are regulated by RNF126-mediated ubiquitination and proteasomal degradation, which in ovarian cancer cells reprograms lipid metabolism to promote proliferation and metastasis [PMID:40251363]."},"prefetch_data":{"uniprot":{"accession":"Q15057","full_name":"Arf-GAP with coiled-coil, ANK repeat and PH domain-containing protein 2","aliases":["Centaurin-beta-2","Cnt-b2"],"length_aa":778,"mass_kda":88.0,"function":"GTPase-activating protein (GAP) for ADP ribosylation factor 6 (ARF6). Doesn't show GAP activity for RAB35 (PubMed:30905672)","subcellular_location":"Cell membrane; Endosome membrane","url":"https://www.uniprot.org/uniprotkb/Q15057/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/ACAP2","classification":"Not Classified","n_dependent_lines":7,"n_total_lines":1208,"dependency_fraction":0.005794701986754967},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000114331","cell_line_id":"CID000674","localizations":[{"compartment":"cytoplasmic","grade":3}],"interactors":[{"gene":"ACAP3","stoichiometry":4.0},{"gene":"CAPZB","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/target/CID000674","total_profiled":1310},"omim":[{"mim_id":"607766","title":"ARF-GAP WITH COILED-COIL, ANKYRIN REPEAT, AND PLECKSTRIN HOMOLOGY DOMAINS 2; ACAP2","url":"https://www.omim.org/entry/607766"},{"mim_id":"607763","title":"ARF-GAP WITH COILED-COIL, ANKYRIN REPEAT, AND PLECKSTRIN HOMOLOGY DOMAINS 1; ACAP1","url":"https://www.omim.org/entry/607763"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Endosomes","reliability":"Approved"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/ACAP2"},"hgnc":{"alias_symbol":["KIAA0041","CNT-B2"],"prev_symbol":["CENTB2"]},"alphafold":{"accession":"Q15057","domains":[{"cath_id":"1.20.1270.60","chopping":"6-235","consensus_level":"high","plddt":95.0591,"start":6,"end":235},{"cath_id":"2.30.29.30","chopping":"268-364","consensus_level":"high","plddt":87.9385,"start":268,"end":364},{"cath_id":"1.10.220.150","chopping":"409-507","consensus_level":"high","plddt":93.3514,"start":409,"end":507},{"cath_id":"1.25.40.20","chopping":"605-767","consensus_level":"medium","plddt":91.3645,"start":605,"end":767}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q15057","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q15057-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q15057-F1-predicted_aligned_error_v6.png","plddt_mean":81.38},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=ACAP2","jax_strain_url":"https://www.jax.org/strain/search?query=ACAP2"},"sequence":{"accession":"Q15057","fasta_url":"https://rest.uniprot.org/uniprotkb/Q15057.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q15057/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q15057"}},"corpus_meta":[{"pmid":"22344257","id":"PMC_22344257","title":"Rab35 regulates Arf6 activity through centaurin-β2 (ACAP2) during neurite outgrowth.","date":"2012","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/22344257","citation_count":124,"is_preprint":false},{"pmid":"7584044","id":"PMC_7584044","title":"Prediction of the coding sequences of unidentified human genes. II. The coding sequences of 40 new genes (KIAA0041-KIAA0080) deduced by analysis of cDNA clones from human cell line KG-1.","date":"1994","source":"DNA research : an international journal for rapid publication of reports on genes and genomes","url":"https://pubmed.ncbi.nlm.nih.gov/7584044","citation_count":122,"is_preprint":false},{"pmid":"30212824","id":"PMC_30212824","title":"The CircRNA-ACAP2/Hsa-miR-21-5p/ Tiam1 Regulatory Feedback Circuit Affects the Proliferation, Migration, and Invasion of Colon Cancer SW480 Cells.","date":"2018","source":"Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/30212824","citation_count":113,"is_preprint":false},{"pmid":"22045739","id":"PMC_22045739","title":"Rab35 regulates phagosome formation through recruitment of ACAP2 in macrophages during FcγR-mediated phagocytosis.","date":"2011","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/22045739","citation_count":72,"is_preprint":false},{"pmid":"34671193","id":"PMC_34671193","title":"Cancer-associated fibroblasts-derived exosomal miR-3656 promotes the development and progression of esophageal squamous cell carcinoma via the ACAP2/PI3K-AKT signaling pathway.","date":"2021","source":"International journal of biological sciences","url":"https://pubmed.ncbi.nlm.nih.gov/34671193","citation_count":55,"is_preprint":false},{"pmid":"16806385","id":"PMC_16806385","title":"Vaccinia virus K1L protein supports viral replication in human and rabbit cells through a cell-type-specific set of its ankyrin repeat residues that are distinct from its binding site for ACAP2.","date":"2006","source":"Virology","url":"https://pubmed.ncbi.nlm.nih.gov/16806385","citation_count":38,"is_preprint":false},{"pmid":"24600047","id":"PMC_24600047","title":"Rab35, acting through ACAP2 switching off Arf6, negatively regulates oligodendrocyte differentiation and myelination.","date":"2014","source":"Molecular biology of the cell","url":"https://pubmed.ncbi.nlm.nih.gov/24600047","citation_count":37,"is_preprint":false},{"pmid":"32406223","id":"PMC_32406223","title":"CircRNA ACAP2 induces myocardial apoptosis after myocardial infarction by sponging miR-29.","date":"2020","source":"Minerva medica","url":"https://pubmed.ncbi.nlm.nih.gov/32406223","citation_count":31,"is_preprint":false},{"pmid":"34085707","id":"PMC_34085707","title":"Circ-ACAP2 facilitates the progression of colorectal cancer through mediating miR-143-3p/FZD4 axis.","date":"2021","source":"European journal of clinical investigation","url":"https://pubmed.ncbi.nlm.nih.gov/34085707","citation_count":28,"is_preprint":false},{"pmid":"33363013","id":"PMC_33363013","title":"CircRNA_ACAP2 Suppresses EMT in Head and Neck Squamous Cell Carcinoma by Targeting the miR-21-5p/STAT3 Signaling Axis.","date":"2020","source":"Frontiers in oncology","url":"https://pubmed.ncbi.nlm.nih.gov/33363013","citation_count":21,"is_preprint":false},{"pmid":"34139744","id":"PMC_34139744","title":"CircRNA ACAP2 Is Overexpressed in Myocardial Infarction and Promotes the Maturation of miR-532 to Induce the Apoptosis of Cardiomyocyte.","date":"2021","source":"Journal of cardiovascular pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/34139744","citation_count":15,"is_preprint":false},{"pmid":"25694427","id":"PMC_25694427","title":"Structure-function analyses of the small GTPase Rab35 and its effector protein centaurin-β2/ACAP2 during neurite outgrowth of PC12 cells.","date":"2015","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/25694427","citation_count":15,"is_preprint":false},{"pmid":"35070838","id":"PMC_35070838","title":"CircRNA-ACAP2 contributes to the invasion, migration, and anti-apoptosis of neuroblastoma cells through targeting the miRNA-143-3p-hexokinase 2 axis.","date":"2021","source":"Translational pediatrics","url":"https://pubmed.ncbi.nlm.nih.gov/35070838","citation_count":13,"is_preprint":false},{"pmid":"25853217","id":"PMC_25853217","title":"Human ACAP2 is a homolog of C. elegans CNT-1 that promotes apoptosis in cancer cells.","date":"2015","source":"Cell cycle (Georgetown, Tex.)","url":"https://pubmed.ncbi.nlm.nih.gov/25853217","citation_count":8,"is_preprint":false},{"pmid":"40251363","id":"PMC_40251363","title":"RNF126 Promotes Ovarian Cancer Progression by Reprogramming Lipid Metabolism Through Degradation of ACAP2.","date":"2025","source":"Biochemical genetics","url":"https://pubmed.ncbi.nlm.nih.gov/40251363","citation_count":1,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.05.02.25326847","title":"A Fusion-Based Multiomics Classification Approach for Enhanced Gene Discovery in Non-Small Cell Lung Cancer","date":"2025-05-05","source":"bioRxiv","url":"https://doi.org/10.1101/2025.05.02.25326847","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":9561,"output_tokens":1770,"usd":0.027616},"stage2":{"model":"claude-opus-4-6","input_tokens":5041,"output_tokens":2249,"usd":0.122145},"total_usd":0.149761,"stage1_batch_id":"msgbatch_01TqNM6p6FsoMBT2jzQTwKBr","stage2_batch_id":"msgbatch_01Ya473eco8sTHbvRymQaU8o","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2012,\n      \"finding\": \"ACAP2 (centaurin-β2) functions as a Rab35 effector and as an Arf6-GTPase-activating protein (GAP) during neurite outgrowth. Rab35 recruits ACAP2 to Arf6-positive endosomes in response to NGF stimulation, and ACAP2's Arf6-GAP activity at these endosomes is required for neurite outgrowth in PC12 cells.\",\n      \"method\": \"Knockdown and rescue experiments, live-cell imaging, co-localization, GTPase activity assays\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — reciprocal functional assays, knockdown/rescue, replicated across multiple cellular contexts\",\n      \"pmids\": [\"22344257\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Rab35 recruits ACAP2 (an ARF6-GAP) to phagocytic cups during FcγR-mediated phagocytosis in macrophages; GTP-Rab35-dependent recruitment of ACAP2 regulates actin-dependent phagosome formation, and overexpression of ACAP2 together with GTP-locked Rab35 synergistically inhibits phagocytosis.\",\n      \"method\": \"Live-cell imaging, RNAi knockdown, expression of GDP/GTP-locked Rab35 mutants, co-expression assays\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — clean KO/KD with defined cellular phenotype, multiple mutant constructs, functional epistasis\",\n      \"pmids\": [\"22045739\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Rab35 and its effector ACAP2 negatively regulate oligodendrocyte morphological differentiation and myelination by inactivating (switching off) Arf6; knockdown of Rab35 or ACAP2 promotes differentiation and myelination, while Arf6 knockdown inhibits it, placing Rab35/ACAP2 upstream of Arf6 in this pathway.\",\n      \"method\": \"siRNA knockdown, oligodendrocyte-neuronal co-culture, GTPase activity assays, pharmacological inhibition of cytohesin-2\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — genetic epistasis with multiple knockdowns and pharmacological validation\",\n      \"pmids\": [\"24600047\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Two threonine residues (Thr-76 and Thr-81) in the switch II region of Rab35 are required for binding ACAP2, and two asparagine residues (Asn-610 and Asn-691) in ACAP2 are key for specific Rab35 recognition; binding-deficient mutants of either protein fail to support neurite outgrowth in PC12 cells.\",\n      \"method\": \"Deletion and point mutation analyses, co-immunoprecipitation, knockdown-rescue experiments\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — mutagenesis with functional validation and rescue experiments\",\n      \"pmids\": [\"25694427\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Human ACAP2 is a functional homolog of C. elegans CNT-1; ACAP2 has a pro-apoptotic function and shares an identical phosphoinositide-binding pattern with truncated CNT-1 (tCNT-1). Knockdown of ACAP2 blocks apoptosis in cancer cells in response to 5-fluorouracil.\",\n      \"method\": \"siRNA knockdown, apoptosis assay, phosphoinositide-binding assay\",\n      \"journal\": \"Cell cycle\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — single lab, knockdown with defined phenotype plus binding assay\",\n      \"pmids\": [\"25853217\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Vaccinia virus K1L protein binds ACAP2 (a GAP for ARF6); however, mutational analysis showed that residues required for VV replication in human or rabbit cells are distinct from the ACAP2-binding site on K1L, indicating K1L's host-range function is independent of its ACAP2 interaction.\",\n      \"method\": \"Mutagenesis, co-immunoprecipitation/binding assays, viral replication assays\",\n      \"journal\": \"Virology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — mutagenesis plus binding assay in single study; identifies ACAP2 as a K1L-binding partner\",\n      \"pmids\": [\"16806385\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"RNF126 (a ubiquitin E3 ligase) ubiquitinates and promotes degradation of ACAP2 protein in ovarian cancer cells; RNF126-mediated ACAP2 degradation reprograms lipid metabolism to promote ovarian cancer proliferation, migration, and metastasis.\",\n      \"method\": \"Co-immunoprecipitation, cycloheximide chase assay, siRNA knockdown, in vivo xenograft model\",\n      \"journal\": \"Biochemical genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — Co-IP plus protein stability assay in single study identifying RNF126 as ACAP2 ubiquitin ligase\",\n      \"pmids\": [\"40251363\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"ACAP2 (centaurin-β2) is an Arf6 GTPase-activating protein (GAP) that is recruited to specific membrane compartments (Arf6-positive endosomes, phagocytic cups) by acting as an effector of GTP-bound Rab35 — via specific residues in Rab35's switch II region and ACAP2's C-terminal domain — to inactivate Arf6 and thereby regulate neurite outgrowth, phagocytosis, and oligodendrocyte differentiation/myelination; additionally, ACAP2 has a pro-apoptotic phosphoinositide-binding function and is subject to ubiquitin-mediated proteasomal degradation by the E3 ligase RNF126.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"ACAP2 (centaurin-β2) is an Arf6 GTPase-activating protein that functions as an effector of GTP-bound Rab35, coupling Rab35-dependent membrane recruitment to local Arf6 inactivation at endosomes and phagocytic cups to regulate neurite outgrowth, Fcγ receptor–mediated phagocytosis, and oligodendrocyte differentiation/myelination [PMID:22344257, PMID:22045739, PMID:24600047]. Specificity of the Rab35–ACAP2 interaction is determined by two threonine residues in Rab35 switch II (Thr-76, Thr-81) and two asparagine residues in the ACAP2 C-terminal domain (Asn-610, Asn-691), and disruption of this interface abolishes neurite outgrowth [PMID:25694427]. ACAP2 also possesses a phosphoinositide-binding–dependent pro-apoptotic function, as its knockdown blocks 5-fluorouracil–induced apoptosis in cancer cells [PMID:25853217]. ACAP2 protein levels are regulated by RNF126-mediated ubiquitination and proteasomal degradation, which in ovarian cancer cells reprograms lipid metabolism to promote proliferation and metastasis [PMID:40251363].\",\n  \"teleology\": [\n    {\n      \"year\": 2006,\n      \"claim\": \"ACAP2 was identified as a physical interactor of vaccinia virus K1L protein, establishing it as a GAP for ARF6 accessible to viral manipulation, though K1L's host-range function proved independent of this binding.\",\n      \"evidence\": \"Mutagenesis and co-immunoprecipitation with vaccinia K1L in cell culture\",\n      \"pmids\": [\"16806385\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Only a single study; no independent confirmation of the K1L–ACAP2 interaction\",\n        \"Biological significance of the K1L–ACAP2 interaction remains undefined\",\n        \"No endogenous cellular phenotype linked to this interaction\"\n      ]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"The upstream activator Rab35 was shown to recruit ACAP2 to phagocytic cups during Fcγ receptor–mediated phagocytosis, establishing the Rab35→ACAP2→Arf6 axis as a regulatory module controlling actin-dependent phagosome formation in macrophages.\",\n      \"evidence\": \"Live-cell imaging, RNAi knockdown, and expression of GDP/GTP-locked Rab35 mutants in macrophages\",\n      \"pmids\": [\"22045739\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"The precise step at which Arf6 inactivation blocks phagosome closure was not resolved\",\n        \"Whether other Rab35 effectors contribute in parallel was not tested\"\n      ]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"The same Rab35→ACAP2→Arf6 cascade was extended to neuronal differentiation, showing that NGF-stimulated Rab35 recruits ACAP2 to Arf6-positive endosomes and that ACAP2 GAP activity is required for neurite outgrowth.\",\n      \"evidence\": \"Knockdown-rescue, live-cell imaging, and GTPase activity assays in PC12 cells\",\n      \"pmids\": [\"22344257\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Downstream effectors of Arf6 inactivation that drive neurite extension were not identified\",\n        \"Whether ACAP2 acts at recycling endosomes versus the plasma membrane was not fully distinguished\"\n      ]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"The Rab35/ACAP2 module was shown to negatively regulate oligodendrocyte differentiation and myelination by inactivating Arf6, broadening the pathway's physiological scope to CNS glia.\",\n      \"evidence\": \"siRNA knockdown epistasis and oligodendrocyte–neuronal co-culture with pharmacological validation\",\n      \"pmids\": [\"24600047\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"In vivo myelination phenotypes in knockout animals were not examined\",\n        \"The Arf6 effector(s) controlling morphological differentiation downstream were not identified\"\n      ]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"The molecular basis of Rab35–ACAP2 specificity was resolved: Thr-76/Thr-81 in Rab35 switch II and Asn-610/Asn-691 in ACAP2's C-terminal domain are essential for binding, and binding-deficient mutants fail to support neurite outgrowth.\",\n      \"evidence\": \"Point mutagenesis, co-immunoprecipitation, and knockdown-rescue in PC12 cells\",\n      \"pmids\": [\"25694427\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"No structural model of the Rab35–ACAP2 complex exists\",\n        \"Whether post-translational modifications modulate this interface is unknown\"\n      ]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"A GAP-independent pro-apoptotic role for ACAP2 was revealed: ACAP2 shares phosphoinositide-binding properties with its C. elegans ortholog CNT-1, and its knockdown blocks 5-FU–induced apoptosis, linking ACAP2 to programmed cell death.\",\n      \"evidence\": \"siRNA knockdown and phosphoinositide-binding assays in human cancer cell lines\",\n      \"pmids\": [\"25853217\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Single-lab finding; independent replication in other cancer cell types is lacking\",\n        \"Whether the pro-apoptotic function requires GAP activity or only phosphoinositide binding is unresolved\",\n        \"Mechanism connecting phosphoinositide binding to apoptosis execution is unknown\"\n      ]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"ACAP2 protein stability was shown to be controlled by RNF126-mediated ubiquitination and proteasomal degradation, and this degradation reprograms lipid metabolism to promote ovarian cancer progression.\",\n      \"evidence\": \"Co-immunoprecipitation, cycloheximide chase, siRNA knockdown, and in vivo xenograft in ovarian cancer cells\",\n      \"pmids\": [\"40251363\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"The specific ubiquitination sites on ACAP2 were not mapped\",\n        \"Mechanism linking ACAP2 loss to lipid metabolic reprogramming is not delineated\",\n        \"Whether RNF126-ACAP2 regulation occurs in normal physiology or is cancer-specific is unknown\"\n      ]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"A high-resolution structural understanding of ACAP2's GAP domain in complex with Arf6 and of the Rab35–ACAP2 interface is lacking, and the in vivo physiological consequences of ACAP2 loss in mammalian development and myelination have not been characterized genetically.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"No crystal or cryo-EM structure of ACAP2 alone or in complex\",\n        \"No conditional or constitutive ACAP2 knockout mouse phenotype reported\",\n        \"Relationship between GAP-dependent and GAP-independent (apoptotic) functions is unresolved\"\n      ]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [0, 1, 2]},\n      {\"term_id\": \"GO:0008289\", \"supporting_discovery_ids\": [4]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005768\", \"supporting_discovery_ids\": [0, 1]},\n      {\"term_id\": \"GO:0031410\", \"supporting_discovery_ids\": [0, 1]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-5653656\", \"supporting_discovery_ids\": [0, 1, 2]},\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [4]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [1]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\n      \"RAB35\",\n      \"ARF6\",\n      \"RNF126\"\n    ],\n    \"other_free_text\": []\n  }\n}\n```"}