{"gene":"WASHC2C","run_date":"2026-06-11T09:02:06","timeline":{"discoveries":[{"year":2008,"finding":"VPEF (WASHC2C/FAM21C) was detected on cell surface lipid rafts and on vesicle-like structures in the cytoplasm, and knockdown of VPEF blocked vaccinia virus penetration and intracellular transport of dextran, indicating VPEF mediates fluid phase (macropinocytic) endocytosis. Intracellular VPEF-containing vesicles partially colocalized with Rab11 but not Rab5a or caveolin, placing VPEF in recycling endosome trafficking.","method":"siRNA knockdown with virus penetration and dextran uptake assays; confocal colocalization with Rab5a, Rab11, caveolin markers; cell surface lipid raft fractionation","journal":"Journal of virology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — clean knockdown with defined cellular phenotype (fluid-phase endocytosis block) and colocalization to Rab11+ vesicles, single lab but multiple orthogonal methods","pmids":["18550675"],"is_preprint":false},{"year":2015,"finding":"VPEF/FAM21C (WASHC2C) recruits WASH and retromer protein complexes to early endosomes, and this WASH–VPEF/FAM21–retromer complex mediates endosomal membrane fission and cargo sorting into Rab11- and Rab22-positive recycling pathways. Functional interference with VPEF/FAM21 blocked sorting of vaccinia virus-containing vesicles prior to membrane fusion.","method":"Live single-particle fluorescence imaging; functional interference assays (dominant-negative constructs, siRNA knockdown) of WASH, FAM21, retromer components; colocalization with Rab11, Rab22, early endosome markers","journal":"Journal of virology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal functional interference methods plus live imaging, single lab, specific cellular phenotype readout","pmids":["26041286"],"is_preprint":false},{"year":2016,"finding":"The C-tail of FAM21C (WASHC2C) is required for complexing retromer to the WASH pentamer on endosomal membranes, and this FAM21C–retromer–WASH interaction is necessary for recycling of WT β1-AR and β2-AR from endosomes to the plasma membrane. Additionally, FKBP15 was identified as a FAM21C-binding endosomal protein that selectively regulates WT β1-AR (but not β2-AR) recycling.","method":"siRNA knockdown of FAM21C and FKBP15; receptor recycling assays (radioligand binding); epistasis with SNX3, Rab7a, SNX27","journal":"Cellular signalling","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — siRNA knockdown with defined recycling phenotype, epistasis with multiple pathway components, single lab","pmids":["27816670"],"is_preprint":false},{"year":2020,"finding":"Retromer-binding segments of WASHC2C were used in a reconstituted supported-lipid-bilayer system, and neither WASHC2C segments nor other accessory factors substantially affected retromer oligomerization state, indicating that WASHC2C does not potentiate retromer oligomerization on membranes.","method":"Quantitative single-particle fluorescence microscopy on supported lipid bilayers with purified fluorescently-labeled retromer, SNX3, RAB7, and retromer-binding segments of WASHC2C","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — reconstituted in vitro system with defined components, but this is a negative result for WASHC2C's effect on retromer oligomerization, single lab","pmids":["32651229"],"is_preprint":false},{"year":2022,"finding":"FAM21C (WASHC2C) interacts with CAPZA1 (capping protein alpha-1) via its CP-interacting (CPI) domain, and this binding inhibits the actin-filament capping capacity of CAPZA1, thereby promoting F-actin cytoskeleton remodeling, cell invasion and migration. Mutation of the CPI domain on FAM21C abolished its interaction with CAPZA1 and eliminated the pro-invasive effect.","method":"Co-immunoprecipitation; actin capping assays; CPI-domain mutagenesis; in vitro invasion/migration assays; in vivo xenograft models","journal":"Frontiers in oncology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP plus domain mutagenesis plus functional actin capping assay, single lab, multiple orthogonal methods","pmids":["35096613"],"is_preprint":false}],"current_model":"WASHC2C (FAM21C/VPEF) is a subunit of the WASH complex that localizes to endosomal membranes, where its C-terminal tail directly bridges retromer to the WASH pentamer to drive actin-dependent endosomal fission and cargo sorting into recycling pathways (Rab11/Rab22), while its CPI domain binds and inhibits the actin-capping protein CAPZA1 to promote actin cytoskeleton remodeling; it also localizes to cell-surface lipid rafts and facilitates fluid-phase (macropinocytic) endocytosis."},"narrative":{"mechanistic_narrative":"WASHC2C (FAM21C/VPEF) is an endosomal trafficking factor that couples the retromer cargo-recognition machinery to the actin-nucleating WASH complex to drive cargo sorting into recycling pathways [PMID:26041286, PMID:27816670]. Its C-terminal tail directly bridges retromer to the WASH pentamer on endosomal membranes, and this interaction is required for recycling of cargo such as β1- and β2-adrenergic receptors from endosomes to the plasma membrane, with FKBP15 acting as a WASHC2C-binding partner that selectively governs β1-AR recycling [PMID:27816670]. The WASH–WASHC2C–retromer assembly mediates endosomal membrane fission and routes cargo into Rab11- and Rab22-positive recycling compartments [PMID:26041286]. Through a CP-interacting (CPI) domain, WASHC2C binds and inhibits the actin-capping protein CAPZA1, promoting F-actin remodeling that supports cell invasion and migration; mutation of the CPI domain abolishes both CAPZA1 binding and the pro-invasive effect [PMID:35096613]. WASHC2C additionally localizes to cell-surface lipid rafts and cytoplasmic vesicles and is required for fluid-phase (macropinocytic) endocytosis [PMID:18550675]. In reconstituted supported-lipid-bilayer assays, WASHC2C does not alter retromer oligomerization state, indicating its bridging role does not operate by potentiating retromer assembly [PMID:32651229].","teleology":[{"year":2008,"claim":"Established WASHC2C as a membrane trafficking factor by showing it localizes to lipid rafts and recycling-endosome vesicles and is required for fluid-phase endocytosis, defining its cellular compartment before its molecular role was known.","evidence":"siRNA knockdown with virus penetration and dextran uptake assays plus confocal colocalization with Rab5a/Rab11/caveolin and lipid raft fractionation","pmids":["18550675"],"confidence":"Medium","gaps":["Molecular partners and the basis of vesicle recruitment not yet identified","Mechanism linking surface lipid-raft pool to intracellular recycling vesicles unresolved"]},{"year":2015,"claim":"Placed WASHC2C within a defined molecular machine by showing it recruits both WASH and retromer to early endosomes to drive membrane fission and sorting into Rab11/Rab22 recycling pathways, answering how it acts mechanistically in trafficking.","evidence":"Live single-particle imaging plus dominant-negative and siRNA functional interference of WASH/FAM21/retromer with Rab11/Rab22 colocalization","pmids":["26041286"],"confidence":"Medium","gaps":["Which structural region mediates each interaction not yet mapped","Direct versus indirect recruitment of retromer not distinguished"]},{"year":2016,"claim":"Mapped the bridging function to the WASHC2C C-tail and connected it to physiological cargo, showing the C-tail couples retromer to the WASH pentamer to recycle β-adrenergic receptors, and identified FKBP15 as a cargo-selective binding partner.","evidence":"siRNA knockdown of FAM21C and FKBP15 with radioligand receptor-recycling assays and epistasis against SNX3, Rab7a, SNX27","pmids":["27816670"],"confidence":"Medium","gaps":["Structural basis of the C-tail–retromer interface not resolved","How FKBP15 confers cargo selectivity for β1-AR unknown"]},{"year":2020,"claim":"Tested whether WASHC2C bridging works by promoting retromer assembly and found it does not affect retromer oligomerization in vitro, refining the model of how the C-tail engages retromer.","evidence":"Quantitative single-particle fluorescence microscopy on supported lipid bilayers with purified retromer, SNX3, RAB7, and WASHC2C retromer-binding segments","pmids":["32651229"],"confidence":"Medium","gaps":["Negative result; does not define what the bridging interaction does accomplish mechanically","Full-length WASHC2C and the WASH pentamer were not included in the reconstitution"]},{"year":2022,"claim":"Extended WASHC2C beyond cargo sorting to direct actin regulation by showing its CPI domain binds and inhibits CAPZA1 to promote F-actin remodeling and tumor cell invasion, linking it to cytoskeletal dynamics and disease relevant phenotypes.","evidence":"Co-immunoprecipitation, actin capping assays, CPI-domain mutagenesis, invasion/migration assays, and xenograft models","pmids":["35096613"],"confidence":"Medium","gaps":["Whether CAPZA1 inhibition is coupled to retromer/WASH-dependent sorting at endosomes not established","Reciprocal/endogenous validation of the interaction limited to single-lab co-IP"]},{"year":null,"claim":"How WASHC2C integrates its retromer-bridging, WASH-recruitment, and CAPZA1-inhibitory activities into a single coordinated fission/sorting event, and the structural basis of these interfaces, remain unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No structural model of the C-tail–retromer or CPI–CAPZA1 interfaces","Whether actin regulation and cargo sorting are mechanistically coupled is untested"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[1,2]},{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[4]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[4]}],"localization":[{"term_id":"GO:0005768","term_label":"endosome","supporting_discovery_ids":[0,1,2]},{"term_id":"GO:0031410","term_label":"cytoplasmic vesicle","supporting_discovery_ids":[0]},{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[0]}],"pathway":[{"term_id":"R-HSA-5653656","term_label":"Vesicle-mediated transport","supporting_discovery_ids":[1,2]},{"term_id":"R-HSA-9609507","term_label":"Protein localization","supporting_discovery_ids":[2]}],"complexes":["WASH complex"],"partners":["CAPZA1","FKBP15"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9Y4E1","full_name":"WASH complex subunit 2C","aliases":["Vaccinia virus penetration factor","VPEF"],"length_aa":1341,"mass_kda":147.2,"function":"Acts as a component of the WASH core complex that functions as a nucleation-promoting factor (NPF) at the surface of endosomes, where it recruits and activates the Arp2/3 complex to induce actin polymerization, playing a key role in the fission of tubules that serve as transport intermediates during endosome sorting. Mediates the recruitment of the WASH core complex to endosome membranes via binding to phospholipids and VPS35 of the retromer CSC. Mediates the recruitment of the F-actin-capping protein dimer to the WASH core complex probably promoting localized F-actin polymerization needed for vesicle scission (PubMed:19922874, PubMed:20498093, PubMed:22513087, PubMed:23331060). Via its C-terminus binds various phospholipids, most strongly phosphatidylinositol 4-phosphate (PtdIns-(4)P), phosphatidylinositol 5-phosphate (PtdIns-(5)P) and phosphatidylinositol 3,5-bisphosphate (PtdIns-(3,5)P2). Involved in the endosome-to-plasma membrane trafficking and recycling of SNX27-retromer-dependent cargo proteins, such as GLUT1 (PubMed:25278552). Required for the association of DNAJC13, ENTR1, ANKRD50 with retromer CSC subunit VPS35 (PubMed:24980502). Required for the endosomal recruitment of CCC and retriever complexes subunits COMMD1 and CCDC93 as well as the retrievere complex subunit VPS35L (PubMed:25355947, PubMed:28892079) (Microbial infection) Plays a role in fluid-phase endocytosis, a process exploited by vaccinia intracellular mature virus (IMV) to enter cells. As a result, may facilitate the penetration of IMV into cells","subcellular_location":"Early endosome membrane; Cell membrane","url":"https://www.uniprot.org/uniprotkb/Q9Y4E1/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/WASHC2C","classification":"Common Essential","n_dependent_lines":729,"n_total_lines":1165,"dependency_fraction":0.6257510729613734},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"CAPZB","stoichiometry":0.2},{"gene":"VPS35","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/WASHC2C","total_profiled":1310},"omim":[{"mim_id":"613631","title":"WASH COMPLEX, SUBUNIT 2C; WASHC2C","url":"https://www.omim.org/entry/613631"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Vesicles","reliability":"Supported"},{"location":"Nucleoli","reliability":"Additional"},{"location":"Cytosol","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/WASHC2C"},"hgnc":{"alias_symbol":["VPEF","KIAA0592"],"prev_symbol":["FAM21C"]},"alphafold":{"accession":"Q9Y4E1","domains":[],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9Y4E1","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9Y4E1-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9Y4E1-F1-predicted_aligned_error_v6.png","plddt_mean":47.88},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=WASHC2C","jax_strain_url":"https://www.jax.org/strain/search?query=WASHC2C"},"sequence":{"accession":"Q9Y4E1","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9Y4E1.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9Y4E1/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9Y4E1"}},"corpus_meta":[{"pmid":"18550675","id":"PMC_18550675","title":"A novel cellular protein, VPEF, facilitates vaccinia virus penetration into HeLa cells through fluid phase endocytosis.","date":"2008","source":"Journal of virology","url":"https://pubmed.ncbi.nlm.nih.gov/18550675","citation_count":80,"is_preprint":false},{"pmid":"20714399","id":"PMC_20714399","title":"Evolutionary conservation of the WASH complex, an actin polymerization machine involved in endosomal fission.","date":"2010","source":"Communicative & integrative biology","url":"https://pubmed.ncbi.nlm.nih.gov/20714399","citation_count":33,"is_preprint":false},{"pmid":"26041286","id":"PMC_26041286","title":"Intracellular Transport of Vaccinia Virus in HeLa Cells Requires WASH-VPEF/FAM21-Retromer Complexes and Recycling Molecules Rab11 and Rab22.","date":"2015","source":"Journal of virology","url":"https://pubmed.ncbi.nlm.nih.gov/26041286","citation_count":30,"is_preprint":false},{"pmid":"29193877","id":"PMC_29193877","title":"Effects of probiotics on ghrelin and lungs in children with acute lung injury: A double-blind randomized, controlled trial.","date":"2017","source":"Pediatric pulmonology","url":"https://pubmed.ncbi.nlm.nih.gov/29193877","citation_count":18,"is_preprint":false},{"pmid":"27816670","id":"PMC_27816670","title":"Two barcodes encoded by the type-1 PDZ and by phospho-Ser312 regulate retromer/WASH-mediated sorting of the ß1-adrenergic receptor from endosomes to the plasma membrane.","date":"2016","source":"Cellular signalling","url":"https://pubmed.ncbi.nlm.nih.gov/27816670","citation_count":16,"is_preprint":false},{"pmid":"32651229","id":"PMC_32651229","title":"Retromer forms low order oligomers on supported lipid bilayers.","date":"2020","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/32651229","citation_count":14,"is_preprint":false},{"pmid":"35096613","id":"PMC_35096613","title":"FAM21C Promotes Hepatocellular Carcinoma Invasion and Metastasis by Driving Actin Cytoskeleton Remodeling via Inhibiting Capping Ability of CAPZA1.","date":"2022","source":"Frontiers in oncology","url":"https://pubmed.ncbi.nlm.nih.gov/35096613","citation_count":6,"is_preprint":false},{"pmid":"28551275","id":"PMC_28551275","title":"Genetic analysis of VCP and WASH complex genes in a German cohort of sporadic ALS-FTD patients.","date":"2017","source":"Neurobiology of aging","url":"https://pubmed.ncbi.nlm.nih.gov/28551275","citation_count":6,"is_preprint":false},{"pmid":"41301950","id":"PMC_41301950","title":"Transcriptome Analysis Reveals Circadian Rhythmic Regulation of Lipid Metabolism and Immune Function in Chicken Livers.","date":"2025","source":"Animals : an open access journal from MDPI","url":"https://pubmed.ncbi.nlm.nih.gov/41301950","citation_count":1,"is_preprint":false},{"pmid":"41081810","id":"PMC_41081810","title":"Association of the endosomal sorting processes cargo selection and membrane tubulation with human reward system reactivity.","date":"2025","source":"European archives of psychiatry and clinical neuroscience","url":"https://pubmed.ncbi.nlm.nih.gov/41081810","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":7534,"output_tokens":1520,"usd":0.022701,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":8238,"output_tokens":2423,"usd":0.050882,"stage2_stop_reason":"end_turn"},"total_usd":0.073583,"stage1_batch_id":"msgbatch_01UT1AahFPCAyn2uVweS7A9z","stage2_batch_id":"msgbatch_01HuKWtKNT4Pqj1bc8eEP5RV","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2008,\n      \"finding\": \"VPEF (WASHC2C/FAM21C) was detected on cell surface lipid rafts and on vesicle-like structures in the cytoplasm, and knockdown of VPEF blocked vaccinia virus penetration and intracellular transport of dextran, indicating VPEF mediates fluid phase (macropinocytic) endocytosis. Intracellular VPEF-containing vesicles partially colocalized with Rab11 but not Rab5a or caveolin, placing VPEF in recycling endosome trafficking.\",\n      \"method\": \"siRNA knockdown with virus penetration and dextran uptake assays; confocal colocalization with Rab5a, Rab11, caveolin markers; cell surface lipid raft fractionation\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — clean knockdown with defined cellular phenotype (fluid-phase endocytosis block) and colocalization to Rab11+ vesicles, single lab but multiple orthogonal methods\",\n      \"pmids\": [\"18550675\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"VPEF/FAM21C (WASHC2C) recruits WASH and retromer protein complexes to early endosomes, and this WASH–VPEF/FAM21–retromer complex mediates endosomal membrane fission and cargo sorting into Rab11- and Rab22-positive recycling pathways. Functional interference with VPEF/FAM21 blocked sorting of vaccinia virus-containing vesicles prior to membrane fusion.\",\n      \"method\": \"Live single-particle fluorescence imaging; functional interference assays (dominant-negative constructs, siRNA knockdown) of WASH, FAM21, retromer components; colocalization with Rab11, Rab22, early endosome markers\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal functional interference methods plus live imaging, single lab, specific cellular phenotype readout\",\n      \"pmids\": [\"26041286\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"The C-tail of FAM21C (WASHC2C) is required for complexing retromer to the WASH pentamer on endosomal membranes, and this FAM21C–retromer–WASH interaction is necessary for recycling of WT β1-AR and β2-AR from endosomes to the plasma membrane. Additionally, FKBP15 was identified as a FAM21C-binding endosomal protein that selectively regulates WT β1-AR (but not β2-AR) recycling.\",\n      \"method\": \"siRNA knockdown of FAM21C and FKBP15; receptor recycling assays (radioligand binding); epistasis with SNX3, Rab7a, SNX27\",\n      \"journal\": \"Cellular signalling\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — siRNA knockdown with defined recycling phenotype, epistasis with multiple pathway components, single lab\",\n      \"pmids\": [\"27816670\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Retromer-binding segments of WASHC2C were used in a reconstituted supported-lipid-bilayer system, and neither WASHC2C segments nor other accessory factors substantially affected retromer oligomerization state, indicating that WASHC2C does not potentiate retromer oligomerization on membranes.\",\n      \"method\": \"Quantitative single-particle fluorescence microscopy on supported lipid bilayers with purified fluorescently-labeled retromer, SNX3, RAB7, and retromer-binding segments of WASHC2C\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — reconstituted in vitro system with defined components, but this is a negative result for WASHC2C's effect on retromer oligomerization, single lab\",\n      \"pmids\": [\"32651229\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"FAM21C (WASHC2C) interacts with CAPZA1 (capping protein alpha-1) via its CP-interacting (CPI) domain, and this binding inhibits the actin-filament capping capacity of CAPZA1, thereby promoting F-actin cytoskeleton remodeling, cell invasion and migration. Mutation of the CPI domain on FAM21C abolished its interaction with CAPZA1 and eliminated the pro-invasive effect.\",\n      \"method\": \"Co-immunoprecipitation; actin capping assays; CPI-domain mutagenesis; in vitro invasion/migration assays; in vivo xenograft models\",\n      \"journal\": \"Frontiers in oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP plus domain mutagenesis plus functional actin capping assay, single lab, multiple orthogonal methods\",\n      \"pmids\": [\"35096613\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"WASHC2C (FAM21C/VPEF) is a subunit of the WASH complex that localizes to endosomal membranes, where its C-terminal tail directly bridges retromer to the WASH pentamer to drive actin-dependent endosomal fission and cargo sorting into recycling pathways (Rab11/Rab22), while its CPI domain binds and inhibits the actin-capping protein CAPZA1 to promote actin cytoskeleton remodeling; it also localizes to cell-surface lipid rafts and facilitates fluid-phase (macropinocytic) endocytosis.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"WASHC2C (FAM21C/VPEF) is an endosomal trafficking factor that couples the retromer cargo-recognition machinery to the actin-nucleating WASH complex to drive cargo sorting into recycling pathways [#1, #2]. Its C-terminal tail directly bridges retromer to the WASH pentamer on endosomal membranes, and this interaction is required for recycling of cargo such as β1- and β2-adrenergic receptors from endosomes to the plasma membrane, with FKBP15 acting as a WASHC2C-binding partner that selectively governs β1-AR recycling [#2]. The WASH–WASHC2C–retromer assembly mediates endosomal membrane fission and routes cargo into Rab11- and Rab22-positive recycling compartments [#1]. Through a CP-interacting (CPI) domain, WASHC2C binds and inhibits the actin-capping protein CAPZA1, promoting F-actin remodeling that supports cell invasion and migration; mutation of the CPI domain abolishes both CAPZA1 binding and the pro-invasive effect [#4]. WASHC2C additionally localizes to cell-surface lipid rafts and cytoplasmic vesicles and is required for fluid-phase (macropinocytic) endocytosis [#0]. In reconstituted supported-lipid-bilayer assays, WASHC2C does not alter retromer oligomerization state, indicating its bridging role does not operate by potentiating retromer assembly [#3].\",\n  \"teleology\": [\n    {\n      \"year\": 2008,\n      \"claim\": \"Established WASHC2C as a membrane trafficking factor by showing it localizes to lipid rafts and recycling-endosome vesicles and is required for fluid-phase endocytosis, defining its cellular compartment before its molecular role was known.\",\n      \"evidence\": \"siRNA knockdown with virus penetration and dextran uptake assays plus confocal colocalization with Rab5a/Rab11/caveolin and lipid raft fractionation\",\n      \"pmids\": [\"18550675\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular partners and the basis of vesicle recruitment not yet identified\", \"Mechanism linking surface lipid-raft pool to intracellular recycling vesicles unresolved\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Placed WASHC2C within a defined molecular machine by showing it recruits both WASH and retromer to early endosomes to drive membrane fission and sorting into Rab11/Rab22 recycling pathways, answering how it acts mechanistically in trafficking.\",\n      \"evidence\": \"Live single-particle imaging plus dominant-negative and siRNA functional interference of WASH/FAM21/retromer with Rab11/Rab22 colocalization\",\n      \"pmids\": [\"26041286\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Which structural region mediates each interaction not yet mapped\", \"Direct versus indirect recruitment of retromer not distinguished\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Mapped the bridging function to the WASHC2C C-tail and connected it to physiological cargo, showing the C-tail couples retromer to the WASH pentamer to recycle β-adrenergic receptors, and identified FKBP15 as a cargo-selective binding partner.\",\n      \"evidence\": \"siRNA knockdown of FAM21C and FKBP15 with radioligand receptor-recycling assays and epistasis against SNX3, Rab7a, SNX27\",\n      \"pmids\": [\"27816670\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural basis of the C-tail–retromer interface not resolved\", \"How FKBP15 confers cargo selectivity for β1-AR unknown\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Tested whether WASHC2C bridging works by promoting retromer assembly and found it does not affect retromer oligomerization in vitro, refining the model of how the C-tail engages retromer.\",\n      \"evidence\": \"Quantitative single-particle fluorescence microscopy on supported lipid bilayers with purified retromer, SNX3, RAB7, and WASHC2C retromer-binding segments\",\n      \"pmids\": [\"32651229\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Negative result; does not define what the bridging interaction does accomplish mechanically\", \"Full-length WASHC2C and the WASH pentamer were not included in the reconstitution\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Extended WASHC2C beyond cargo sorting to direct actin regulation by showing its CPI domain binds and inhibits CAPZA1 to promote F-actin remodeling and tumor cell invasion, linking it to cytoskeletal dynamics and disease relevant phenotypes.\",\n      \"evidence\": \"Co-immunoprecipitation, actin capping assays, CPI-domain mutagenesis, invasion/migration assays, and xenograft models\",\n      \"pmids\": [\"35096613\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether CAPZA1 inhibition is coupled to retromer/WASH-dependent sorting at endosomes not established\", \"Reciprocal/endogenous validation of the interaction limited to single-lab co-IP\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How WASHC2C integrates its retromer-bridging, WASH-recruitment, and CAPZA1-inhibitory activities into a single coordinated fission/sorting event, and the structural basis of these interfaces, remain unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No structural model of the C-tail–retromer or CPI–CAPZA1 interfaces\", \"Whether actin regulation and cargo sorting are mechanistically coupled is untested\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [1, 2]},\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [4]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [4]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005768\", \"supporting_discovery_ids\": [0, 1, 2]},\n      {\"term_id\": \"GO:0031410\", \"supporting_discovery_ids\": [0]},\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [0]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-5653656\", \"supporting_discovery_ids\": [1, 2]},\n      {\"term_id\": \"R-HSA-9609507\", \"supporting_discovery_ids\": [2]}\n    ],\n    \"complexes\": [\"WASH complex\"],\n    \"partners\": [\"CAPZA1\", \"FKBP15\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}