{"gene":"ATP6V1D","run_date":"2026-06-09T22:02:44","timeline":{"discoveries":[{"year":1995,"finding":"Subunit D (VMA8/ATP6V1D) is an integral subunit of the V1 catalytic sector of V-ATPase; disruption of yeast VMA8 abolishes V-ATPase activity and prevents growth at neutral pH, and complementation with VMA8 restores wild-type phenotype, establishing its essential catalytic role.","method":"Yeast gene disruption (null mutant delta vma8::URA3), quinacrine accumulation assay, complementation with plasmid-borne VMA8; bovine cDNA sequencing and structural homology to F-ATPase gamma subunit","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic null mutant with functional rescue, enzymatic activity assay, replicated by two independent 1995 papers (PMID 7831318 and 7797485)","pmids":["7831318"],"is_preprint":false},{"year":1995,"finding":"VMA8p (yeast subunit D) is required for assembly of the V1 sector onto vacuolar membranes: in vma8Δ cells, V1 subunits fail to associate with the vacuolar membrane while Vo subunits are correctly targeted; Vma8p itself assembles only as part of the fully assembled V-ATPase and not with an isolated Vo subcomplex.","method":"Vacuolar membrane isolation, immunoblot analysis of V1/Vo subunit distribution, density gradient fractionation, V-ATPase activity assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal biochemical methods (fractionation, immunoblot, activity assay) in a single rigorous study, consistent with companion paper PMID 7831318","pmids":["7797485"],"is_preprint":false},{"year":2024,"finding":"ATP6V1D promotes lysosomal acidification and enhances the interaction between CHMP4B and IST1 to foster ESCRT-III complex assembly, thereby facilitating autophagosome-lysosome fusion and maintaining autophagic flux, which drives hepatocellular carcinoma stemness and progression.","method":"CRISPR-Cas9 metabolic knockout screen, ATP6V1D knockdown in vitro and in vivo, co-immunoprecipitation (co-IP) of CHMP4B and IST1, autophagic flux assays, lysosomal acidification measurements","journal":"Autophagy","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP for ESCRT-III interaction, CRISPR/knockdown with functional readouts; single lab, mechanistic follow-up limited to abstract-level detail","pmids":["39316516"],"is_preprint":false},{"year":2026,"finding":"CRISPR/Cas9 disruption of ATP6V1D in rat macrophages impairs v-ATPase function and lysosomal acidification, leading to increased intracellular Toxoplasma gondii growth; overexpression of GIMAP5 or GIMAP6 partially compensates for loss of ATP6V1D, demonstrating that ATP6V1D-dependent lysosomal acidification and GIMAP-driven lysosome-PVM fusion act additively to restrict parasite replication.","method":"CRISPR/Cas9 knockout of ATP6V1D in NR8383 rat macrophage cell line, v-ATPase activity assay, lysosomal acidification measurement, GIMAP transgene overexpression, intracellular parasite growth quantification","journal":"Microbiology spectrum","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — CRISPR KO with enzymatic readout plus genetic epistasis with GIMAP overexpression; single lab, single study","pmids":["41972776"],"is_preprint":false}],"current_model":"ATP6V1D encodes the D subunit of the V1 catalytic sector of the vacuolar H⁺-ATPase (V-ATPase); it is essential for V1 assembly onto vacuolar membranes and for proton-pumping activity, and in mammalian cells it additionally promotes lysosomal acidification, ESCRT-III (CHMP4B–IST1) complex assembly, and autophagosome-lysosome fusion, thereby regulating autophagic flux, cancer cell stemness, and innate immunity against intracellular pathogens."},"narrative":{"mechanistic_narrative":"ATP6V1D encodes the D subunit of the V1 catalytic sector of the vacuolar H⁺-ATPase, where it is an integral subunit essential for proton-pumping activity and for organellar acidification [PMID:7831318]. Genetic ablation studies establish that subunit D is required for assembly of the V1 sector onto the vacuolar membrane: in its absence, V1 subunits fail to associate with the membrane while the Vo sector is correctly targeted, and subunit D itself incorporates only into the fully assembled enzyme rather than with an isolated Vo subcomplex [PMID:7797485]. In mammalian cells, ATP6V1D-dependent V-ATPase function drives lysosomal acidification that supports downstream membrane-trafficking and host-defense programs: it promotes CHMP4B–IST1 interaction and ESCRT-III assembly to facilitate autophagosome-lysosome fusion and sustain autophagic flux, a process that contributes to hepatocellular carcinoma stemness and progression [PMID:39316516], and it restricts intracellular Toxoplasma gondii replication, acting additively with GIMAP-driven lysosome-parasitophorous vacuole fusion [PMID:41972776].","teleology":[{"year":1995,"claim":"Established that subunit D is an essential catalytic component of the V-ATPase, defining the gene's core function before any role in higher-order cellular processes was known.","evidence":"Yeast VMA8 null mutant with functional rescue, quinacrine accumulation and V-ATPase activity assays, with bovine cDNA sequencing","pmids":["7831318"],"confidence":"High","gaps":["Did not resolve the structural position of subunit D within the V1 rotor mechanism in mammalian cells","No direct mammalian loss-of-function characterization at this stage"]},{"year":1995,"claim":"Defined the assembly requirement of subunit D, showing it is needed for V1 docking onto the membrane and only joins as part of the complete enzyme, clarifying when and how the subunit is incorporated.","evidence":"Vacuolar membrane fractionation, immunoblot of V1/Vo distribution, density gradient fractionation and activity assays in yeast","pmids":["7797485"],"confidence":"High","gaps":["Mechanism of regulated V1/Vo reversible assembly not addressed","Did not test whether mammalian ATP6V1D follows the identical assembly logic"]},{"year":2024,"claim":"Connected ATP6V1D-driven lysosomal acidification to ESCRT-III–mediated autophagosome-lysosome fusion and to a disease phenotype, extending the subunit from a housekeeping role to a regulator of autophagic flux and cancer cell stemness.","evidence":"CRISPR-Cas9 metabolic knockout screen, knockdown in vitro/in vivo, CHMP4B–IST1 co-IP, autophagic flux and lysosomal acidification assays in hepatocellular carcinoma","pmids":["39316516"],"confidence":"Medium","gaps":["Co-IP for CHMP4B–IST1 lacks reciprocal and reconstitution validation","Whether ATP6V1D acts on ESCRT-III directly or only via acidification is unresolved","Single-lab study with abstract-level mechanistic detail"]},{"year":2026,"claim":"Showed ATP6V1D-dependent lysosomal acidification contributes to innate restriction of an intracellular pathogen, acting additively with GIMAP-driven membrane fusion, broadening the subunit's role into host defense.","evidence":"CRISPR/Cas9 knockout in NR8383 rat macrophages, V-ATPase activity and acidification assays, GIMAP5/6 overexpression epistasis, intracellular parasite growth quantification","pmids":["41972776"],"confidence":"Medium","gaps":["Single lab, single study using one rat macrophage line","Molecular link between acidification and parasite restriction not mechanistically dissected","Genetic interaction with GIMAP shown by overexpression rather than endogenous epistasis"]},{"year":null,"claim":"How ATP6V1D coordinates V-ATPase proton pumping with ESCRT-III assembly and pathogen restriction at the molecular level, and whether these are direct or acidification-dependent effects, remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No structural model of mammalian ATP6V1D within an assembled V-ATPase in the corpus","Direct vs. indirect role in CHMP4B–IST1 recruitment not separated from acidification","Reversible V1/Vo assembly regulation in mammalian cells uncharacterized"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140657","term_label":"ATP-dependent activity","supporting_discovery_ids":[0,1]},{"term_id":"GO:0005215","term_label":"transporter activity","supporting_discovery_ids":[0,1,3]}],"localization":[{"term_id":"GO:0005773","term_label":"vacuole","supporting_discovery_ids":[0,1]},{"term_id":"GO:0005764","term_label":"lysosome","supporting_discovery_ids":[2,3]}],"pathway":[{"term_id":"R-HSA-9612973","term_label":"Autophagy","supporting_discovery_ids":[2]},{"term_id":"R-HSA-382551","term_label":"Transport of small molecules","supporting_discovery_ids":[0,1]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[3]}],"complexes":["V-ATPase V1 sector"],"partners":["CHMP4B","IST1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9Y5K8","full_name":"V-type proton ATPase subunit D","aliases":["V-ATPase 28 kDa accessory protein","Vacuolar proton pump subunit D"],"length_aa":247,"mass_kda":28.3,"function":"Subunit of the V1 complex of vacuolar(H+)-ATPase (V-ATPase), a multisubunit enzyme composed of a peripheral complex (V1) that hydrolyzes ATP and a membrane integral complex (V0) that translocates protons (PubMed:33065002). V-ATPase is responsible for acidifying and maintaining the pH of intracellular compartments and in some cell types, is targeted to the plasma membrane, where it is responsible for acidifying the extracellular environment (By similarity). May play a role in cilium biogenesis through regulation of the transport and the localization of proteins to the cilium (PubMed:21844891)","subcellular_location":"Membrane; Cytoplasmic vesicle, clathrin-coated vesicle membrane; Cytoplasm, cytoskeleton, microtubule organizing center, centrosome; Cell projection, cilium","url":"https://www.uniprot.org/uniprotkb/Q9Y5K8/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/ATP6V1D","classification":"Common Essential","n_dependent_lines":1050,"n_total_lines":1208,"dependency_fraction":0.8692052980132451},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000100554","cell_line_id":"CID001924","localizations":[{"compartment":"cytoplasmic","grade":3},{"compartment":"nucleoplasm","grade":2},{"compartment":"vesicles","grade":2}],"interactors":[{"gene":"ATP6AP2","stoichiometry":10.0},{"gene":"ATP6V1A","stoichiometry":10.0},{"gene":"ATP6V1B2","stoichiometry":10.0},{"gene":"ATP6V1H","stoichiometry":10.0},{"gene":"ATP6V1E1","stoichiometry":10.0},{"gene":"ATP6V1G1","stoichiometry":10.0},{"gene":"DAG1","stoichiometry":4.0},{"gene":"ATP6V0A1","stoichiometry":0.2},{"gene":"ATP6V0D1","stoichiometry":0.2},{"gene":"CEP135","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/target/CID001924","total_profiled":1310},"omim":[{"mim_id":"614780","title":"SORTING NEXIN 10; SNX10","url":"https://www.omim.org/entry/614780"},{"mim_id":"609398","title":"ATPase, H+ TRANSPORTING, LYSOSOMAL, 34-KD, V1 SUBUNIT D; ATP6V1D","url":"https://www.omim.org/entry/609398"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Plasma membrane","reliability":"Approved"},{"location":"Nucleoplasm","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/ATP6V1D"},"hgnc":{"alias_symbol":["VATD","VMA8"],"prev_symbol":["ATP6M"]},"alphafold":{"accession":"Q9Y5K8","domains":[{"cath_id":"1.10.287.3240","chopping":"12-184","consensus_level":"high","plddt":90.9329,"start":12,"end":184}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9Y5K8","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9Y5K8-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9Y5K8-F1-predicted_aligned_error_v6.png","plddt_mean":86.25},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=ATP6V1D","jax_strain_url":"https://www.jax.org/strain/search?query=ATP6V1D"},"sequence":{"accession":"Q9Y5K8","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9Y5K8.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9Y5K8/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9Y5K8"}},"corpus_meta":[{"pmid":"7831318","id":"PMC_7831318","title":"A bovine cDNA and a yeast gene (VMA8) encoding the subunit D of the vacuolar H(+)-ATPase.","date":"1995","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/7831318","citation_count":78,"is_preprint":false},{"pmid":"10868806","id":"PMC_10868806","title":"Quinupristin/dalfopristin-resistant enterococci of the satA (vatD) and satG (vatE) genotypes from different ecological origins in Germany.","date":"2000","source":"Microbial drug resistance (Larchmont, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/10868806","citation_count":57,"is_preprint":false},{"pmid":"11841212","id":"PMC_11841212","title":"Crystal structure of Vat(D): an acetyltransferase that inactivates streptogramin group A antibiotics.","date":"2002","source":"Biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/11841212","citation_count":52,"is_preprint":false},{"pmid":"7797485","id":"PMC_7797485","title":"VMA8 encodes a 32-kDa V1 subunit of the Saccharomyces cerevisiae vacuolar H(+)-ATPase required for function and assembly of the enzyme complex.","date":"1995","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/7797485","citation_count":48,"is_preprint":false},{"pmid":"11600385","id":"PMC_11600385","title":"Indication of transposition of a mobile DNA element containing the vat(D) and erm(B) genes in Enterococcus faecium.","date":"2001","source":"Antimicrobial agents and chemotherapy","url":"https://pubmed.ncbi.nlm.nih.gov/11600385","citation_count":27,"is_preprint":false},{"pmid":"24135791","id":"PMC_24135791","title":"Role of periplasmic binding proteins, FatB and VatD, in the vulnibactin utilization system of Vibrio vulnificus M2799.","date":"2013","source":"Microbial pathogenesis","url":"https://pubmed.ncbi.nlm.nih.gov/24135791","citation_count":9,"is_preprint":false},{"pmid":"39316516","id":"PMC_39316516","title":"ATP6V1D drives hepatocellular carcinoma stemness and progression via both lysosome acidification-dependent and -independent mechanisms.","date":"2024","source":"Autophagy","url":"https://pubmed.ncbi.nlm.nih.gov/39316516","citation_count":8,"is_preprint":false},{"pmid":"11435709","id":"PMC_11435709","title":"cDNA cloning, chromosomal localization and evolutionary analysis of mouse vacuolar ATPase subunit D, Atp6m.","date":"2001","source":"Cytogenetics and cell genetics","url":"https://pubmed.ncbi.nlm.nih.gov/11435709","citation_count":4,"is_preprint":false},{"pmid":"40309610","id":"PMC_40309610","title":"Association and functional study of ATP6V1D and GPHN gene polymorphisms with depression in Chinese population.","date":"2025","source":"World journal of psychiatry","url":"https://pubmed.ncbi.nlm.nih.gov/40309610","citation_count":1,"is_preprint":false},{"pmid":"26249703","id":"PMC_26249703","title":"Expression, purification, crystallization and X-ray crystallographic analysis of the periplasmic binding protein VatD from Vibrio vulnificus M2799.","date":"2015","source":"Acta crystallographica. Section F, Structural biology communications","url":"https://pubmed.ncbi.nlm.nih.gov/26249703","citation_count":1,"is_preprint":false},{"pmid":"40911183","id":"PMC_40911183","title":"Differential Expression of ATP6V1D and Its Diagnostic Potential in IgA Nephropathy.","date":"2025","source":"Current medical science","url":"https://pubmed.ncbi.nlm.nih.gov/40911183","citation_count":0,"is_preprint":false},{"pmid":"40693774","id":"PMC_40693774","title":"Detection of a streptogramin A O-acetyltransferase gene (vatD) in the chromosome of Clostridium botulinum isolated from infants in the United States.","date":"2025","source":"Applied and environmental microbiology","url":"https://pubmed.ncbi.nlm.nih.gov/40693774","citation_count":0,"is_preprint":false},{"pmid":"41972776","id":"PMC_41972776","title":"Synergy between upregulated small GTPase immunity-associated proteins and lysosomal ATP6V1D in restricting intracellular Toxoplasma growth.","date":"2026","source":"Microbiology spectrum","url":"https://pubmed.ncbi.nlm.nih.gov/41972776","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.01.31.635826","title":"Initial leads to combat streptogramin resistance generated from X-ray fragment screening against VatD","date":"2025-02-02","source":"bioRxiv","url":"https://doi.org/10.1101/2025.01.31.635826","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":9270,"output_tokens":1253,"usd":0.023303,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":7850,"output_tokens":2170,"usd":0.04675,"stage2_stop_reason":"end_turn"},"total_usd":0.070053,"stage1_batch_id":"msgbatch_0179Z2pxRYBsXi2xADneafRw","stage2_batch_id":"msgbatch_01LaY9ZaVdgjWArcZo9eqQE5","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1995,\n      \"finding\": \"Subunit D (VMA8/ATP6V1D) is an integral subunit of the V1 catalytic sector of V-ATPase; disruption of yeast VMA8 abolishes V-ATPase activity and prevents growth at neutral pH, and complementation with VMA8 restores wild-type phenotype, establishing its essential catalytic role.\",\n      \"method\": \"Yeast gene disruption (null mutant delta vma8::URA3), quinacrine accumulation assay, complementation with plasmid-borne VMA8; bovine cDNA sequencing and structural homology to F-ATPase gamma subunit\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic null mutant with functional rescue, enzymatic activity assay, replicated by two independent 1995 papers (PMID 7831318 and 7797485)\",\n      \"pmids\": [\"7831318\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1995,\n      \"finding\": \"VMA8p (yeast subunit D) is required for assembly of the V1 sector onto vacuolar membranes: in vma8Δ cells, V1 subunits fail to associate with the vacuolar membrane while Vo subunits are correctly targeted; Vma8p itself assembles only as part of the fully assembled V-ATPase and not with an isolated Vo subcomplex.\",\n      \"method\": \"Vacuolar membrane isolation, immunoblot analysis of V1/Vo subunit distribution, density gradient fractionation, V-ATPase activity assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal biochemical methods (fractionation, immunoblot, activity assay) in a single rigorous study, consistent with companion paper PMID 7831318\",\n      \"pmids\": [\"7797485\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"ATP6V1D promotes lysosomal acidification and enhances the interaction between CHMP4B and IST1 to foster ESCRT-III complex assembly, thereby facilitating autophagosome-lysosome fusion and maintaining autophagic flux, which drives hepatocellular carcinoma stemness and progression.\",\n      \"method\": \"CRISPR-Cas9 metabolic knockout screen, ATP6V1D knockdown in vitro and in vivo, co-immunoprecipitation (co-IP) of CHMP4B and IST1, autophagic flux assays, lysosomal acidification measurements\",\n      \"journal\": \"Autophagy\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP for ESCRT-III interaction, CRISPR/knockdown with functional readouts; single lab, mechanistic follow-up limited to abstract-level detail\",\n      \"pmids\": [\"39316516\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"CRISPR/Cas9 disruption of ATP6V1D in rat macrophages impairs v-ATPase function and lysosomal acidification, leading to increased intracellular Toxoplasma gondii growth; overexpression of GIMAP5 or GIMAP6 partially compensates for loss of ATP6V1D, demonstrating that ATP6V1D-dependent lysosomal acidification and GIMAP-driven lysosome-PVM fusion act additively to restrict parasite replication.\",\n      \"method\": \"CRISPR/Cas9 knockout of ATP6V1D in NR8383 rat macrophage cell line, v-ATPase activity assay, lysosomal acidification measurement, GIMAP transgene overexpression, intracellular parasite growth quantification\",\n      \"journal\": \"Microbiology spectrum\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — CRISPR KO with enzymatic readout plus genetic epistasis with GIMAP overexpression; single lab, single study\",\n      \"pmids\": [\"41972776\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"ATP6V1D encodes the D subunit of the V1 catalytic sector of the vacuolar H⁺-ATPase (V-ATPase); it is essential for V1 assembly onto vacuolar membranes and for proton-pumping activity, and in mammalian cells it additionally promotes lysosomal acidification, ESCRT-III (CHMP4B–IST1) complex assembly, and autophagosome-lysosome fusion, thereby regulating autophagic flux, cancer cell stemness, and innate immunity against intracellular pathogens.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"ATP6V1D encodes the D subunit of the V1 catalytic sector of the vacuolar H\\u207a-ATPase, where it is an integral subunit essential for proton-pumping activity and for organellar acidification [#0]. Genetic ablation studies establish that subunit D is required for assembly of the V1 sector onto the vacuolar membrane: in its absence, V1 subunits fail to associate with the membrane while the Vo sector is correctly targeted, and subunit D itself incorporates only into the fully assembled enzyme rather than with an isolated Vo subcomplex [#1]. In mammalian cells, ATP6V1D-dependent V-ATPase function drives lysosomal acidification that supports downstream membrane-trafficking and host-defense programs: it promotes CHMP4B\\u2013IST1 interaction and ESCRT-III assembly to facilitate autophagosome-lysosome fusion and sustain autophagic flux, a process that contributes to hepatocellular carcinoma stemness and progression [#2], and it restricts intracellular Toxoplasma gondii replication, acting additively with GIMAP-driven lysosome-parasitophorous vacuole fusion [#3].\",\n  \"teleology\": [\n    {\n      \"year\": 1995,\n      \"claim\": \"Established that subunit D is an essential catalytic component of the V-ATPase, defining the gene's core function before any role in higher-order cellular processes was known.\",\n      \"evidence\": \"Yeast VMA8 null mutant with functional rescue, quinacrine accumulation and V-ATPase activity assays, with bovine cDNA sequencing\",\n      \"pmids\": [\"7831318\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Did not resolve the structural position of subunit D within the V1 rotor mechanism in mammalian cells\",\n        \"No direct mammalian loss-of-function characterization at this stage\"\n      ]\n    },\n    {\n      \"year\": 1995,\n      \"claim\": \"Defined the assembly requirement of subunit D, showing it is needed for V1 docking onto the membrane and only joins as part of the complete enzyme, clarifying when and how the subunit is incorporated.\",\n      \"evidence\": \"Vacuolar membrane fractionation, immunoblot of V1/Vo distribution, density gradient fractionation and activity assays in yeast\",\n      \"pmids\": [\"7797485\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Mechanism of regulated V1/Vo reversible assembly not addressed\",\n        \"Did not test whether mammalian ATP6V1D follows the identical assembly logic\"\n      ]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Connected ATP6V1D-driven lysosomal acidification to ESCRT-III\\u2013mediated autophagosome-lysosome fusion and to a disease phenotype, extending the subunit from a housekeeping role to a regulator of autophagic flux and cancer cell stemness.\",\n      \"evidence\": \"CRISPR-Cas9 metabolic knockout screen, knockdown in vitro/in vivo, CHMP4B\\u2013IST1 co-IP, autophagic flux and lysosomal acidification assays in hepatocellular carcinoma\",\n      \"pmids\": [\"39316516\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Co-IP for CHMP4B\\u2013IST1 lacks reciprocal and reconstitution validation\",\n        \"Whether ATP6V1D acts on ESCRT-III directly or only via acidification is unresolved\",\n        \"Single-lab study with abstract-level mechanistic detail\"\n      ]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Showed ATP6V1D-dependent lysosomal acidification contributes to innate restriction of an intracellular pathogen, acting additively with GIMAP-driven membrane fusion, broadening the subunit's role into host defense.\",\n      \"evidence\": \"CRISPR/Cas9 knockout in NR8383 rat macrophages, V-ATPase activity and acidification assays, GIMAP5/6 overexpression epistasis, intracellular parasite growth quantification\",\n      \"pmids\": [\"41972776\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Single lab, single study using one rat macrophage line\",\n        \"Molecular link between acidification and parasite restriction not mechanistically dissected\",\n        \"Genetic interaction with GIMAP shown by overexpression rather than endogenous epistasis\"\n      ]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How ATP6V1D coordinates V-ATPase proton pumping with ESCRT-III assembly and pathogen restriction at the molecular level, and whether these are direct or acidification-dependent effects, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"No structural model of mammalian ATP6V1D within an assembled V-ATPase in the corpus\",\n        \"Direct vs. indirect role in CHMP4B\\u2013IST1 recruitment not separated from acidification\",\n        \"Reversible V1/Vo assembly regulation in mammalian cells uncharacterized\"\n      ]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140657\", \"supporting_discovery_ids\": [0, 1]},\n      {\"term_id\": \"GO:0005215\", \"supporting_discovery_ids\": [0, 1, 3]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005773\", \"supporting_discovery_ids\": [0, 1]},\n      {\"term_id\": \"GO:0005764\", \"supporting_discovery_ids\": [2, 3]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-9612973\", \"supporting_discovery_ids\": [2]},\n      {\"term_id\": \"R-HSA-382551\", \"supporting_discovery_ids\": [0, 1]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [3]}\n    ],\n    \"complexes\": [\n      \"V-ATPase V1 sector\"\n    ],\n    \"partners\": [\n      \"CHMP4B\",\n      \"IST1\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":3,"faith_total":3,"faith_pct":100.0}}