{"gene":"ARMC3","run_date":"2026-06-09T22:02:44","timeline":{"discoveries":[{"year":1998,"finding":"Yeast Vac8 (ARMC3 ortholog) localizes to the vacuolar membrane via N-terminal myristylation; deletion of the myristylation site abolishes vacuolar localization. Loss of Vac8 causes accumulation of small fragmented vacuoles and defective vacuolar inheritance.","method":"GFP fusion live imaging, N-terminal myristylation mutant analysis, deletion genetics","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct localization experiment with functional consequence and mutagenesis, single lab","pmids":["9664035"],"is_preprint":false},{"year":2005,"finding":"Vac8 palmitoylation on isolated yeast vacuoles is mediated by the R-SNARE Ykt6 and is part of a SNARE subcomplex distinct from the Nyv1-containing complex; this reaction is ATP-independent, restricted to a narrow time window, and stimulated by EDTA (ion chelation). Palmitoylation is required for vacuole fusion.","method":"In vitro vacuole fusion assay with purified vacuoles, antibody inhibition, protein complex analysis","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vitro reconstitution on purified vacuoles with biochemical complex analysis, single lab","pmids":["15701652"],"is_preprint":false},{"year":2005,"finding":"The DHHC palmitoyl acyltransferase Pfa3 is specifically required for efficient vacuolar localization of Vac8 in vivo; Pfa3 deletion impairs Vac8 palmitoylation and reduces vacuole fusion, while vacuole morphology and inheritance appear normal.","method":"DHHC deletion mutant analysis, in vivo localization, vacuole fusion assay","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic deletion combined with localization and functional fusion assay, single lab","pmids":["16301533"],"is_preprint":false},{"year":2006,"finding":"Stable vacuolar membrane binding of Vac8 requires two N-terminal cysteines for palmitoylation regardless of their combination; palmitoylation adds functional roles beyond membrane anchoring — a basic-residue replacement mutant that still localizes to vacuoles can support cytoplasm-to-vacuole transport but requires at least one palmitoylation cysteine for vacuolar morphology and inheritance functions.","method":"Cysteine mutagenesis, vacuolar inheritance assay, Cvt pathway assay, fluorescence microscopy","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — systematic mutagenesis with multiple functional readouts, single lab","pmids":["16720644"],"is_preprint":false},{"year":2006,"finding":"In Pichia pastoris, Vac8 ARM repeat domains (central region) are required for formation of vacuolar arm-like extensions that engulf peroxisomes during micropexophagy, and Vac8 is essential for recruitment of Atg11 to the vacuolar membrane during glucose-induced pexophagy; palmitoylation/myristoylation sites are required for protein stability and vacuolar association.","method":"Domain deletion mutants, GFP localization, pexophagy assay (alcohol oxidase activity), genetic knockouts","journal":"Autophagy","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — domain-deletion mutants with multiple functional readouts in Pichia pastoris, single lab","pmids":["16921262"],"is_preprint":false},{"year":2006,"finding":"In Pichia pastoris, the ARM repeat domain of Vac8 is required for vacuolar inheritance but not for micropexophagy; deletion of both ARM and C-terminal domains abolishes vacuolar sequestering membrane formation and abolishes recruitment of Atg11 to the vacuolar membrane during micropexophagy.","method":"Domain deletion mutants, fluorescence microscopy, vacuolar inheritance assay","journal":"Autophagy","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — domain-specific deletion mutants with distinct functional readouts, single lab","pmids":["16874085"],"is_preprint":false},{"year":2009,"finding":"Pfa3 palmitoylates each of the three N-terminal cysteines of Vac8 in vitro, with efficiency enhanced by prior N-myristoylation; the 11th armadillo repeat of Vac8 is a key determinant for specific recognition by Pfa3, as shown by chimeric protein and competition experiments.","method":"In vitro palmitoylation assay, chimeric protein analysis, competition assay with truncation mutants","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution of palmitoylation with mutagenesis, chimeric proteins, and competition assays in single rigorous study","pmids":["19416974"],"is_preprint":false},{"year":2019,"finding":"Crystal structure of Vac8 bound to Atg13 reveals that the Atg13 extended loop (70 Å) binds the ARM domain of Vac8 in an antiparallel manner; the N-terminal H1 helix of Vac8 intramolecularly associates with ARM1 and regulates Vac8 self-association (dimerization), which is required differentially for Cvt and PMN autophagy pathways. Different quaternary structures of Vac8 (Atg13-bound heterotetramer vs. Nvj1-bound complex) mediate distinct autophagic functions.","method":"X-ray crystallography, analytical ultracentrifugation, isothermal titration calorimetry, SAXS, in vivo functional assays","journal":"Autophagy","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure combined with ITC, AUC, SAXS, and in vivo validation with multiple orthogonal methods","pmids":["31512555"],"is_preprint":false},{"year":2019,"finding":"The Atg13 C-terminus binds lipid membranes via electrostatic interactions and hydrophobic insertion of a Phe residue; this phospholipid binding and Vac8 binding are mutually exclusive because they involve overlapping residues in the Atg13 IDR, and both interactions are required for efficient autophagy.","method":"Isothermal titration calorimetry, circular dichroism, lipid-binding assays, mutagenesis","journal":"Autophagy","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — in vitro binding assays with mutagenesis, single lab, mechanistically defines Vac8-Atg13 interaction as mutually exclusive with membrane binding","pmids":["31352862"],"is_preprint":false},{"year":2020,"finding":"Vac8 anchors the phagophore assembly site (PAS) to the vacuolar membrane by binding Atg13 and thereby recruiting the Atg1 initiation complex; VAC8 deletion or Vac8 mislocalization reduce autophagy activity, establishing Vac8 as required for correct vacuolar localization of the PAS.","method":"VAC8 deletion, auxin-inducible protein degradation (AID), fluorescence microscopy, autophagy activity assay","journal":"Autophagy","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — conditional deletion with fluorescence microscopy and functional autophagy readout, single lab","pmids":["32508216"],"is_preprint":false},{"year":2016,"finding":"A 1 bp frameshift deletion (p.A451fs26) in bovine ARMC3 that truncates the protein by 401 amino acids (46%) is causally associated with a sterilizing tail stump sperm defect characterized by severely disorganized, immotile spermatozoa tails, establishing ARMC3 as required for normal spermatogenesis.","method":"Homozygosity mapping, whole-genome re-sequencing, segregation analysis in affected bulls","journal":"BMC genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — causal variant identification with segregation analysis in multiple affected animals, but no in vitro functional rescue","pmids":["26923438"],"is_preprint":false},{"year":2021,"finding":"Mouse ARMC3 is the functional homolog of yeast Vac8; its ARM domains recruit PtdIns3K-CI (class III PI3K complex I) to the phagophore assembly site to initiate autophagosome formation via PtdIns3P generation. Armc3 knockout mice show blocked ribophagy in spermatids, low mitochondrial energy levels, immotile flagella, and male infertility.","method":"Mouse knockout, ribophagy assay, mitochondrial function assay, yeast complementation studies","journal":"Autophagy","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — knockout mouse with multiple cellular phenotype readouts and cross-species complementation, single lab","pmids":["34705610"],"is_preprint":false},{"year":2023,"finding":"X-ray crystal structure of the Vac8-Vac17 complex reveals a bipartite interaction interface; binding of Vac17 to Vac8 clamps the H1 helix to ARM1, preventing Vac8 dimerization and thereby competitively inhibiting Vac8 interactions with Nvj1 and Atg13. Mutation of key interface residues severely impairs vacuole inheritance in vivo.","method":"X-ray crystallography, mutagenesis, binding affinity measurements, in vivo vacuole inheritance assay","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure with mutagenesis and in vivo functional validation, mechanistically explaining how Vac8-mediated processes are mutually exclusive","pmids":["37094131"],"is_preprint":false},{"year":2023,"finding":"PI3KCI interacts with the vacuolar membrane anchor Vac8 via the Atg14 C-terminal region in a constitutive manner; this interaction cooperates with Atg38-Atg1 complex and Vps30-Atg9 interactions to target PI3KCI to the PAS for autophagosome biogenesis.","method":"Co-immunoprecipitation, deletion mutant analysis, autophagy flux assays, genetic epistasis","journal":"The Journal of cell biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal co-IP with deletion mutants and functional autophagy readouts, single lab","pmids":["37436710"],"is_preprint":false},{"year":2024,"finding":"A homozygous splicing variant (c.916+1G>A) in human ARMC3 causes exon 8 skipping, producing a truncated protein undetectable by Western blot in patient sperm, and results in asthenozoospermia with disrupted flagellar ultrastructure including vacuolated sperm mitochondria at the midpiece.","method":"Whole-exome sequencing, RT-PCR, Western blot, transmission electron microscopy of patient sperm","journal":"Clinical genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — patient variant with protein-level confirmation and TEM ultrastructural analysis, single family","pmids":["39221575"],"is_preprint":false},{"year":2025,"finding":"The vacuole-specific adaptor Vac17 interacts with Myo2 (yeast myosin V) through two distinct binding sites (handhold mechanism); cryo-EM and structure prediction show one of these sites links to Vac8 on the vacuole membrane, forming the Myo2-Vac17-Vac8 complex for vacuole transport to daughter cells.","method":"Cryo-electron microscopy, AlphaFold structure prediction, genetic analysis, cell-based transport assays","journal":"bioRxiv","confidence":"Low","confidence_rationale":"Tier 2 / Weak — preprint, cryo-EM with structure prediction, single lab, no peer review","pmids":["bio_10.1101_2025.03.24.645041"],"is_preprint":true}],"current_model":"ARMC3 (mammalian) / Vac8 (yeast ortholog) is an armadillo-repeat protein anchored to the vacuolar/lysosomal membrane via N-terminal myristoylation and palmitoylation (performed by the DHHC enzyme Pfa3, with the 11th ARM repeat conferring substrate specificity); through its ARM domain it recruits PI3K complex I (via Atg14) and the Atg1 initiation complex (via Atg13) to the phagophore assembly site to initiate autophagy, while it also forms a tripartite Myo2-Vac17-Vac8 complex for myosin V-driven vacuole inheritance; distinct quaternary structures of Vac8 (monomer vs. dimer) differentially regulate these binding partners, ensuring vacuole inheritance, NVJ formation, and selective autophagy (Cvt, ribophagy, pexophagy) are mutually exclusive; in mammals, ARMC3 loss blocks ribophagy in spermatids, disrupts flagellar ultrastructure, and causes male infertility."},"narrative":{"mechanistic_narrative":"ARMC3 (mammalian) and its yeast ortholog Vac8 are armadillo-repeat proteins that act as membrane-anchored scaffolds coupling the vacuolar/lysosomal membrane to autophagic and membrane-inheritance machinery [PMID:31512555, PMID:34705610]. Vac8 is targeted to the vacuolar membrane by N-terminal myristoylation, with deletion of the myristoylation site abolishing localization and producing fragmented vacuoles and inheritance defects [PMID:9664035]; stable membrane attachment further requires palmitoylation of N-terminal cysteines, a modification installed by the DHHC acyltransferase Pfa3 — whose recognition of Vac8 is conferred specifically by the 11th armadillo repeat — and which adds functional roles beyond mere anchoring [PMID:16301533, PMID:16720644, PMID:19416974]. Through its ARM domain, Vac8 anchors the phagophore assembly site to the vacuole by binding Atg13, thereby recruiting the Atg1 initiation complex, and additionally recruits class III PI3K complex I via the Atg14 C-terminus to drive autophagosome biogenesis [PMID:31512555, PMID:32508216, PMID:37436710]. The same scaffold supports vacuole inheritance through a tripartite Myo2–Vac17–Vac8 complex [PMID:bio_10.1101_2025.03.24.645041]. These activities are made mutually exclusive by the protein's quaternary state: an intramolecular H1-helix/ARM1 contact governs Vac8 self-association, and Vac17 binding clamps H1 to ARM1 to block dimerization and thereby competitively exclude the Nvj1 and Atg13 interactions [PMID:31512555, PMID:37094131]. In mammals, ARMC3 is the functional homolog of Vac8, with its ARM domains recruiting PtdIns3K-CI to initiate autophagosome formation; loss of ARMC3 blocks ribophagy in spermatids, lowers mitochondrial energy, and produces immotile flagella [PMID:34705610]. Truncating ARMC3 mutations cause sterilizing sperm tail defects in cattle and asthenozoospermia with disrupted flagellar ultrastructure in humans, establishing ARMC3 as required for normal spermatogenesis [PMID:26923438, PMID:39221575].","teleology":[{"year":1998,"claim":"Established how Vac8 reaches its site of action and that it is functionally required for vacuole maintenance, answering where this protein works.","evidence":"GFP fusion live imaging and N-terminal myristylation mutant analysis with deletion genetics in yeast","pmids":["9664035"],"confidence":"Medium","gaps":["Did not resolve the molecular partners through which Vac8 directs inheritance","Role of additional lipid modifications not yet defined"]},{"year":2005,"claim":"Identified that Vac8 also undergoes palmitoylation and that this modification is required for vacuole fusion, distinguishing lipid anchoring from fusion function.","evidence":"In vitro vacuole fusion assay on purified yeast vacuoles with antibody inhibition and complex analysis; DHHC deletion mutant analysis identifying Pfa3","pmids":["15701652","16301533"],"confidence":"Medium","gaps":["Direct enzyme-substrate mechanism of Pfa3 on Vac8 not yet reconstituted","Whether palmitoylation has roles separable from fusion unresolved"]},{"year":2006,"claim":"Dissected which Vac8 modifications and domains support which functions, separating membrane anchoring from inheritance, Cvt transport, and pexophagy.","evidence":"Cysteine and domain-deletion mutagenesis with vacuolar inheritance, Cvt, and pexophagy (alcohol oxidase) readouts in yeast and Pichia pastoris","pmids":["16720644","16921262","16874085"],"confidence":"Medium","gaps":["Structural basis of ARM-domain partner recognition not yet defined","Identity of all ARM-domain effectors incomplete"]},{"year":2009,"claim":"Defined the molecular specificity of Vac8 palmitoylation, showing N-myristoylation primes the reaction and the 11th ARM repeat dictates Pfa3 recognition.","evidence":"In vitro palmitoylation assay with chimeric proteins and competition/truncation mutants","pmids":["19416974"],"confidence":"High","gaps":["In vivo dynamics of sequential modification not addressed","Whether mammalian ARMC3 is similarly modified untested here"]},{"year":2019,"claim":"Provided the structural logic of how Vac8 self-association and Atg13 binding underlie distinct autophagy pathways, explaining functional partitioning.","evidence":"X-ray crystallography of Vac8-Atg13, AUC, ITC, SAXS with in vivo assays; complementary ITC/CD lipid-binding analysis of Atg13","pmids":["31512555","31352862"],"confidence":"High","gaps":["How upstream signals switch Vac8 quaternary state not defined","Mammalian structural conservation untested"]},{"year":2020,"claim":"Showed Vac8 anchors the phagophore assembly site to the vacuole by recruiting the Atg1 initiation complex via Atg13, defining its role in autophagy initiation.","evidence":"VAC8 deletion and auxin-inducible degradation with fluorescence microscopy and autophagy activity assays in yeast","pmids":["32508216"],"confidence":"Medium","gaps":["Quantitative contribution of Vac8 anchoring versus other PAS tethers unresolved","Regulation of PAS positioning by signals not addressed"]},{"year":2023,"claim":"Resolved how PI3K complex I is brought to the PAS through Vac8 and how Vac17 enforces mutual exclusivity among Vac8 functions.","evidence":"Co-IP with deletion mutants and autophagy flux assays for the Atg14-Vac8 interaction; crystal structure of Vac8-Vac17 with mutagenesis and in vivo inheritance assays","pmids":["37436710","37094131"],"confidence":"High","gaps":["Temporal coordination of competing partner binding in vivo not mapped","How the cell selects between inheritance and autophagy states unclear"]},{"year":2021,"claim":"Established mammalian ARMC3 as the functional Vac8 homolog and linked its autophagy function to spermatogenesis, bridging yeast mechanism to mammalian physiology.","evidence":"Armc3 knockout mice with ribophagy and mitochondrial assays plus yeast complementation","pmids":["34705610"],"confidence":"Medium","gaps":["Direct biochemical demonstration of ARMC3 partner binding in mammalian cells limited","Subcellular membrane anchor of ARMC3 in spermatids not defined"]},{"year":2024,"claim":"Connected ARMC3 loss-of-function to human disease, showing truncating variants cause asthenozoospermia with flagellar and mitochondrial ultrastructural defects.","evidence":"Bovine homozygosity mapping/whole-genome sequencing of tail-stump defect; human whole-exome sequencing, RT-PCR, Western blot, and TEM of patient sperm","pmids":["26923438","39221575"],"confidence":"Medium","gaps":["No functional rescue confirming causality in human cells","Mechanism linking autophagy defect to flagellar disassembly not directly demonstrated"]},{"year":2025,"claim":"Structurally defined the Myo2-Vac17-Vac8 transport complex, clarifying how Vac8 couples to myosin V for vacuole inheritance.","evidence":"Cryo-EM with AlphaFold prediction, genetics, and transport assays (preprint)","pmids":["bio_10.1101_2025.03.24.645041"],"confidence":"Low","gaps":["Preprint, not peer reviewed","Vac8-Vac17 linkage inferred partly from structure prediction","Mammalian relevance of the transport complex untested"]},{"year":null,"claim":"How ARMC3-dependent autophagy is mechanistically required for mammalian flagellar assembly and what regulates the switch between its competing scaffolding states remains open.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No direct mammalian biochemistry of ARMC3 partner recruitment","Signal controlling Vac8/ARMC3 quaternary state unknown","Causal chain from ribophagy block to flagellar defect undefined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[7,9,11,13]},{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[15]}],"localization":[{"term_id":"GO:0005773","term_label":"vacuole","supporting_discovery_ids":[0,4,5]}],"pathway":[{"term_id":"R-HSA-9612973","term_label":"Autophagy","supporting_discovery_ids":[7,9,11,13]},{"term_id":"R-HSA-9609507","term_label":"Protein localization","supporting_discovery_ids":[15]}],"complexes":["Myo2-Vac17-Vac8 vacuole inheritance complex","Vac8-Atg13-Atg1 initiation complex","Vac8-PI3K complex I (via Atg14)"],"partners":["ATG13","ATG14","VAC17","MYO2","NVJ1","PFA3","YKT6","ATG11"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q5W041","full_name":"Armadillo repeat-containing protein 3","aliases":["Beta-catenin-like protein","Cancer/testis antigen 81","CT81","KU-CT-1"],"length_aa":872,"mass_kda":96.4,"function":"Essential for male fertility and sperm motility (By similarity). During spermatogenesis, promotes the autophagic degradation of excessive ribosomes, providing energy resources for mitochondria and thus ensuring sperm flagellar motility (PubMed:34428398)","subcellular_location":"","url":"https://www.uniprot.org/uniprotkb/Q5W041/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/ARMC3","classification":"Not Classified","n_dependent_lines":0,"n_total_lines":1208,"dependency_fraction":0.0},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/ARMC3","total_profiled":1310},"omim":[{"mim_id":"611226","title":"ARMADILLO REPEAT-CONTAINING PROTEIN 3; ARMC3","url":"https://www.omim.org/entry/611226"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Group enriched","tissue_distribution":"Detected in some","driving_tissues":[{"tissue":"choroid plexus","ntpm":48.7},{"tissue":"fallopian tube","ntpm":41.9},{"tissue":"testis","ntpm":31.4}],"url":"https://www.proteinatlas.org/search/ARMC3"},"hgnc":{"alias_symbol":["FLJ32827","CT81","VAC8"],"prev_symbol":[]},"alphafold":{"accession":"Q5W041","domains":[{"cath_id":"1.25.10.10","chopping":"446-551","consensus_level":"medium","plddt":95.5454,"start":446,"end":551},{"cath_id":"-","chopping":"602-608_718-872","consensus_level":"high","plddt":90.2962,"start":602,"end":872}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q5W041","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q5W041-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q5W041-F1-predicted_aligned_error_v6.png","plddt_mean":84.81},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=ARMC3","jax_strain_url":"https://www.jax.org/strain/search?query=ARMC3"},"sequence":{"accession":"Q5W041","fasta_url":"https://rest.uniprot.org/uniprotkb/Q5W041.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q5W041/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q5W041"}},"corpus_meta":[{"pmid":"9664035","id":"PMC_9664035","title":"YEB3/VAC8 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Vac8 at the yeast vacuole.","date":"2006","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/16720644","citation_count":44,"is_preprint":false},{"pmid":"26923438","id":"PMC_26923438","title":"A frameshift mutation in ARMC3 is associated with a tail stump sperm defect in Swedish Red (Bos taurus) cattle.","date":"2016","source":"BMC genetics","url":"https://pubmed.ncbi.nlm.nih.gov/26923438","citation_count":34,"is_preprint":false},{"pmid":"37436710","id":"PMC_37436710","title":"The Atg1 complex, Atg9, and Vac8 recruit PI3K complex I to the pre-autophagosomal structure.","date":"2023","source":"The Journal of cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/37436710","citation_count":28,"is_preprint":false},{"pmid":"32508216","id":"PMC_32508216","title":"Vac8 determines phagophore assembly site vacuolar localization during nitrogen starvation-induced 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gene.","date":"2006","source":"Genetika","url":"https://pubmed.ncbi.nlm.nih.gov/16915934","citation_count":3,"is_preprint":false},{"pmid":"39045779","id":"PMC_39045779","title":"The emerging significance of Vac8, a multi-purpose armadillo-repeat protein in yeast.","date":"2024","source":"Autophagy","url":"https://pubmed.ncbi.nlm.nih.gov/39045779","citation_count":2,"is_preprint":false},{"pmid":"41440702","id":"PMC_41440702","title":"The Vacuolar Protein 8 (Vac8) Homolog in Cryptococcus neoformans Impacts Stress Responses and Virulence Traits Through Conserved and Unique Roles.","date":"2025","source":"Journal of fungi (Basel, Switzerland)","url":"https://pubmed.ncbi.nlm.nih.gov/41440702","citation_count":0,"is_preprint":false},{"pmid":"40661350","id":"PMC_40661350","title":"The vacuolar protein 8 (Vac8) homolog in Cryptococcus neoformans plays conserved and unique roles in vacuolar and cellular morphology, impacting important stress responses and virulence traits.","date":"2025","source":"bioRxiv : the preprint server for biology","url":"https://pubmed.ncbi.nlm.nih.gov/40661350","citation_count":0,"is_preprint":false},{"pmid":"37465006","id":"PMC_37465006","title":"Erratum to \"Chemical Complementarity of Breast Cancer Resident, T-Cell Receptor CDR3 Domains and the Cancer Antigen, ARMC3, is Associated With Higher Levels of Survival and Granzyme Expression\".","date":"2023","source":"Cancer informatics","url":"https://pubmed.ncbi.nlm.nih.gov/37465006","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.05.22.655601","title":"The vacuolar protein 8 (Vac8) homolog in  <i>Cryptococcus neoformans</i>  plays conserved and unique roles in vacuolar and cellular morphology, impacting important stress responses and virulence traits","date":"2025-05-22","source":"bioRxiv","url":"https://doi.org/10.1101/2025.05.22.655601","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2025.03.24.645041","title":"Cargo adaptors use a handhold mechanism to engage with myosin V for organelle transport","date":"2025-03-25","source":"bioRxiv","url":"https://doi.org/10.1101/2025.03.24.645041","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":14215,"output_tokens":4243,"usd":0.053145,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":11607,"output_tokens":3670,"usd":0.074893,"stage2_stop_reason":"end_turn"},"total_usd":0.128038,"stage1_batch_id":"msgbatch_0179Z2pxRYBsXi2xADneafRw","stage2_batch_id":"msgbatch_01LaY9ZaVdgjWArcZo9eqQE5","note":"batch pricing = 50% of standard"},"stage1_raw":"I need to carefully classify each paper. The query gene is ARMC3 (human/mammalian), with aliases including VAC8. \n\n**Classification analysis:**\n- Papers [1]-[14], [18], [21]-[23], [25]-[26]: These are about yeast/fungal VAC8. The alias \"VAC8\" is listed as an HGNC alias for ARMC3, but I need to determine if yeast Vac8 is an ortholog of human ARMC3. Paper [16] explicitly states \"mouse ARMC3 is the homolog of yeast Vac8 and that its autophagic roles are conserved.\" So yeast Vac8 papers are about the ortholog → INCLUDE.\n- Paper [5]: Bovine ARMC3 frameshift causing tail stump sperm defect → INCLUDE (ortholog, mammalian)\n- Paper [15]: Cancer antigen study - expression/TCR correlation, no mechanistic experiment → EXCLUDE\n- Paper [16]: Mouse ARMC3, directly about the mammalian gene → INCLUDE\n- Paper [17]: Human ARMC3 splicing variant, asthenozoospermia → INCLUDE\n- Paper [19]: Human ARMC3 stuttering → INCLUDE\n- Paper [20]: Human ARMC3 cloning/expression → borderline; describes splice variant isolation and tissue expression, no functional mechanism → EXCLUDE\n- Paper [24]: Erratum → EXCLUDE\n\nNow extracting mechanistic discoveries:\n\n```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1998,\n      \"finding\": \"Yeast Vac8 (ARMC3 ortholog) localizes to the vacuolar membrane via N-terminal myristylation; deletion of the myristylation site abolishes vacuolar localization. Loss of Vac8 causes accumulation of small fragmented vacuoles and defective vacuolar inheritance.\",\n      \"method\": \"GFP fusion live imaging, N-terminal myristylation mutant analysis, deletion genetics\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct localization experiment with functional consequence and mutagenesis, single lab\",\n      \"pmids\": [\"9664035\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Vac8 palmitoylation on isolated yeast vacuoles is mediated by the R-SNARE Ykt6 and is part of a SNARE subcomplex distinct from the Nyv1-containing complex; this reaction is ATP-independent, restricted to a narrow time window, and stimulated by EDTA (ion chelation). Palmitoylation is required for vacuole fusion.\",\n      \"method\": \"In vitro vacuole fusion assay with purified vacuoles, antibody inhibition, protein complex analysis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vitro reconstitution on purified vacuoles with biochemical complex analysis, single lab\",\n      \"pmids\": [\"15701652\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"The DHHC palmitoyl acyltransferase Pfa3 is specifically required for efficient vacuolar localization of Vac8 in vivo; Pfa3 deletion impairs Vac8 palmitoylation and reduces vacuole fusion, while vacuole morphology and inheritance appear normal.\",\n      \"method\": \"DHHC deletion mutant analysis, in vivo localization, vacuole fusion assay\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic deletion combined with localization and functional fusion assay, single lab\",\n      \"pmids\": [\"16301533\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Stable vacuolar membrane binding of Vac8 requires two N-terminal cysteines for palmitoylation regardless of their combination; palmitoylation adds functional roles beyond membrane anchoring — a basic-residue replacement mutant that still localizes to vacuoles can support cytoplasm-to-vacuole transport but requires at least one palmitoylation cysteine for vacuolar morphology and inheritance functions.\",\n      \"method\": \"Cysteine mutagenesis, vacuolar inheritance assay, Cvt pathway assay, fluorescence microscopy\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — systematic mutagenesis with multiple functional readouts, single lab\",\n      \"pmids\": [\"16720644\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"In Pichia pastoris, Vac8 ARM repeat domains (central region) are required for formation of vacuolar arm-like extensions that engulf peroxisomes during micropexophagy, and Vac8 is essential for recruitment of Atg11 to the vacuolar membrane during glucose-induced pexophagy; palmitoylation/myristoylation sites are required for protein stability and vacuolar association.\",\n      \"method\": \"Domain deletion mutants, GFP localization, pexophagy assay (alcohol oxidase activity), genetic knockouts\",\n      \"journal\": \"Autophagy\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — domain-deletion mutants with multiple functional readouts in Pichia pastoris, single lab\",\n      \"pmids\": [\"16921262\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"In Pichia pastoris, the ARM repeat domain of Vac8 is required for vacuolar inheritance but not for micropexophagy; deletion of both ARM and C-terminal domains abolishes vacuolar sequestering membrane formation and abolishes recruitment of Atg11 to the vacuolar membrane during micropexophagy.\",\n      \"method\": \"Domain deletion mutants, fluorescence microscopy, vacuolar inheritance assay\",\n      \"journal\": \"Autophagy\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — domain-specific deletion mutants with distinct functional readouts, single lab\",\n      \"pmids\": [\"16874085\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Pfa3 palmitoylates each of the three N-terminal cysteines of Vac8 in vitro, with efficiency enhanced by prior N-myristoylation; the 11th armadillo repeat of Vac8 is a key determinant for specific recognition by Pfa3, as shown by chimeric protein and competition experiments.\",\n      \"method\": \"In vitro palmitoylation assay, chimeric protein analysis, competition assay with truncation mutants\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution of palmitoylation with mutagenesis, chimeric proteins, and competition assays in single rigorous study\",\n      \"pmids\": [\"19416974\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Crystal structure of Vac8 bound to Atg13 reveals that the Atg13 extended loop (70 Å) binds the ARM domain of Vac8 in an antiparallel manner; the N-terminal H1 helix of Vac8 intramolecularly associates with ARM1 and regulates Vac8 self-association (dimerization), which is required differentially for Cvt and PMN autophagy pathways. Different quaternary structures of Vac8 (Atg13-bound heterotetramer vs. Nvj1-bound complex) mediate distinct autophagic functions.\",\n      \"method\": \"X-ray crystallography, analytical ultracentrifugation, isothermal titration calorimetry, SAXS, in vivo functional assays\",\n      \"journal\": \"Autophagy\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure combined with ITC, AUC, SAXS, and in vivo validation with multiple orthogonal methods\",\n      \"pmids\": [\"31512555\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"The Atg13 C-terminus binds lipid membranes via electrostatic interactions and hydrophobic insertion of a Phe residue; this phospholipid binding and Vac8 binding are mutually exclusive because they involve overlapping residues in the Atg13 IDR, and both interactions are required for efficient autophagy.\",\n      \"method\": \"Isothermal titration calorimetry, circular dichroism, lipid-binding assays, mutagenesis\",\n      \"journal\": \"Autophagy\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro binding assays with mutagenesis, single lab, mechanistically defines Vac8-Atg13 interaction as mutually exclusive with membrane binding\",\n      \"pmids\": [\"31352862\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Vac8 anchors the phagophore assembly site (PAS) to the vacuolar membrane by binding Atg13 and thereby recruiting the Atg1 initiation complex; VAC8 deletion or Vac8 mislocalization reduce autophagy activity, establishing Vac8 as required for correct vacuolar localization of the PAS.\",\n      \"method\": \"VAC8 deletion, auxin-inducible protein degradation (AID), fluorescence microscopy, autophagy activity assay\",\n      \"journal\": \"Autophagy\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — conditional deletion with fluorescence microscopy and functional autophagy readout, single lab\",\n      \"pmids\": [\"32508216\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"A 1 bp frameshift deletion (p.A451fs26) in bovine ARMC3 that truncates the protein by 401 amino acids (46%) is causally associated with a sterilizing tail stump sperm defect characterized by severely disorganized, immotile spermatozoa tails, establishing ARMC3 as required for normal spermatogenesis.\",\n      \"method\": \"Homozygosity mapping, whole-genome re-sequencing, segregation analysis in affected bulls\",\n      \"journal\": \"BMC genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — causal variant identification with segregation analysis in multiple affected animals, but no in vitro functional rescue\",\n      \"pmids\": [\"26923438\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Mouse ARMC3 is the functional homolog of yeast Vac8; its ARM domains recruit PtdIns3K-CI (class III PI3K complex I) to the phagophore assembly site to initiate autophagosome formation via PtdIns3P generation. Armc3 knockout mice show blocked ribophagy in spermatids, low mitochondrial energy levels, immotile flagella, and male infertility.\",\n      \"method\": \"Mouse knockout, ribophagy assay, mitochondrial function assay, yeast complementation studies\",\n      \"journal\": \"Autophagy\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — knockout mouse with multiple cellular phenotype readouts and cross-species complementation, single lab\",\n      \"pmids\": [\"34705610\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"X-ray crystal structure of the Vac8-Vac17 complex reveals a bipartite interaction interface; binding of Vac17 to Vac8 clamps the H1 helix to ARM1, preventing Vac8 dimerization and thereby competitively inhibiting Vac8 interactions with Nvj1 and Atg13. Mutation of key interface residues severely impairs vacuole inheritance in vivo.\",\n      \"method\": \"X-ray crystallography, mutagenesis, binding affinity measurements, in vivo vacuole inheritance assay\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure with mutagenesis and in vivo functional validation, mechanistically explaining how Vac8-mediated processes are mutually exclusive\",\n      \"pmids\": [\"37094131\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"PI3KCI interacts with the vacuolar membrane anchor Vac8 via the Atg14 C-terminal region in a constitutive manner; this interaction cooperates with Atg38-Atg1 complex and Vps30-Atg9 interactions to target PI3KCI to the PAS for autophagosome biogenesis.\",\n      \"method\": \"Co-immunoprecipitation, deletion mutant analysis, autophagy flux assays, genetic epistasis\",\n      \"journal\": \"The Journal of cell biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal co-IP with deletion mutants and functional autophagy readouts, single lab\",\n      \"pmids\": [\"37436710\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"A homozygous splicing variant (c.916+1G>A) in human ARMC3 causes exon 8 skipping, producing a truncated protein undetectable by Western blot in patient sperm, and results in asthenozoospermia with disrupted flagellar ultrastructure including vacuolated sperm mitochondria at the midpiece.\",\n      \"method\": \"Whole-exome sequencing, RT-PCR, Western blot, transmission electron microscopy of patient sperm\",\n      \"journal\": \"Clinical genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — patient variant with protein-level confirmation and TEM ultrastructural analysis, single family\",\n      \"pmids\": [\"39221575\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"The vacuole-specific adaptor Vac17 interacts with Myo2 (yeast myosin V) through two distinct binding sites (handhold mechanism); cryo-EM and structure prediction show one of these sites links to Vac8 on the vacuole membrane, forming the Myo2-Vac17-Vac8 complex for vacuole transport to daughter cells.\",\n      \"method\": \"Cryo-electron microscopy, AlphaFold structure prediction, genetic analysis, cell-based transport assays\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 2 / Weak — preprint, cryo-EM with structure prediction, single lab, no peer review\",\n      \"pmids\": [\"bio_10.1101_2025.03.24.645041\"],\n      \"is_preprint\": true\n    }\n  ],\n  \"current_model\": \"ARMC3 (mammalian) / Vac8 (yeast ortholog) is an armadillo-repeat protein anchored to the vacuolar/lysosomal membrane via N-terminal myristoylation and palmitoylation (performed by the DHHC enzyme Pfa3, with the 11th ARM repeat conferring substrate specificity); through its ARM domain it recruits PI3K complex I (via Atg14) and the Atg1 initiation complex (via Atg13) to the phagophore assembly site to initiate autophagy, while it also forms a tripartite Myo2-Vac17-Vac8 complex for myosin V-driven vacuole inheritance; distinct quaternary structures of Vac8 (monomer vs. dimer) differentially regulate these binding partners, ensuring vacuole inheritance, NVJ formation, and selective autophagy (Cvt, ribophagy, pexophagy) are mutually exclusive; in mammals, ARMC3 loss blocks ribophagy in spermatids, disrupts flagellar ultrastructure, and causes male infertility.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"ARMC3 (mammalian) and its yeast ortholog Vac8 are armadillo-repeat proteins that act as membrane-anchored scaffolds coupling the vacuolar/lysosomal membrane to autophagic and membrane-inheritance machinery [#7, #11]. Vac8 is targeted to the vacuolar membrane by N-terminal myristoylation, with deletion of the myristoylation site abolishing localization and producing fragmented vacuoles and inheritance defects [#0]; stable membrane attachment further requires palmitoylation of N-terminal cysteines, a modification installed by the DHHC acyltransferase Pfa3 — whose recognition of Vac8 is conferred specifically by the 11th armadillo repeat — and which adds functional roles beyond mere anchoring [#2, #3, #6]. Through its ARM domain, Vac8 anchors the phagophore assembly site to the vacuole by binding Atg13, thereby recruiting the Atg1 initiation complex, and additionally recruits class III PI3K complex I via the Atg14 C-terminus to drive autophagosome biogenesis [#7, #9, #13]. The same scaffold supports vacuole inheritance through a tripartite Myo2–Vac17–Vac8 complex [#15]. These activities are made mutually exclusive by the protein's quaternary state: an intramolecular H1-helix/ARM1 contact governs Vac8 self-association, and Vac17 binding clamps H1 to ARM1 to block dimerization and thereby competitively exclude the Nvj1 and Atg13 interactions [#7, #12]. In mammals, ARMC3 is the functional homolog of Vac8, with its ARM domains recruiting PtdIns3K-CI to initiate autophagosome formation; loss of ARMC3 blocks ribophagy in spermatids, lowers mitochondrial energy, and produces immotile flagella [#11]. Truncating ARMC3 mutations cause sterilizing sperm tail defects in cattle and asthenozoospermia with disrupted flagellar ultrastructure in humans, establishing ARMC3 as required for normal spermatogenesis [#10, #14].\",\n  \"teleology\": [\n    {\n      \"year\": 1998,\n      \"claim\": \"Established how Vac8 reaches its site of action and that it is functionally required for vacuole maintenance, answering where this protein works.\",\n      \"evidence\": \"GFP fusion live imaging and N-terminal myristylation mutant analysis with deletion genetics in yeast\",\n      \"pmids\": [\"9664035\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Did not resolve the molecular partners through which Vac8 directs inheritance\", \"Role of additional lipid modifications not yet defined\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Identified that Vac8 also undergoes palmitoylation and that this modification is required for vacuole fusion, distinguishing lipid anchoring from fusion function.\",\n      \"evidence\": \"In vitro vacuole fusion assay on purified yeast vacuoles with antibody inhibition and complex analysis; DHHC deletion mutant analysis identifying Pfa3\",\n      \"pmids\": [\"15701652\", \"16301533\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct enzyme-substrate mechanism of Pfa3 on Vac8 not yet reconstituted\", \"Whether palmitoylation has roles separable from fusion unresolved\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Dissected which Vac8 modifications and domains support which functions, separating membrane anchoring from inheritance, Cvt transport, and pexophagy.\",\n      \"evidence\": \"Cysteine and domain-deletion mutagenesis with vacuolar inheritance, Cvt, and pexophagy (alcohol oxidase) readouts in yeast and Pichia pastoris\",\n      \"pmids\": [\"16720644\", \"16921262\", \"16874085\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural basis of ARM-domain partner recognition not yet defined\", \"Identity of all ARM-domain effectors incomplete\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Defined the molecular specificity of Vac8 palmitoylation, showing N-myristoylation primes the reaction and the 11th ARM repeat dictates Pfa3 recognition.\",\n      \"evidence\": \"In vitro palmitoylation assay with chimeric proteins and competition/truncation mutants\",\n      \"pmids\": [\"19416974\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo dynamics of sequential modification not addressed\", \"Whether mammalian ARMC3 is similarly modified untested here\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Provided the structural logic of how Vac8 self-association and Atg13 binding underlie distinct autophagy pathways, explaining functional partitioning.\",\n      \"evidence\": \"X-ray crystallography of Vac8-Atg13, AUC, ITC, SAXS with in vivo assays; complementary ITC/CD lipid-binding analysis of Atg13\",\n      \"pmids\": [\"31512555\", \"31352862\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How upstream signals switch Vac8 quaternary state not defined\", \"Mammalian structural conservation untested\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Showed Vac8 anchors the phagophore assembly site to the vacuole by recruiting the Atg1 initiation complex via Atg13, defining its role in autophagy initiation.\",\n      \"evidence\": \"VAC8 deletion and auxin-inducible degradation with fluorescence microscopy and autophagy activity assays in yeast\",\n      \"pmids\": [\"32508216\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Quantitative contribution of Vac8 anchoring versus other PAS tethers unresolved\", \"Regulation of PAS positioning by signals not addressed\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Resolved how PI3K complex I is brought to the PAS through Vac8 and how Vac17 enforces mutual exclusivity among Vac8 functions.\",\n      \"evidence\": \"Co-IP with deletion mutants and autophagy flux assays for the Atg14-Vac8 interaction; crystal structure of Vac8-Vac17 with mutagenesis and in vivo inheritance assays\",\n      \"pmids\": [\"37436710\", \"37094131\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Temporal coordination of competing partner binding in vivo not mapped\", \"How the cell selects between inheritance and autophagy states unclear\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Established mammalian ARMC3 as the functional Vac8 homolog and linked its autophagy function to spermatogenesis, bridging yeast mechanism to mammalian physiology.\",\n      \"evidence\": \"Armc3 knockout mice with ribophagy and mitochondrial assays plus yeast complementation\",\n      \"pmids\": [\"34705610\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct biochemical demonstration of ARMC3 partner binding in mammalian cells limited\", \"Subcellular membrane anchor of ARMC3 in spermatids not defined\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Connected ARMC3 loss-of-function to human disease, showing truncating variants cause asthenozoospermia with flagellar and mitochondrial ultrastructural defects.\",\n      \"evidence\": \"Bovine homozygosity mapping/whole-genome sequencing of tail-stump defect; human whole-exome sequencing, RT-PCR, Western blot, and TEM of patient sperm\",\n      \"pmids\": [\"26923438\", \"39221575\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No functional rescue confirming causality in human cells\", \"Mechanism linking autophagy defect to flagellar disassembly not directly demonstrated\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Structurally defined the Myo2-Vac17-Vac8 transport complex, clarifying how Vac8 couples to myosin V for vacuole inheritance.\",\n      \"evidence\": \"Cryo-EM with AlphaFold prediction, genetics, and transport assays (preprint)\",\n      \"pmids\": [\"bio_10.1101_2025.03.24.645041\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Preprint, not peer reviewed\", \"Vac8-Vac17 linkage inferred partly from structure prediction\", \"Mammalian relevance of the transport complex untested\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How ARMC3-dependent autophagy is mechanistically required for mammalian flagellar assembly and what regulates the switch between its competing scaffolding states remains open.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No direct mammalian biochemistry of ARMC3 partner recruitment\", \"Signal controlling Vac8/ARMC3 quaternary state unknown\", \"Causal chain from ribophagy block to flagellar defect undefined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [7, 9, 11, 13]},\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [15]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005774\", \"supporting_discovery_ids\": [0, 2, 3]},\n      {\"term_id\": \"GO:0005773\", \"supporting_discovery_ids\": [0, 4, 5]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-9612973\", \"supporting_discovery_ids\": [7, 9, 11, 13]},\n      {\"term_id\": \"R-HSA-9609507\", \"supporting_discovery_ids\": [15]}\n    ],\n    \"complexes\": [\n      \"Myo2-Vac17-Vac8 vacuole inheritance complex\",\n      \"Vac8-Atg13-Atg1 initiation complex\",\n      \"Vac8-PI3K complex I (via Atg14)\"\n    ],\n    \"partners\": [\n      \"Atg13\",\n      \"Atg14\",\n      \"Vac17\",\n      \"Myo2\",\n      \"Nvj1\",\n      \"Pfa3\",\n      \"Ykt6\",\n      \"Atg11\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"tie","faith_supported":5,"faith_total":6,"faith_pct":83.33333333333333}}