{"gene":"ATG101","run_date":"2026-06-09T22:02:44","timeline":{"discoveries":[{"year":2009,"finding":"ATG101 (FLJ11773) is a novel mammalian Atg13-binding protein that associates with the ULK1-Atg13-FIP200 complex, most likely through direct interaction with Atg13. In Atg13 siRNA-treated cells, ATG101 is present solely as a monomer, indicating that complex incorporation depends on Atg13. ATG101 is important for the stability and basal phosphorylation of Atg13 and ULK1. GFP-ATG101 localizes to the isolation membrane/phagophore, and ATG101 knockdown suppresses GFP-LC3 dot formation and causes accumulation of LC3-I, establishing ATG101 as essential for autophagy initiation.","method":"Co-immunoprecipitation, siRNA knockdown, GFP-localization (live imaging), immunoblot for LC3 processing","journal":"Autophagy","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP and multiple orthogonal methods (localization, knockdown with defined phenotype, phosphorylation assay), independently replicated in a second concurrent paper (PMID:19287211)","pmids":["19597335"],"is_preprint":false},{"year":2009,"finding":"ATG101 (FLJ11773) interacts with ULK1 in an Atg13-dependent manner, stabilizes Atg13 expression by protecting it from proteasomal degradation, and is essential for macroautophagy. Intracellular localization of the ULK1 complex is regulated by nutrient conditions.","method":"Co-immunoprecipitation, siRNA knockdown, proteasome inhibitor rescue, autophagy flux assays","journal":"Autophagy","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP plus proteasomal rescue experiment, replicated independently (PMID:19597335)","pmids":["19287211"],"is_preprint":false},{"year":2012,"finding":"The C. elegans ATG101 homolog EPG-9 directly interacts with EPG-1/Atg13 and is essential for autophagic degradation of protein aggregates and starvation survival, placing ATG101 function in the Atg1/Atg13 pathway across metazoans.","method":"Genetic loss-of-function screen, direct protein interaction assay, autophagy flux assays in C. elegans","journal":"Autophagy","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis plus direct interaction assay in a model organism, single lab","pmids":["22885670"],"is_preprint":false},{"year":2014,"finding":"Drosophila Atg101 dimerizes and is predicted to fold into a HORMA domain. Loss of Atg101 impairs both starvation-induced and basal autophagy, leading to accumulation of ref(2)P/p62-positive aggregates. Mapping experiments show Atg101 binds the N-terminal HORMA domain of Atg13 and may also interact with two unstructured regions of Atg1. Atg101 also interacts with ref(2)P.","method":"Genetic loss-of-function (Drosophila mutant), domain-mapping pulldowns, dimerization assay, immunofluorescence for selective autophagy cargo","journal":"BioMed research international","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function with clear phenotypic readout plus domain-mapping in a model organism, single lab","pmids":["24895579"],"is_preprint":false},{"year":2015,"finding":"Crystal structure of the human Atg13 HORMA domain in complex with full-length ATG101 HORMA domain was determined. The two HORMA domains assemble with an architecture conserved in the Mad2 conformational heterodimer. The WF finger motif essential for ATG101 function is sequestered in a hydrophobic pocket, suggesting its exposure is regulated. Two benzamidine-marked hydrophobic pockets unique to animals suggest additional protein interaction sites, identifying the Atg13-ATG101 subcomplex as an interaction hub.","method":"X-ray crystallography, structural comparison, functional mapping of WF finger motif","journal":"Structure","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure with functional motif identification and structural conservation analysis across species","pmids":["26299944"],"is_preprint":false},{"year":2016,"finding":"Structural and cell biological analysis established that ATG101 is required for stabilization of 'uncapped' Atg13 in most eukaryotes (because Atg13 HORMA domain is exposed/uncapped without ATG101), and ATG101 recruits downstream Atg proteins through its WF motif. By contrast, S. cerevisiae Atg13 is stably 'capped' and does not require Atg101.","method":"Structural analysis, cell biology (reviewed synthesis of prior structural/functional studies)","journal":"Cell structure and function","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — synthesis of structural data with cell biological validation, single review paper integrating multiple prior experiments","pmids":["26754330"],"is_preprint":false},{"year":2018,"finding":"The C-terminal region of ATG101, which adopts a β-strand conformation in free ATG101 but a different conformation in the ATG101-ATG13HORMA complex, is responsible for binding PtdIns3K complex components (PIK3C3/VPS34, PIK3R4/VPS15, BECN1, UVRAG). C-terminal deletion of ATG101 shows significant defects in PtdIns3K interaction and impairs autophagosome formation, establishing ATG101 as a bridge between the ULK1 and PtdIns3K complexes.","method":"Crystal structure, SEC-SAXS, co-immunoprecipitation with deletion mutants, autophagosome formation assay, KO/reconstitution","journal":"Autophagy","confidence":"High","confidence_rationale":"Tier 1 / Strong — structural determination plus deletion mutagenesis plus Co-IP plus functional autophagy assay, single rigorous study with multiple orthogonal methods","pmids":["30081750"],"is_preprint":false},{"year":2018,"finding":"ATG101 physically interacts with the C-terminal domain (CTD) of the Hedgehog receptor PATCHED1 (PTCH1), connecting PTCH1 to the ULK complex. This interaction results in a blockade of basal autophagic flux and accumulation of autophagosomes with undegraded cargo, independent of PTCH1's repressive activity on SMO.","method":"Co-immunoprecipitation, autophagic flux assays, SMO-deficient cells and SMO inhibitor controls","journal":"Molecular cancer research","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Co-IP plus functional flux assay with multiple controls, single lab","pmids":["29453315"],"is_preprint":false},{"year":2019,"finding":"Drosophila Atg101 loss-of-function mutants are semi-lethal, with defective developmental and starvation-induced autophagy, accumulation of ubiquitin-positive aggregates in brains (neuronal defect), shortened/thickened midguts with enlarged enterocytes, and impaired differentiation of intestinal stem cells to enterocytes. Cell type-specific rescue showed ATG101 functions in enterocytes to limit their growth.","method":"Drosophila loss-of-function genetics, cell-type-specific rescue, immunofluorescence, lifespan and mobility assays","journal":"Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — clean loss-of-function with cell-type-specific rescue and multiple phenotypic readouts, single lab","pmids":["30760524"],"is_preprint":false},{"year":2021,"finding":"HUWE1 is the major E3 ubiquitin ligase targeting ATG101 for ubiquitination and proteasomal degradation, with the C-terminal region of ATG101 identified as the key ubiquitination domain. HUWE1 depletion stabilizes ATG101 and increases autophagy activity; this enhanced autophagy is reversed by siRNA-mediated ATG101 knockdown, placing ATG101 downstream of HUWE1 in autophagy regulation.","method":"CRISPR knockout, co-immunoprecipitation, siRNA knockdown, ubiquitination assays with C-terminal deletion mutants, autophagy flux assays","journal":"International journal of molecular sciences","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP plus domain mapping plus epistatic rescue, single lab with multiple orthogonal methods","pmids":["34502089"],"is_preprint":false},{"year":2021,"finding":"ATG9A interacts specifically with the ATG13-ATG101 dimer independently of ULK1, as demonstrated by knockout/reconstitution and split-mVenus approaches. Deletion of ATG13 or ATG101 causes aberrant accumulation of ATG9A at stalled p62/SQSTM1-ubiquitin clusters, rescuable by a ULK1 binding-deficient ATG13 mutant, establishing a ULK1-independent ATG13-ATG101 complex function in regulating ATG9A distribution.","method":"BioID quantitative proteomics, knockout/reconstitution, split-mVenus bimolecular complementation, immunofluorescence","journal":"EMBO reports","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (BioID, KO/reconstitution, split-mVenus, rescue with specific mutant), single lab with rigorous controls","pmids":["34369648"],"is_preprint":false},{"year":2025,"finding":"ATG101 HORMA domain forms a tight complex with PI3P-binding proteins WIPI3 and WIPI2. Bound to WIPI2/3, the ATG13:ATG101 dimer aligns with membranes to insert its WF (Trp-Phe) finger into the membrane. Molecular dynamics simulations show cooperative stabilization of the complex on membranes by WIPIs and the ATG101 WF finger. Biochemical reconstitution and cell-based assays show that WIPI3:ATG13 engagement is required for ATG16L1 phosphorylation by ULK1, ATG13 puncta formation, and bulk autophagic flux. A PVP motif in the ULK1 IDR docks onto the ATG13:ATG101 HORMA dimer surface, bringing the ULK1 kinase domain close to the membrane.","method":"Biochemical reconstitution, molecular dynamics simulation, in vitro kinase assay (ATG16L1 phosphorylation), cell-based autophagic flux assay, structural modeling","journal":"bioRxiv","confidence":"High","confidence_rationale":"Tier 1 / Strong — reconstitution plus MD simulation plus in vitro kinase assay plus cell-based validation with multiple orthogonal methods","pmids":["bio_10.1101_2025.11.07.687251"],"is_preprint":true},{"year":2025,"finding":"ATG101 HORMA domain undergoes a conformational change (fold change/metamorphosis) that enables interaction with ATG9A and ATG13 to form the essential ATG9A-ATG13-ATG101 complex. ATG101 homo-dimerization, initiated by ULK1 phosphorylation, dramatically accelerates complex formation. This creates an auto-catalytic positive feedback where ATG101 dimers propagate activation to further ATG101 molecules. Memory of ATG101 activation persists for many hours after dephosphorylation and continues to accelerate ATG9A-ATG13-ATG101 assembly.","method":"Interaction kinetics assays, phosphorylation assays, homodimerization assays, complex formation rate measurements","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple biochemical assays in single preprint lab, awaiting peer review","pmids":["bio_10.1101_2025.06.27.661946"],"is_preprint":true},{"year":2025,"finding":"ATG101 is required for the ATG13-ATG9 interaction in mammals but is dispensable for this interaction in Aspergillus oryzae, due to a shift in the AoAtg9-binding site in AoAtg13. Yeast two-hybrid assays established this species-specific dependency, and evolutionary analysis showed that ATG101 was lost in some Holomycota lineages after acquisition of Atg29/Atg31 and a cap structure in Atg13.","method":"Yeast two-hybrid assay, evolutionary BLAST analysis, genetic deletion (atg101 and atg31 in K. phaffii)","journal":"Autophagy","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — yeast two-hybrid plus genetic epistasis, single lab, peer-reviewed","pmids":["40931865"],"is_preprint":false}],"current_model":"ATG101 is a HORMA domain protein that forms a constitutive heterodimer with the Atg13 HORMA domain within the ULK1-Atg13-FIP200 complex; it stabilizes Atg13 against proteasomal degradation (regulated by HUWE1-mediated ubiquitination), recruits downstream autophagy proteins via its exposed WF finger that inserts into membranes in complex with WIPI2/3, bridges the ULK1 complex to the PtdIns3K complex through its C-terminal region, interacts with ATG9A independently of ULK1 to regulate ATG9A distribution, and undergoes ULK1 phosphorylation-dependent homo-dimerization that autocatalytically accelerates assembly of the ATG9A-ATG13-ATG101 initiation complex."},"narrative":{"mechanistic_narrative":"ATG101 is a HORMA-domain protein essential for autophagy initiation in metazoans, functioning as an integral subunit of the ULK1-Atg13-FIP200 complex that nucleates phagophore formation [PMID:19597335, PMID:19287211]. It associates with the complex through a constitutive heterodimer with the Atg13 HORMA domain, an interaction conserved from C. elegans (EPG-9/EPG-1) and Drosophila to humans, and assembles into an architecture analogous to the Mad2 conformational heterodimer [PMID:22885670, PMID:26299944]. A central function of ATG101 is to stabilize Atg13: it protects Atg13 from proteasomal degradation and caps the otherwise exposed Atg13 HORMA domain, and its own levels are controlled by HUWE1-mediated ubiquitination of its C-terminal region [PMID:19287211, PMID:26754330, PMID:34502089]. ATG101 acts as an interaction hub that couples the ULK1 complex to downstream autophagy machinery: its WF (Trp-Phe) finger, normally sequestered in a hydrophobic pocket, becomes exposed to recruit downstream factors and, together with PI3P-binding WIPI2/WIPI3 proteins, inserts into the membrane to position the ULK1 kinase for ATG16L1 phosphorylation [PMID:26299944, PMID:26754330, PMID:bio_10.1101_2025.11.07.687251]. Its C-terminal region bridges the ULK1 complex to the PtdIns3K complex (VPS34, VPS15, BECN1, UVRAG) [PMID:30081750]. Independently of ULK1, the ATG13-ATG101 dimer binds ATG9A to govern ATG9A distribution, and loss of ATG101 causes aberrant ATG9A accumulation at stalled cargo clusters [PMID:34369648]. Loss of ATG101 across model organisms impairs basal and starvation-induced autophagy, causing accumulation of ubiquitin/p62-positive aggregates and tissue-specific developmental defects [PMID:24895579, PMID:30760524]. ATG101 additionally links autophagy to other pathways through physical interaction with the Hedgehog receptor PTCH1, which blocks autophagic flux independently of SMO [PMID:29453315].","teleology":[{"year":2009,"claim":"Established ATG101 as a previously unknown core component of the mammalian autophagy initiation complex, answering whether the ULK1-Atg13-FIP200 machinery had additional essential subunits.","evidence":"Reciprocal Co-IP, siRNA knockdown with LC3 readout, GFP-localization to the phagophore, and proteasome-inhibitor rescue in mammalian cells (two concurrent papers)","pmids":["19597335","19287211"],"confidence":"High","gaps":["Structural basis of the Atg13-ATG101 interaction not defined","Mechanism by which ATG101 stabilizes Atg13 against degradation unresolved"]},{"year":2012,"claim":"Showed the Atg13-binding and autophagy function of ATG101 is conserved across metazoans, generalizing the mammalian finding beyond a single system.","evidence":"Genetic loss-of-function and direct interaction assays of EPG-9/EPG-1 in C. elegans","pmids":["22885670"],"confidence":"Medium","gaps":["Single-organism genetic data","No structural or biochemical mapping of the interaction interface"]},{"year":2014,"claim":"Identified ATG101 as a HORMA-domain protein that dimerizes and maps binding to the Atg13 HORMA domain, framing the molecular nature of the subcomplex.","evidence":"Drosophila loss-of-function genetics, domain-mapping pulldowns, and dimerization assays","pmids":["24895579"],"confidence":"Medium","gaps":["HORMA fold inferred by prediction, not solved","Functional significance of putative Atg1 interactions unclear"]},{"year":2015,"claim":"Solved the human Atg13HORMA-ATG101 heterodimer structure, revealing a Mad2-like architecture and a sequestered WF finger whose regulated exposure underlies downstream recruitment.","evidence":"X-ray crystallography with functional mapping of the WF finger motif","pmids":["26299944"],"confidence":"High","gaps":["Trigger for WF finger exposure not defined","Identity of downstream partners docking at the unique hydrophobic pockets unknown"]},{"year":2016,"claim":"Synthesized the structural data into a model explaining why ATG101 is required in most eukaryotes (to cap an exposed Atg13 HORMA) but dispensable in budding yeast, defining the evolutionary logic of its essentiality.","evidence":"Structural and cell-biological synthesis of prior work","pmids":["26754330"],"confidence":"Medium","gaps":["Review synthesis rather than new primary data","Direct test of the capping model in diverse species not provided"]},{"year":2018,"claim":"Defined how ATG101 physically couples autophagy initiation to lipid-kinase activation by mapping its C-terminal region to PtdIns3K binding.","evidence":"Crystal structure, SEC-SAXS, Co-IP with deletion mutants, and autophagosome formation assays with KO/reconstitution","pmids":["30081750"],"confidence":"High","gaps":["Conformational switch driving C-terminal availability not kinetically resolved","Stoichiometry of the ULK1-PtdIns3K bridge unknown"]},{"year":2018,"claim":"Extended ATG101 function beyond core autophagy by showing it physically links the Hedgehog receptor PTCH1 to the ULK complex to restrain autophagic flux.","evidence":"Co-IP and autophagic flux assays with SMO-deficient cells and SMO inhibitor controls","pmids":["29453315"],"confidence":"Medium","gaps":["Interaction interface on ATG101 not mapped","Mechanism by which PTCH1 binding blocks flux unresolved"]},{"year":2019,"claim":"Demonstrated physiological consequences of ATG101 loss in a whole organism, including neuronal aggregate accumulation and cell-type-specific roles in tissue homeostasis.","evidence":"Drosophila loss-of-function genetics with cell-type-specific rescue and phenotypic assays","pmids":["30760524"],"confidence":"Medium","gaps":["Molecular basis of tissue-specific requirements not defined","Mammalian in vivo phenotype not addressed"]},{"year":2021,"claim":"Identified HUWE1 as the E3 ligase controlling ATG101 turnover via its C-terminal region, establishing a ubiquitin-dependent layer of autophagy regulation acting through ATG101 levels.","evidence":"CRISPR knockout, Co-IP, ubiquitination assays with C-terminal deletion mutants, and epistatic siRNA rescue","pmids":["34502089"],"confidence":"Medium","gaps":["Signals regulating HUWE1-ATG101 activity unknown","Single-lab data"]},{"year":2021,"claim":"Uncovered a ULK1-independent function of the ATG13-ATG101 dimer in binding ATG9A and controlling its distribution, separating ATG101's roles within and outside ULK1 kinase signaling.","evidence":"BioID proteomics, KO/reconstitution, split-mVenus complementation, and rescue with a ULK1-binding-deficient ATG13 mutant","pmids":["34369648"],"confidence":"High","gaps":["Structural basis of ATG9A engagement not defined at this stage","How ATG9A trafficking is mechanistically corrected unclear"]},{"year":2025,"claim":"Reconstituted how the ATG13:ATG101 dimer engages PI3P-binding WIPI2/3 and inserts its WF finger into membranes to position ULK1 for ATG16L1 phosphorylation, linking membrane targeting to downstream conjugation machinery.","evidence":"Biochemical reconstitution, molecular dynamics, in vitro kinase assay, and cell-based flux assays (preprint)","pmids":["bio_10.1101_2025.11.07.687251"],"confidence":"High","gaps":["Peer review pending","In vivo relevance of the membrane-insertion model not yet tested"]},{"year":2025,"claim":"Proposed an autocatalytic mechanism in which ULK1-phosphorylation-driven ATG101 homodimerization accelerates assembly of an ATG9A-ATG13-ATG101 initiation complex, providing a kinetic switch and memory for autophagy onset.","evidence":"Interaction kinetics, phosphorylation and homodimerization assays, and complex formation rate measurements (preprint)","pmids":["bio_10.1101_2025.06.27.661946"],"confidence":"Medium","gaps":["Preprint awaiting peer review","Physiological role of the proposed activation memory not established","Structural basis of the metamorphic HORMA change not solved"]},{"year":2025,"claim":"Defined the species-specific basis for ATG101's requirement in the ATG13-ATG9 interaction, clarifying why ATG101 was retained in metazoans but lost in some fungal lineages.","evidence":"Yeast two-hybrid assays, evolutionary BLAST analysis, and genetic deletions in K. phaffii","pmids":["40931865"],"confidence":"Medium","gaps":["Mechanism of the Atg13 binding-site shift not structurally resolved","Selective pressures driving ATG101 loss inferred, not demonstrated"]},{"year":null,"claim":"How the regulated exposure of the WF finger, the C-terminal conformational switch, and ATG101 homodimerization are coordinated in time to control autophagy initiation in vivo remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No integrated kinetic model linking phosphorylation, dimerization, and membrane engagement","Mammalian in vivo loss-of-function phenotype not characterized in the corpus","Disease relevance not established in the corpus"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[0,4,6]},{"term_id":"GO:0008289","term_label":"lipid binding","supporting_discovery_ids":[11]},{"term_id":"GO:0140313","term_label":"molecular sequestering activity","supporting_discovery_ids":[1,5]}],"localization":[{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[0]}],"pathway":[{"term_id":"R-HSA-9612973","term_label":"Autophagy","supporting_discovery_ids":[0,1,6,10]}],"complexes":["ULK1-ATG13-FIP200 complex","ATG13-ATG101 HORMA heterodimer","ATG9A-ATG13-ATG101 complex"],"partners":["ATG13","ULK1","ATG9A","WIPI2","WIPI3","HUWE1","PTCH1","PIK3C3"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9BSB4","full_name":"Autophagy-related protein 101","aliases":[],"length_aa":218,"mass_kda":25.0,"function":"Autophagy factor required for autophagosome formation. Stabilizes ATG13, protecting it from proteasomal degradation","subcellular_location":"Cytoplasm; Preautophagosomal structure","url":"https://www.uniprot.org/uniprotkb/Q9BSB4/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/ATG101","classification":"Not Classified","n_dependent_lines":37,"n_total_lines":1208,"dependency_fraction":0.030629139072847682},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000123395","cell_line_id":"CID001806","localizations":[{"compartment":"cytoplasmic","grade":3},{"compartment":"vesicles","grade":3},{"compartment":"nucleoplasm","grade":1}],"interactors":[{"gene":"ATG13","stoichiometry":10.0},{"gene":"ULK1","stoichiometry":10.0},{"gene":"RB1CC1","stoichiometry":4.0},{"gene":"HSBP1","stoichiometry":0.2},{"gene":"EMC8","stoichiometry":0.2},{"gene":"CYP51A1","stoichiometry":0.2},{"gene":"RBMX;RBMXL2","stoichiometry":0.2},{"gene":"PRMT1","stoichiometry":0.2},{"gene":"DOCK6","stoichiometry":0.2},{"gene":"CCPG1","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/target/CID001806","total_profiled":1310},"omim":[{"mim_id":"617074","title":"SMITH-MAGENIS SYNDROME CHROMOSOME REGION, CANDIDATE GENE 8; SMCR8","url":"https://www.omim.org/entry/617074"},{"mim_id":"615089","title":"AUTOPHAGY-RELATED PROTEIN 101; ATG101","url":"https://www.omim.org/entry/615089"},{"mim_id":"615088","title":"AUTOPHAGY-RELATED 13; ATG13","url":"https://www.omim.org/entry/615088"},{"mim_id":"614260","title":"CHROMOSOME 9 OPEN READING FRAME 72; C9ORF72","url":"https://www.omim.org/entry/614260"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/ATG101"},"hgnc":{"alias_symbol":["FLJ11773"],"prev_symbol":["C12orf44"]},"alphafold":{"accession":"Q9BSB4","domains":[{"cath_id":"3.30.900","chopping":"3-208","consensus_level":"high","plddt":92.7569,"start":3,"end":208}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9BSB4","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9BSB4-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9BSB4-F1-predicted_aligned_error_v6.png","plddt_mean":90.12},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=ATG101","jax_strain_url":"https://www.jax.org/strain/search?query=ATG101"},"sequence":{"accession":"Q9BSB4","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9BSB4.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9BSB4/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9BSB4"}},"corpus_meta":[{"pmid":"19597335","id":"PMC_19597335","title":"Atg101, a novel mammalian autophagy protein interacting with Atg13.","date":"2009","source":"Autophagy","url":"https://pubmed.ncbi.nlm.nih.gov/19597335","citation_count":397,"is_preprint":false},{"pmid":"19287211","id":"PMC_19287211","title":"A novel, human Atg13 binding protein, Atg101, interacts with ULK1 and is essential for macroautophagy.","date":"2009","source":"Autophagy","url":"https://pubmed.ncbi.nlm.nih.gov/19287211","citation_count":338,"is_preprint":false},{"pmid":"26299944","id":"PMC_26299944","title":"Structure of the Human Atg13-Atg101 HORMA Heterodimer: an Interaction Hub within the ULK1 Complex.","date":"2015","source":"Structure (London, England : 1993)","url":"https://pubmed.ncbi.nlm.nih.gov/26299944","citation_count":81,"is_preprint":false},{"pmid":"30081750","id":"PMC_30081750","title":"The C-terminal region of ATG101 bridges ULK1 and PtdIns3K complex in autophagy initiation.","date":"2018","source":"Autophagy","url":"https://pubmed.ncbi.nlm.nih.gov/30081750","citation_count":51,"is_preprint":false},{"pmid":"22885670","id":"PMC_22885670","title":"The C. elegans ATG101 homolog EPG-9 directly interacts with EPG-1/Atg13 and is essential for autophagy.","date":"2012","source":"Autophagy","url":"https://pubmed.ncbi.nlm.nih.gov/22885670","citation_count":46,"is_preprint":false},{"pmid":"34369648","id":"PMC_34369648","title":"BioID reveals an ATG9A interaction with ATG13-ATG101 in the degradation of p62/SQSTM1-ubiquitin clusters.","date":"2021","source":"EMBO reports","url":"https://pubmed.ncbi.nlm.nih.gov/34369648","citation_count":42,"is_preprint":false},{"pmid":"30760524","id":"PMC_30760524","title":"The autophagy-related gene Atg101 in Drosophila regulates both neuron and midgut homeostasis.","date":"2019","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/30760524","citation_count":38,"is_preprint":false},{"pmid":"24895579","id":"PMC_24895579","title":"The putative HORMA domain protein Atg101 dimerizes and is required for starvation-induced and selective autophagy in Drosophila.","date":"2014","source":"BioMed research international","url":"https://pubmed.ncbi.nlm.nih.gov/24895579","citation_count":38,"is_preprint":false},{"pmid":"33027196","id":"PMC_33027196","title":"Circ-HIPK2 Accelerates Cell Apoptosis and Autophagy in Myocardial Oxidative Injury by Sponging miR-485-5p and Targeting ATG101.","date":"2020","source":"Journal of cardiovascular pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/33027196","citation_count":28,"is_preprint":false},{"pmid":"26754330","id":"PMC_26754330","title":"Atg101: Not Just an Accessory Subunit in the Autophagy-initiation Complex.","date":"2016","source":"Cell structure and 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interfaces","url":"https://pubmed.ncbi.nlm.nih.gov/29154530","citation_count":17,"is_preprint":false},{"pmid":"40487652","id":"PMC_40487652","title":"First ATG101-recruiting small molecule degrader for selective CDK9 degradation via autophagy-lysosome pathway.","date":"2025","source":"Acta pharmaceutica Sinica. 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In Atg13 siRNA-treated cells, ATG101 is present solely as a monomer, indicating that complex incorporation depends on Atg13. ATG101 is important for the stability and basal phosphorylation of Atg13 and ULK1. GFP-ATG101 localizes to the isolation membrane/phagophore, and ATG101 knockdown suppresses GFP-LC3 dot formation and causes accumulation of LC3-I, establishing ATG101 as essential for autophagy initiation.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, GFP-localization (live imaging), immunoblot for LC3 processing\",\n      \"journal\": \"Autophagy\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP and multiple orthogonal methods (localization, knockdown with defined phenotype, phosphorylation assay), independently replicated in a second concurrent paper (PMID:19287211)\",\n      \"pmids\": [\"19597335\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"ATG101 (FLJ11773) interacts with ULK1 in an Atg13-dependent manner, stabilizes Atg13 expression by protecting it from proteasomal degradation, and is essential for macroautophagy. Intracellular localization of the ULK1 complex is regulated by nutrient conditions.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, proteasome inhibitor rescue, autophagy flux assays\",\n      \"journal\": \"Autophagy\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP plus proteasomal rescue experiment, replicated independently (PMID:19597335)\",\n      \"pmids\": [\"19287211\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"The C. elegans ATG101 homolog EPG-9 directly interacts with EPG-1/Atg13 and is essential for autophagic degradation of protein aggregates and starvation survival, placing ATG101 function in the Atg1/Atg13 pathway across metazoans.\",\n      \"method\": \"Genetic loss-of-function screen, direct protein interaction assay, autophagy flux assays in C. elegans\",\n      \"journal\": \"Autophagy\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis plus direct interaction assay in a model organism, single lab\",\n      \"pmids\": [\"22885670\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Drosophila Atg101 dimerizes and is predicted to fold into a HORMA domain. Loss of Atg101 impairs both starvation-induced and basal autophagy, leading to accumulation of ref(2)P/p62-positive aggregates. Mapping experiments show Atg101 binds the N-terminal HORMA domain of Atg13 and may also interact with two unstructured regions of Atg1. Atg101 also interacts with ref(2)P.\",\n      \"method\": \"Genetic loss-of-function (Drosophila mutant), domain-mapping pulldowns, dimerization assay, immunofluorescence for selective autophagy cargo\",\n      \"journal\": \"BioMed research international\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function with clear phenotypic readout plus domain-mapping in a model organism, single lab\",\n      \"pmids\": [\"24895579\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Crystal structure of the human Atg13 HORMA domain in complex with full-length ATG101 HORMA domain was determined. The two HORMA domains assemble with an architecture conserved in the Mad2 conformational heterodimer. The WF finger motif essential for ATG101 function is sequestered in a hydrophobic pocket, suggesting its exposure is regulated. Two benzamidine-marked hydrophobic pockets unique to animals suggest additional protein interaction sites, identifying the Atg13-ATG101 subcomplex as an interaction hub.\",\n      \"method\": \"X-ray crystallography, structural comparison, functional mapping of WF finger motif\",\n      \"journal\": \"Structure\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure with functional motif identification and structural conservation analysis across species\",\n      \"pmids\": [\"26299944\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Structural and cell biological analysis established that ATG101 is required for stabilization of 'uncapped' Atg13 in most eukaryotes (because Atg13 HORMA domain is exposed/uncapped without ATG101), and ATG101 recruits downstream Atg proteins through its WF motif. By contrast, S. cerevisiae Atg13 is stably 'capped' and does not require Atg101.\",\n      \"method\": \"Structural analysis, cell biology (reviewed synthesis of prior structural/functional studies)\",\n      \"journal\": \"Cell structure and function\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — synthesis of structural data with cell biological validation, single review paper integrating multiple prior experiments\",\n      \"pmids\": [\"26754330\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"The C-terminal region of ATG101, which adopts a β-strand conformation in free ATG101 but a different conformation in the ATG101-ATG13HORMA complex, is responsible for binding PtdIns3K complex components (PIK3C3/VPS34, PIK3R4/VPS15, BECN1, UVRAG). C-terminal deletion of ATG101 shows significant defects in PtdIns3K interaction and impairs autophagosome formation, establishing ATG101 as a bridge between the ULK1 and PtdIns3K complexes.\",\n      \"method\": \"Crystal structure, SEC-SAXS, co-immunoprecipitation with deletion mutants, autophagosome formation assay, KO/reconstitution\",\n      \"journal\": \"Autophagy\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — structural determination plus deletion mutagenesis plus Co-IP plus functional autophagy assay, single rigorous study with multiple orthogonal methods\",\n      \"pmids\": [\"30081750\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"ATG101 physically interacts with the C-terminal domain (CTD) of the Hedgehog receptor PATCHED1 (PTCH1), connecting PTCH1 to the ULK complex. This interaction results in a blockade of basal autophagic flux and accumulation of autophagosomes with undegraded cargo, independent of PTCH1's repressive activity on SMO.\",\n      \"method\": \"Co-immunoprecipitation, autophagic flux assays, SMO-deficient cells and SMO inhibitor controls\",\n      \"journal\": \"Molecular cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Co-IP plus functional flux assay with multiple controls, single lab\",\n      \"pmids\": [\"29453315\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Drosophila Atg101 loss-of-function mutants are semi-lethal, with defective developmental and starvation-induced autophagy, accumulation of ubiquitin-positive aggregates in brains (neuronal defect), shortened/thickened midguts with enlarged enterocytes, and impaired differentiation of intestinal stem cells to enterocytes. Cell type-specific rescue showed ATG101 functions in enterocytes to limit their growth.\",\n      \"method\": \"Drosophila loss-of-function genetics, cell-type-specific rescue, immunofluorescence, lifespan and mobility assays\",\n      \"journal\": \"Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — clean loss-of-function with cell-type-specific rescue and multiple phenotypic readouts, single lab\",\n      \"pmids\": [\"30760524\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"HUWE1 is the major E3 ubiquitin ligase targeting ATG101 for ubiquitination and proteasomal degradation, with the C-terminal region of ATG101 identified as the key ubiquitination domain. HUWE1 depletion stabilizes ATG101 and increases autophagy activity; this enhanced autophagy is reversed by siRNA-mediated ATG101 knockdown, placing ATG101 downstream of HUWE1 in autophagy regulation.\",\n      \"method\": \"CRISPR knockout, co-immunoprecipitation, siRNA knockdown, ubiquitination assays with C-terminal deletion mutants, autophagy flux assays\",\n      \"journal\": \"International journal of molecular sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP plus domain mapping plus epistatic rescue, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"34502089\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"ATG9A interacts specifically with the ATG13-ATG101 dimer independently of ULK1, as demonstrated by knockout/reconstitution and split-mVenus approaches. Deletion of ATG13 or ATG101 causes aberrant accumulation of ATG9A at stalled p62/SQSTM1-ubiquitin clusters, rescuable by a ULK1 binding-deficient ATG13 mutant, establishing a ULK1-independent ATG13-ATG101 complex function in regulating ATG9A distribution.\",\n      \"method\": \"BioID quantitative proteomics, knockout/reconstitution, split-mVenus bimolecular complementation, immunofluorescence\",\n      \"journal\": \"EMBO reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (BioID, KO/reconstitution, split-mVenus, rescue with specific mutant), single lab with rigorous controls\",\n      \"pmids\": [\"34369648\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"ATG101 HORMA domain forms a tight complex with PI3P-binding proteins WIPI3 and WIPI2. Bound to WIPI2/3, the ATG13:ATG101 dimer aligns with membranes to insert its WF (Trp-Phe) finger into the membrane. Molecular dynamics simulations show cooperative stabilization of the complex on membranes by WIPIs and the ATG101 WF finger. Biochemical reconstitution and cell-based assays show that WIPI3:ATG13 engagement is required for ATG16L1 phosphorylation by ULK1, ATG13 puncta formation, and bulk autophagic flux. A PVP motif in the ULK1 IDR docks onto the ATG13:ATG101 HORMA dimer surface, bringing the ULK1 kinase domain close to the membrane.\",\n      \"method\": \"Biochemical reconstitution, molecular dynamics simulation, in vitro kinase assay (ATG16L1 phosphorylation), cell-based autophagic flux assay, structural modeling\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — reconstitution plus MD simulation plus in vitro kinase assay plus cell-based validation with multiple orthogonal methods\",\n      \"pmids\": [\"bio_10.1101_2025.11.07.687251\"],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"ATG101 HORMA domain undergoes a conformational change (fold change/metamorphosis) that enables interaction with ATG9A and ATG13 to form the essential ATG9A-ATG13-ATG101 complex. ATG101 homo-dimerization, initiated by ULK1 phosphorylation, dramatically accelerates complex formation. This creates an auto-catalytic positive feedback where ATG101 dimers propagate activation to further ATG101 molecules. Memory of ATG101 activation persists for many hours after dephosphorylation and continues to accelerate ATG9A-ATG13-ATG101 assembly.\",\n      \"method\": \"Interaction kinetics assays, phosphorylation assays, homodimerization assays, complex formation rate measurements\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple biochemical assays in single preprint lab, awaiting peer review\",\n      \"pmids\": [\"bio_10.1101_2025.06.27.661946\"],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"ATG101 is required for the ATG13-ATG9 interaction in mammals but is dispensable for this interaction in Aspergillus oryzae, due to a shift in the AoAtg9-binding site in AoAtg13. Yeast two-hybrid assays established this species-specific dependency, and evolutionary analysis showed that ATG101 was lost in some Holomycota lineages after acquisition of Atg29/Atg31 and a cap structure in Atg13.\",\n      \"method\": \"Yeast two-hybrid assay, evolutionary BLAST analysis, genetic deletion (atg101 and atg31 in K. phaffii)\",\n      \"journal\": \"Autophagy\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — yeast two-hybrid plus genetic epistasis, single lab, peer-reviewed\",\n      \"pmids\": [\"40931865\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"ATG101 is a HORMA domain protein that forms a constitutive heterodimer with the Atg13 HORMA domain within the ULK1-Atg13-FIP200 complex; it stabilizes Atg13 against proteasomal degradation (regulated by HUWE1-mediated ubiquitination), recruits downstream autophagy proteins via its exposed WF finger that inserts into membranes in complex with WIPI2/3, bridges the ULK1 complex to the PtdIns3K complex through its C-terminal region, interacts with ATG9A independently of ULK1 to regulate ATG9A distribution, and undergoes ULK1 phosphorylation-dependent homo-dimerization that autocatalytically accelerates assembly of the ATG9A-ATG13-ATG101 initiation complex.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"ATG101 is a HORMA-domain protein essential for autophagy initiation in metazoans, functioning as an integral subunit of the ULK1-Atg13-FIP200 complex that nucleates phagophore formation [#0, #1]. It associates with the complex through a constitutive heterodimer with the Atg13 HORMA domain, an interaction conserved from C. elegans (EPG-9/EPG-1) and Drosophila to humans, and assembles into an architecture analogous to the Mad2 conformational heterodimer [#2, #4]. A central function of ATG101 is to stabilize Atg13: it protects Atg13 from proteasomal degradation and caps the otherwise exposed Atg13 HORMA domain, and its own levels are controlled by HUWE1-mediated ubiquitination of its C-terminal region [#1, #5, #9]. ATG101 acts as an interaction hub that couples the ULK1 complex to downstream autophagy machinery: its WF (Trp-Phe) finger, normally sequestered in a hydrophobic pocket, becomes exposed to recruit downstream factors and, together with PI3P-binding WIPI2/WIPI3 proteins, inserts into the membrane to position the ULK1 kinase for ATG16L1 phosphorylation [#4, #5, #11]. Its C-terminal region bridges the ULK1 complex to the PtdIns3K complex (VPS34, VPS15, BECN1, UVRAG) [#6]. Independently of ULK1, the ATG13-ATG101 dimer binds ATG9A to govern ATG9A distribution, and loss of ATG101 causes aberrant ATG9A accumulation at stalled cargo clusters [#10]. Loss of ATG101 across model organisms impairs basal and starvation-induced autophagy, causing accumulation of ubiquitin/p62-positive aggregates and tissue-specific developmental defects [#3, #8]. ATG101 additionally links autophagy to other pathways through physical interaction with the Hedgehog receptor PTCH1, which blocks autophagic flux independently of SMO [#7].\",\n  \"teleology\": [\n    {\n      \"year\": 2009,\n      \"claim\": \"Established ATG101 as a previously unknown core component of the mammalian autophagy initiation complex, answering whether the ULK1-Atg13-FIP200 machinery had additional essential subunits.\",\n      \"evidence\": \"Reciprocal Co-IP, siRNA knockdown with LC3 readout, GFP-localization to the phagophore, and proteasome-inhibitor rescue in mammalian cells (two concurrent papers)\",\n      \"pmids\": [\"19597335\", \"19287211\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of the Atg13-ATG101 interaction not defined\", \"Mechanism by which ATG101 stabilizes Atg13 against degradation unresolved\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Showed the Atg13-binding and autophagy function of ATG101 is conserved across metazoans, generalizing the mammalian finding beyond a single system.\",\n      \"evidence\": \"Genetic loss-of-function and direct interaction assays of EPG-9/EPG-1 in C. elegans\",\n      \"pmids\": [\"22885670\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-organism genetic data\", \"No structural or biochemical mapping of the interaction interface\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Identified ATG101 as a HORMA-domain protein that dimerizes and maps binding to the Atg13 HORMA domain, framing the molecular nature of the subcomplex.\",\n      \"evidence\": \"Drosophila loss-of-function genetics, domain-mapping pulldowns, and dimerization assays\",\n      \"pmids\": [\"24895579\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"HORMA fold inferred by prediction, not solved\", \"Functional significance of putative Atg1 interactions unclear\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Solved the human Atg13HORMA-ATG101 heterodimer structure, revealing a Mad2-like architecture and a sequestered WF finger whose regulated exposure underlies downstream recruitment.\",\n      \"evidence\": \"X-ray crystallography with functional mapping of the WF finger motif\",\n      \"pmids\": [\"26299944\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Trigger for WF finger exposure not defined\", \"Identity of downstream partners docking at the unique hydrophobic pockets unknown\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Synthesized the structural data into a model explaining why ATG101 is required in most eukaryotes (to cap an exposed Atg13 HORMA) but dispensable in budding yeast, defining the evolutionary logic of its essentiality.\",\n      \"evidence\": \"Structural and cell-biological synthesis of prior work\",\n      \"pmids\": [\"26754330\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Review synthesis rather than new primary data\", \"Direct test of the capping model in diverse species not provided\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Defined how ATG101 physically couples autophagy initiation to lipid-kinase activation by mapping its C-terminal region to PtdIns3K binding.\",\n      \"evidence\": \"Crystal structure, SEC-SAXS, Co-IP with deletion mutants, and autophagosome formation assays with KO/reconstitution\",\n      \"pmids\": [\"30081750\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Conformational switch driving C-terminal availability not kinetically resolved\", \"Stoichiometry of the ULK1-PtdIns3K bridge unknown\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Extended ATG101 function beyond core autophagy by showing it physically links the Hedgehog receptor PTCH1 to the ULK complex to restrain autophagic flux.\",\n      \"evidence\": \"Co-IP and autophagic flux assays with SMO-deficient cells and SMO inhibitor controls\",\n      \"pmids\": [\"29453315\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Interaction interface on ATG101 not mapped\", \"Mechanism by which PTCH1 binding blocks flux unresolved\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Demonstrated physiological consequences of ATG101 loss in a whole organism, including neuronal aggregate accumulation and cell-type-specific roles in tissue homeostasis.\",\n      \"evidence\": \"Drosophila loss-of-function genetics with cell-type-specific rescue and phenotypic assays\",\n      \"pmids\": [\"30760524\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular basis of tissue-specific requirements not defined\", \"Mammalian in vivo phenotype not addressed\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Identified HUWE1 as the E3 ligase controlling ATG101 turnover via its C-terminal region, establishing a ubiquitin-dependent layer of autophagy regulation acting through ATG101 levels.\",\n      \"evidence\": \"CRISPR knockout, Co-IP, ubiquitination assays with C-terminal deletion mutants, and epistatic siRNA rescue\",\n      \"pmids\": [\"34502089\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Signals regulating HUWE1-ATG101 activity unknown\", \"Single-lab data\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Uncovered a ULK1-independent function of the ATG13-ATG101 dimer in binding ATG9A and controlling its distribution, separating ATG101's roles within and outside ULK1 kinase signaling.\",\n      \"evidence\": \"BioID proteomics, KO/reconstitution, split-mVenus complementation, and rescue with a ULK1-binding-deficient ATG13 mutant\",\n      \"pmids\": [\"34369648\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of ATG9A engagement not defined at this stage\", \"How ATG9A trafficking is mechanistically corrected unclear\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Reconstituted how the ATG13:ATG101 dimer engages PI3P-binding WIPI2/3 and inserts its WF finger into membranes to position ULK1 for ATG16L1 phosphorylation, linking membrane targeting to downstream conjugation machinery.\",\n      \"evidence\": \"Biochemical reconstitution, molecular dynamics, in vitro kinase assay, and cell-based flux assays (preprint)\",\n      \"pmids\": [\"bio_10.1101_2025.11.07.687251\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Peer review pending\", \"In vivo relevance of the membrane-insertion model not yet tested\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Proposed an autocatalytic mechanism in which ULK1-phosphorylation-driven ATG101 homodimerization accelerates assembly of an ATG9A-ATG13-ATG101 initiation complex, providing a kinetic switch and memory for autophagy onset.\",\n      \"evidence\": \"Interaction kinetics, phosphorylation and homodimerization assays, and complex formation rate measurements (preprint)\",\n      \"pmids\": [\"bio_10.1101_2025.06.27.661946\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Preprint awaiting peer review\", \"Physiological role of the proposed activation memory not established\", \"Structural basis of the metamorphic HORMA change not solved\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Defined the species-specific basis for ATG101's requirement in the ATG13-ATG9 interaction, clarifying why ATG101 was retained in metazoans but lost in some fungal lineages.\",\n      \"evidence\": \"Yeast two-hybrid assays, evolutionary BLAST analysis, and genetic deletions in K. phaffii\",\n      \"pmids\": [\"40931865\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism of the Atg13 binding-site shift not structurally resolved\", \"Selective pressures driving ATG101 loss inferred, not demonstrated\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How the regulated exposure of the WF finger, the C-terminal conformational switch, and ATG101 homodimerization are coordinated in time to control autophagy initiation in vivo remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No integrated kinetic model linking phosphorylation, dimerization, and membrane engagement\", \"Mammalian in vivo loss-of-function phenotype not characterized in the corpus\", \"Disease relevance not established in the corpus\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [0, 4, 6]},\n      {\"term_id\": \"GO:0008289\", \"supporting_discovery_ids\": [11]},\n      {\"term_id\": \"GO:0140313\", \"supporting_discovery_ids\": [1, 5]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [0]},\n      {\"term_id\": \"GO:0005776\", \"supporting_discovery_ids\": [0]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-9612973\", \"supporting_discovery_ids\": [0, 1, 6, 10]}\n    ],\n    \"complexes\": [\n      \"ULK1-ATG13-FIP200 complex\",\n      \"ATG13-ATG101 HORMA heterodimer\",\n      \"ATG9A-ATG13-ATG101 complex\"\n    ],\n    \"partners\": [\n      \"ATG13\",\n      \"ULK1\",\n      \"ATG9A\",\n      \"WIPI2\",\n      \"WIPI3\",\n      \"HUWE1\",\n      \"PTCH1\",\n      \"PIK3C3\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":8,"faith_total":8,"faith_pct":100.0}}