id: Q96BY7 gene_symbol: ATG2B product_type: PROTEIN status: COMPLETE taxon: id: NCBITaxon:9606 label: Homo sapiens description: >- ATG2B is a large conserved autophagy-related lipid-transfer protein that acts at endoplasmic reticulum-phagophore membrane contact sites during autophagosome biogenesis. It binds membranes and forms an ATG2-WIPI/ATG18 complex with WDR45/WIPI4, which promotes ATG2B recruitment to PI3P-containing autophagic membranes and stimulates lipid-transfer activity. Mammalian ATG2 proteins are also associated with lipid droplets and affect lipid droplet morphology and dispersion, but the best-supported core role of ATG2B is membrane tethering and lipid transfer for phagophore expansion. existing_annotations: - term: id: GO:0000407 label: phagophore assembly site evidence_type: IBA original_reference_id: GO_REF:0000033 qualifier: is_active_in review: summary: >- ATG2B is active at autophagy initiation/elongation sites. Mammalian ATG2 proteins are required at a late step of autophagosome formation, with a direct mechanism of lipid transfer at the ER-phagophore interface. action: ACCEPT reason: >- This cellular-component annotation matches the conserved ATG2/WIPI role at phagophore assembly sites and is supported by mammalian cell evidence for ATG2-dependent autophagosome formation. additional_reference_ids: - PMID:22219374 - PMID:31721365 supported_by: - reference_id: PMID:22219374 supporting_text: both Atg2A and Atg2B are required for autophagy and that they have redundant and overlapping functions - reference_id: PMID:31721365 supporting_text: direct lipid transfer (LT) from ER to IM for IM expansion - term: id: GO:0000422 label: autophagy of mitochondrion evidence_type: IBA original_reference_id: GO_REF:0000033 qualifier: involved_in review: summary: >- ATG2B is part of the core macroautophagy machinery required to build autophagosomal membranes, so it can participate in mitophagy when the macroautophagy machinery is recruited to mitochondria. action: KEEP_AS_NON_CORE reason: >- The annotation is biologically plausible as a phylogenetic inference for a core autophagy factor, but it should not be interpreted as a mitochondria- selective recognition role for ATG2B. The core function remains phagophore lipid transfer. additional_reference_ids: - PMID:22219374 supported_by: - reference_id: PMID:22219374 supporting_text: these results suggest that Atg2A/B play an essential role, probably at a late step of autophagosome formation - term: id: GO:0061908 label: phagophore evidence_type: IBA original_reference_id: GO_REF:0000033 qualifier: is_active_in review: summary: >- ATG2B functions on nascent autophagic membranes during phagophore expansion and forms a WIPI/ATG18-associated lipid-transfer module. action: ACCEPT reason: >- The phagophore is the core location for ATG2B lipid-transfer activity in autophagosome biogenesis. additional_reference_ids: - PMID:22219374 - PMID:31721365 supported_by: - reference_id: PMID:22219374 supporting_text: these results suggest that Atg2A/B play an essential role, probably at a late step of autophagosome formation - reference_id: PMID:31721365 supporting_text: direct lipid transfer (LT) from ER to IM for IM expansion - term: id: GO:0000425 label: pexophagy evidence_type: IBA original_reference_id: GO_REF:0000033 qualifier: involved_in review: summary: >- ATG2B can support pexophagy indirectly as part of the general autophagosome-building machinery used by selective autophagy pathways. action: KEEP_AS_NON_CORE reason: >- This is best retained as a non-core phylogenetic selective-autophagy annotation. The evidence does not show ATG2B as a peroxisome-selective cargo receptor or specificity factor. additional_reference_ids: - PMID:22219374 supported_by: - reference_id: PMID:22219374 supporting_text: both Atg2A and Atg2B are required for autophagy and that they have redundant and overlapping functions - term: id: GO:0043495 label: protein-membrane adaptor activity evidence_type: IBA original_reference_id: GO_REF:0000033 qualifier: enables review: summary: >- ATG2B binds membranes and WDR45/WIPI4 and functions with this WIPI partner at autophagic membranes, consistent with a protein-membrane adaptor or tethering role in addition to lipid transfer. action: ACCEPT reason: >- Direct biochemical and structural evidence supports ATG2B membrane binding, ATG2B-WDR45 complex formation, and membrane tethering/lipid-transfer activity. The more specific core molecular function is lipid transfer activity, but this term captures a supported tether/adaptor aspect. additional_reference_ids: - PMID:28820312 - PMID:31721365 supported_by: - reference_id: PMID:28820312 supporting_text: mammalian WDR45/WIPI4 has a stronger binding capacity for mammalian ATG2A or ATG2B than the other 3 WIPIs - reference_id: PMID:31721365 supporting_text: ATG2B possesses the membrane tethering (MT) and LT activity - term: id: GO:0061723 label: glycophagy evidence_type: IBA original_reference_id: GO_REF:0000033 qualifier: involved_in review: summary: >- ATG2B may be required for glycophagy insofar as glycophagy uses the core macroautophagy membrane expansion machinery. action: KEEP_AS_NON_CORE reason: >- Retain as a non-core inferred selective-autophagy annotation. The direct evidence for ATG2B is autophagosome membrane formation, not glycogen cargo recognition. additional_reference_ids: - PMID:22219374 supported_by: - reference_id: PMID:22219374 supporting_text: both Atg2A and Atg2B are required for autophagy and that they have redundant and overlapping functions - term: id: GO:0032266 label: phosphatidylinositol-3-phosphate binding evidence_type: IBA original_reference_id: GO_REF:0000033 qualifier: enables review: summary: >- The ATG2-WIPI module is recruited to PI3P-positive autophagic membranes, but the direct PI3P-binding effector is the WIPI/ATG18 family partner, not clearly ATG2B itself. action: MARK_AS_OVER_ANNOTATED reason: >- This term overstates the available gene-level evidence for ATG2B. Direct evidence supports liposome binding and WDR45/WIPI4-dependent association with PI3P-containing membranes, but not a specific ATG2B PI3P headgroup binding activity. additional_reference_ids: - PMID:28820312 - PMID:31721365 supported_by: - reference_id: PMID:28820312 supporting_text: PtdIns3P-binding effectors which can form complexes with proteins in the Atg2 family - reference_id: PMID:28820312 supporting_text: ATG2B and found that it could bind to liposomes independently of PtdIns3P or WDR45 - term: id: GO:0034727 label: piecemeal microautophagy of the nucleus evidence_type: IBA original_reference_id: GO_REF:0000033 qualifier: involved_in review: summary: >- ATG2B is a conserved autophagy factor and may support this process where the conserved machinery is used, but human ATG2B is not established as a process-specific PMN factor. action: KEEP_AS_NON_CORE reason: >- Keep as a non-core phylogenetic inference. The strongest mammalian evidence supports general phagophore/autophagosome biogenesis rather than a human PMN-specific role. additional_reference_ids: - PMID:22219374 supported_by: - reference_id: PMID:22219374 supporting_text: both Atg2A and Atg2B are required for autophagy and that they have redundant and overlapping functions - term: id: GO:0061709 label: reticulophagy evidence_type: IBA original_reference_id: GO_REF:0000033 qualifier: involved_in review: summary: >- ATG2B can support reticulophagy as part of the core autophagy membrane expansion system, especially because ATG2B acts at ER-phagophore contact sites. action: KEEP_AS_NON_CORE reason: >- Retain as non-core. ATG2B is not a reticulophagy cargo receptor or ER quality-control factor; its direct role is lipid transfer for phagophore expansion. additional_reference_ids: - PMID:31721365 supported_by: - reference_id: PMID:31721365 supporting_text: direct lipid transfer (LT) from ER to IM for IM expansion - term: id: GO:0005789 label: endoplasmic reticulum membrane evidence_type: IEA original_reference_id: GO_REF:0000044 qualifier: located_in review: summary: >- ER membrane localization is consistent with the accepted model in which ATG2B tethers the isolation membrane/phagophore edge to the ER and transfers lipids from ER-derived membranes to the growing phagophore. action: ACCEPT reason: >- Although this GOA row is electronically inferred from UniProt subcellular location, it is consistent with experimental and biochemical evidence for ATG2-dependent ER-to-phagophore lipid transfer. additional_reference_ids: - PMID:31721365 supported_by: - reference_id: PMID:31721365 supporting_text: direct lipid transfer (LT) from ER to IM for IM expansion - term: id: GO:0005811 label: lipid droplet evidence_type: IEA original_reference_id: GO_REF:0000044 qualifier: located_in review: summary: >- Mammalian ATG2 proteins associate with lipid droplets and affect lipid droplet morphology and dispersion, but this is a secondary cellular context relative to ATG2B's core phagophore lipid-transfer role. action: KEEP_AS_NON_CORE reason: >- Retain the localization, but mark non-core because the publication notes that the precise lipid-droplet mechanism and directness of the association remain unresolved. additional_reference_ids: - PMID:22219374 supported_by: - reference_id: PMID:22219374 supporting_text: One novel aspect of the present study is the finding that mammalian Atg2 proteins are present on lipid droplets - reference_id: PMID:22219374 supporting_text: Further experiments will be required to test whether Atg2 proteins directly or indirectly associate with lipid droplets - term: id: GO:0006914 label: autophagy evidence_type: IEA original_reference_id: GO_REF:0000002 qualifier: involved_in review: summary: >- The broad InterPro-derived autophagy annotation is directionally correct but less informative than the experimentally supported role in autophagosome assembly/phagophore expansion. action: MODIFY reason: >- Modify to GO:0000045 autophagosome assembly. ATG2B is not merely associated with autophagy in general; it is required for formation and closure/expansion of autophagosomal membranes. proposed_replacement_terms: - id: GO:0000045 label: autophagosome assembly additional_reference_ids: - PMID:22219374 - PMID:31721365 supported_by: - reference_id: PMID:22219374 supporting_text: these results suggest that Atg2A/B play an essential role, probably at a late step of autophagosome formation - reference_id: PMID:31721365 supporting_text: direct lipid transfer (LT) from ER to IM for IM expansion - term: id: GO:0034045 label: phagophore assembly site membrane evidence_type: IEA original_reference_id: GO_REF:0000044 qualifier: located_in review: summary: >- The phagophore assembly site membrane is a supported location for ATG2B function and matches its WIPI-associated lipid-transfer role at the growing isolation membrane. action: ACCEPT reason: >- The electronic UniProt location mapping is supported by the experimental ATG2A/B depletion phenotype and by ATG2B membrane tethering/lipid-transfer activity. additional_reference_ids: - PMID:22219374 - PMID:31721365 supported_by: - reference_id: PMID:22219374 supporting_text: these results suggest that Atg2A/B play an essential role, probably at a late step of autophagosome formation - reference_id: PMID:31721365 supporting_text: direct lipid transfer (LT) from ER to IM for IM expansion - term: id: GO:0120009 label: intermembrane lipid transfer evidence_type: IEA original_reference_id: GO_REF:0000108 qualifier: involved_in review: summary: >- ATG2B has membrane tethering and lipid-transfer activity and transfers lipids between membranes during isolation membrane expansion. action: ACCEPT reason: >- This BP annotation follows directly from the experimentally demonstrated lipid-transfer activity and the model of ER-to-isolation-membrane lipid movement. additional_reference_ids: - PMID:31721365 supported_by: - reference_id: PMID:31721365 supporting_text: ATG2B possesses the membrane tethering (MT) and LT activity - reference_id: PMID:31721365 supporting_text: direct lipid transfer (LT) from ER to IM for IM expansion - term: id: GO:0005515 label: protein binding evidence_type: IPI original_reference_id: PMID:20562859 qualifier: enables review: summary: >- This interaction record reflects WDR45/WIPI4 association in the human autophagy interaction network, but generic protein binding is not an informative ATG2B molecular function. action: MARK_AS_OVER_ANNOTATED reason: >- The WDR45 interaction is biologically relevant, but the curation should be represented by the ATG2-ATG18 complex/component context and ATG2B lipid transfer/tethering function rather than by GO:0005515. additional_reference_ids: - PMID:28820312 supported_by: - reference_id: PMID:20562859 supporting_text: association between ATG2A, ATG2B, and WDR45 was unaltered by mTOR inhibition - reference_id: PMID:28820312 supporting_text: mammalian WDR45/WIPI4 has a stronger binding capacity for mammalian ATG2A or ATG2B than the other 3 WIPIs - term: id: GO:0005515 label: protein binding evidence_type: IPI original_reference_id: PMID:33961781 qualifier: enables review: summary: >- This BioPlex interaction row is a high-throughput WDR45 association, but GO:0005515 is too generic for ATG2B. action: MARK_AS_OVER_ANNOTATED reason: >- Retaining a generic protein binding MF would obscure the supported biology: ATG2B forms a WDR45/WIPI4-associated autophagy lipid-transfer complex. The interaction can inform complex membership but should not be treated as a core molecular function by itself. additional_reference_ids: - PMID:28820312 - PMID:31721365 supported_by: - reference_id: PMID:33961781 supporting_text: These networks model the interactome whose structure encodes protein function, localization, and complex membership. - reference_id: PMID:28820312 supporting_text: mammalian WDR45/WIPI4 has a stronger binding capacity for mammalian ATG2A or ATG2B than the other 3 WIPIs - term: id: GO:0005515 label: protein binding evidence_type: IPI original_reference_id: PMID:34524948 qualifier: enables review: summary: >- This proximity-interactome annotation supports ATG2B's placement in the human macroautophagy interaction network, but it is not an adequate molecular-function description. action: MARK_AS_OVER_ANNOTATED reason: >- Generic protein binding should not be propagated as ATG2B's function. The informative curation is lipid transfer activity and ATG2-WIPI complex membership. additional_reference_ids: - PMID:28820312 - PMID:31721365 supported_by: - reference_id: PMID:34524948 supporting_text: Here, we applied BioID to the study of macroautophagy in human cells, generating a proximity interaction map of 39 core macroautophagy proteins. - reference_id: PMID:31721365 supporting_text: ATG2B possesses the membrane tethering (MT) and LT activity - term: id: GO:0005515 label: protein binding evidence_type: IPI original_reference_id: PMID:35271311 qualifier: enables review: summary: >- The OpenCell-derived interaction annotation is useful as interactome context, but GO:0005515 is too broad and uninformative for ATG2B curation. action: MARK_AS_OVER_ANNOTATED reason: >- The interaction should be interpreted conservatively as supporting complex context, not as the gene product's molecular function. ATG2B's curated molecular function is lipid transfer activity. additional_reference_ids: - PMID:28820312 - PMID:31721365 supported_by: - reference_id: PMID:35271311 supporting_text: well-curated localization and interactome measurements - reference_id: PMID:28820312 supporting_text: We also purified the ATG2B-WDR45 complex and then performed 3-dimensional reconstruction of the complex - term: id: GO:0005783 label: endoplasmic reticulum evidence_type: IDA original_reference_id: GO_REF:0000052 qualifier: located_in review: summary: >- ER localization is consistent with ATG2B's role at ER-associated autophagosome biogenesis sites. action: ACCEPT reason: >- The ER is central to ATG2B-mediated lipid transfer to the isolation membrane; this broader ER localization is valid alongside the more specific ER membrane annotation. additional_reference_ids: - PMID:31721365 supported_by: - reference_id: PMID:31721365 supporting_text: direct lipid transfer (LT) from ER to IM for IM expansion - term: id: GO:0005789 label: endoplasmic reticulum membrane evidence_type: ISS original_reference_id: GO_REF:0000024 qualifier: located_in review: summary: >- The ER membrane annotation transferred by sequence similarity is consistent with the experimentally supported ATG2 lipid-transfer model. action: ACCEPT reason: >- ATG2B acts at an ER-to-isolation-membrane interface, so ER membrane localization is a core cellular context. additional_reference_ids: - PMID:31721365 supported_by: - reference_id: PMID:31721365 supporting_text: direct lipid transfer (LT) from ER to IM for IM expansion - term: id: GO:0005811 label: lipid droplet evidence_type: EXP original_reference_id: PMID:22219374 qualifier: located_in review: summary: >- ATG2 proteins were observed on lipid droplets, and ATG2A/B depletion altered lipid droplet size and distribution. action: KEEP_AS_NON_CORE reason: >- This direct localization should be retained, but it is not the main ATG2B core function. The lipid-droplet role is experimentally observed but mechanistically less resolved than ER- phagophore lipid transfer. supported_by: - reference_id: PMID:22219374 supporting_text: One novel aspect of the present study is the finding that mammalian Atg2 proteins are present on lipid droplets - reference_id: PMID:22219374 supporting_text: These data suggest that mammalian Atg2A and Atg2B function both in autophagosome formation and regulation of lipid droplet volume and distribution. - term: id: GO:0034045 label: phagophore assembly site membrane evidence_type: EXP original_reference_id: PMID:22219374 qualifier: located_in review: summary: >- Direct mammalian evidence supports ATG2 localization/function at autophagic membranes and shows defective autophagosome formation after combined ATG2A/B depletion. action: ACCEPT reason: >- This is a core location for ATG2B lipid transfer and membrane tethering during phagophore expansion. supported_by: - reference_id: PMID:22219374 supporting_text: these results suggest that Atg2A/B play an essential role, probably at a late step of autophagosome formation - term: id: GO:0120013 label: lipid transfer activity evidence_type: IDA original_reference_id: PMID:31721365 qualifier: enables review: summary: >- Human ATG2B directly possesses lipid-transfer activity, and this activity is promoted by negatively charged membranes and WIPI4. action: ACCEPT reason: >- This is the most informative supported molecular-function annotation for ATG2B and should be treated as the core molecular function. supported_by: - reference_id: PMID:31721365 supporting_text: ATG2B possesses the membrane tethering (MT) and LT activity - reference_id: PMID:31721365 supporting_text: negatively charged membranes and an Atg18 ortholog WIPI4 - term: id: GO:0062079 label: ATG2-ATG18 complex evidence_type: IDA original_reference_id: PMID:28820312 qualifier: part_of review: summary: >- NEW annotation. ATG2B forms an ATG2-WIPI/ATG18 complex with WDR45/WIPI4, consistent with direct structural and biochemical evidence for complex membership. action: NEW reason: >- This is the conservative way to use the PN projection. The projection is not evidence for a new proteostasis function by itself, but it points to an existing GO complex term that is directly supported by ATG2B-WDR45 structural and biochemical evidence. additional_reference_ids: - PMID:20562859 - PMID:31721365 supported_by: - reference_id: PMID:28820312 supporting_text: We also purified the ATG2B-WDR45 complex and then performed 3-dimensional reconstruction of the complex - reference_id: PMID:28820312 supporting_text: conserved aromatic H/YF motif in the C terminus of ATG2A and ATG2B that is crucial for complex formation - reference_id: PMID:20562859 supporting_text: association between ATG2A, ATG2B, and WDR45 was unaltered by mTOR inhibition references: - id: GO_REF:0000002 title: Gene Ontology annotation through association of InterPro records with GO terms findings: [] - id: GO_REF:0000024 title: Manual transfer of experimentally-verified manual GO annotation data to orthologs by curator judgment of sequence similarity findings: [] - id: GO_REF:0000033 title: Annotation inferences using phylogenetic trees findings: [] - id: GO_REF:0000044 title: Gene Ontology annotation based on UniProtKB/Swiss-Prot Subcellular Location vocabulary mapping, accompanied by conservative changes to GO terms applied by UniProt findings: [] - id: GO_REF:0000052 title: Gene Ontology annotation based on curation of immunofluorescence data findings: [] - id: GO_REF:0000108 title: Automatic assignment of GO terms using logical inference, based on inter-ontology links findings: [] - id: PMID:20562859 title: Network organization of the human autophagy system. findings: - statement: ATG2B is a known autophagy protein in a human autophagy interaction network and associates with WDR45. supporting_text: association between ATG2A, ATG2B, and WDR45 was unaltered by mTOR inhibition - id: PMID:22219374 title: Mammalian Atg2 proteins are essential for autophagosome formation and important for regulation of size and distribution of lipid droplets. findings: - statement: Mammalian ATG2A and ATG2B are required for autophagosome formation and also affect lipid droplet morphology. supporting_text: These data suggest that mammalian Atg2A and Atg2B function both in autophagosome formation and regulation of lipid droplet volume and distribution. - id: PMID:28820312 title: Architecture of the ATG2B-WDR45 complex and an aromatic Y/HF motif crucial for complex formation. full_text_unavailable: true findings: - statement: ATG2B forms a complex with WDR45/WIPI4, supporting ATG2-ATG18 complex membership. supporting_text: We also purified the ATG2B-WDR45 complex and then performed 3-dimensional reconstruction of the complex - id: PMID:31721365 title: Human ATG2B possesses a lipid transfer activity which is accelerated by negatively charged lipids and WIPI4. full_text_unavailable: true findings: - statement: Human ATG2B has membrane tethering and lipid-transfer activity stimulated by WIPI4. supporting_text: ATG2B possesses the membrane tethering (MT) and LT activity - id: PMID:33961781 title: Dual proteome-scale networks reveal cell-specific remodeling of the human interactome. findings: - statement: BioPlex AP-MS data provide proteome-scale interaction context for high-throughput protein-binding annotations. supporting_text: Through affinity-purification mass spectrometry, we have created two proteome-scale, cell-line-specific interaction networks. - id: PMID:34524948 title: Global Proximity Interactome of the Human Macroautophagy Pathway. findings: - statement: BioID mapping places core macroautophagy proteins in a proximity-interaction network and supports cautious interpretation of high-throughput interaction annotations. supporting_text: Here, we applied BioID to the study of macroautophagy in human cells, generating a proximity interaction map of 39 core macroautophagy proteins. - id: PMID:35271311 title: 'OpenCell: Endogenous tagging for the cartography of human cellular organization.' findings: - statement: OpenCell provides endogenous tagging, localization, and interactome measurements for human cellular organization. supporting_text: well-curated localization and interactome measurements - id: PMID:32009292 title: A conserved ATG2-GABARAP family interaction is critical for phagophore formation. full_text_unavailable: true findings: - statement: A conserved ATG2 LIR mediates binding of ATG2A/ATG2B to GABARAP-subfamily ATG8 proteins; this interaction is required for phagophore formation and closure, whereas the ATG2-WIPI4 interaction is dispensable for autophagy flux. - statement: Endogenous ATG2B co-localizes with ATG2A on early autophagic membranes positive for WIPI2 and ATG16L1, providing direct ATG2B localization evidence at phagophore assembly sites. - id: PMID:30952800 title: ATG2 transports lipids to promote autophagosome biogenesis. findings: - statement: ATG2 is a lipid-transfer protein operating at the ER-autophagosome interface; lipid transfer is the major autophagy-dependent activity, since a lipid-transfer-competent ATG2 N-terminal fragment rescues autophagosome biogenesis in ATG2A/ATG2B double-knockout cells whereas lipid-transport-defective mutants do not. - id: PMID:38622126 title: ANKFY1 bridges ATG2A-mediated lipid transfer from endosomes to phagophores. findings: - statement: The endosomal PI3P-binding protein ANKFY1 binds ATG2A and enhances ATG2-mediated lipid transfer, identifying PI3P-enriched endosomes as an additional lipid source for ATG2-mediated phagophore expansion; ANKFY1 depletion impairs autophagosome growth and largely phenocopies ATG2A/ATG2B depletion. - id: PMID:26280900 title: Germline duplication of ATG2B and GSKIP predisposes to familial myeloid malignancies. full_text_unavailable: true findings: - statement: An autosomal-dominant germline 14q32 duplication spanning ATG2B and GSKIP predisposes to familial myeloid malignancies, and overexpression of ATG2B/GSKIP enhances hematopoietic progenitor differentiation and cooperates with JAK2/MPL/CALR driver mutations; the disease effect is a dosage-sensitive locus-level property shared with GSKIP rather than an ATG2B-specific molecular function. core_functions: - description: >- ATG2B is a WDR45/WIPI4-associated lipid-transfer and membrane-tethering protein that transfers lipids between ER-derived membrane sources and the growing isolation membrane/phagophore during autophagosome assembly. molecular_function: id: GO:0120013 label: lipid transfer activity directly_involved_in: - id: GO:0120009 label: intermembrane lipid transfer - id: GO:0000045 label: autophagosome assembly locations: - id: GO:0034045 label: phagophore assembly site membrane - id: GO:0005789 label: endoplasmic reticulum membrane - id: GO:0061908 label: phagophore in_complex: id: GO:0062079 label: ATG2-ATG18 complex supported_by: - reference_id: PMID:31721365 supporting_text: ATG2B possesses the membrane tethering (MT) and LT activity - reference_id: PMID:28820312 supporting_text: We also purified the ATG2B-WDR45 complex and then performed 3-dimensional reconstruction of the complex - reference_id: PMID:22219374 supporting_text: these results suggest that Atg2A/B play an essential role, probably at a late step of autophagosome formation proposed_new_terms: [] suggested_questions: - question: >- Does ATG2B directly associate with lipid droplets at endogenous expression levels, or is the lipid-droplet phenotype secondary to altered ER/phagophore lipid handling? experts: - Noboru Mizushima - Noriyuki Noda - Li Yu - question: >- Is ATG2B's GABARAP/ATG8-family interaction via the conserved ATG2 LIR individually required for ATG2B function at the phagophore (as shown for the ATG2 family), and does it represent a distinct ATG8-binding molecular function separable from WIPI4 binding in mammalian cells? experts: - David G. McEwan - Noboru Mizushima suggested_experiments: - hypothesis: >- ATG2B-WDR45 complex formation is required for efficient ATG2B recruitment to PI3P-positive phagophore membranes and for autophagosome membrane expansion. description: >- Use endogenous ATG2B knockout/rescue cells expressing wild-type ATG2B or WDR45-binding motif mutants, then measure ATG2B-WDR45 colocalization, autophagic flux, and phagophore closure under starvation. experiment_type: genome editing, rescue, live-cell imaging, and autophagy flux assay - hypothesis: >- ATG2B, like ATG2A, can transfer lipids from PI3P-enriched endosomes (via ANKFY1) in addition to the ER, contributing endosome-derived lipid to phagophore expansion. description: >- In ATG2A/ATG2B double-knockout cells reconstituted with ATG2B, test whether ANKFY1 depletion reduces ATG2B-dependent autophagosome growth and PI3P distribution on phagophores, and assay ANKFY1-ATG2B binding and ATG2B-mediated lipid transfer between PI3P-containing liposomes in vitro. experiment_type: knockout/rescue, proximity imaging, in vitro lipid-transfer assay