{"gene":"ANAPC2","run_date":"2026-06-09T22:02:43","timeline":{"discoveries":[{"year":2020,"finding":"ANAPC2, as a core subunit of the APC/C E3 ubiquitin ligase, interacts with the mycobacterial protein Rv0222 and promotes attachment of lysine-11-linked ubiquitin chains to lysine 76 of Rv0222, which facilitates recruitment of the protein tyrosine phosphatase SHP1 to the adaptor protein TRAF6, preventing lysine-63-linked ubiquitination and activation of TRAF6, thereby suppressing proinflammatory cytokine expression.","method":"Co-immunoprecipitation, shRNA knockdown of ANAPC2, site-directed mutagenesis of Rv0222 ubiquitination site, mouse infection model","journal":"Nature","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, shRNA knockdown with defined molecular phenotype, mutagenesis of substrate ubiquitination site, and in vivo validation; multiple orthogonal methods in a single rigorous study","pmids":["31942069"],"is_preprint":false},{"year":2014,"finding":"The ANAPC2-ANAPC11 subcomplex (the cullin-RING ligase module of APC/C) catalyzes ubiquitin chain elongation in combination with the E2 UBE2C/UbcH10, and the post-ZBR region of Emi2 interacts directly with the cullin subunit ANAPC2 to inhibit this activity.","method":"In vitro ubiquitin chain elongation assay with reconstituted ANAPC2-ANAPC11 subcomplex, domain mutagenesis of Emi2 ZBR, pulldown/interaction mapping","journal":"FEBS open bio","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution of enzymatic activity with the ANAPC2-ANAPC11 subcomplex, mutagenesis identifying key residues, two orthogonal methods in a single study","pmids":["25161877"],"is_preprint":false},{"year":2009,"finding":"Knockdown of the ANAPC2 subunit in Xenopus disrupts the polarity of motile cilia and alters the directionality of fluid movement along the epidermis; mechanistically, the APC/C recognizes a D-box motif on Dishevelled (Dvl) and ubiquitylates Dvl on a conserved lysine residue, placing ANAPC2/APC/C upstream of Dvl in the planar cell polarity pathway required for cilia orientation.","method":"Morpholino knockdown of ANAPC2 in Xenopus embryos, D-box motif identification, ubiquitylation assay of Dvl, live imaging of cilia polarity and fluid flow","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic loss-of-function with defined ciliary polarity phenotype, substrate D-box recognition and ubiquitylation assay, multiple orthogonal methods","pmids":["19805045"],"is_preprint":false},{"year":2017,"finding":"Inducible knockout of Anapc2 in mice causes fatal bone marrow failure within 7 days, with rapid loss of hematopoietic stem and progenitor cells (HSPCs); cell cycle analysis showed that quiescent HSPCs shift from quiescence to mitosis followed by apoptosis, associated with dysregulation of Skp2, P27, Cdk2, and Cyclin E1, establishing ANAPC2/APC/C as essential for HSPC quiescence maintenance.","method":"Inducible Anapc2 knockout mouse model, BrdU label-retaining cell assay, colony formation assay, cell cycle regulator protein analysis, bone marrow failure phenotyping","journal":"Oncotarget","confidence":"High","confidence_rationale":"Tier 2 / Moderate — clean conditional knockout with defined hematopoietic phenotype, BrdU quiescence assay, and cell cycle molecular readouts; single lab with multiple orthogonal methods","pmids":["28968996"],"is_preprint":false},{"year":2021,"finding":"ANAPC2, along with ANAPC8 and ANAPC10, directly interacts with FBXO43 (Emi2) in mouse testicular protein extracts, confirming that FBXO43 is a direct inhibitor of the APC/C complex through physical association with core APC/C subunits including ANAPC2.","method":"Co-immunoprecipitation from mouse testicular protein extracts, validated by Western blot","journal":"Clinical genetics","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — single Co-IP experiment from tissue extracts, replicated across three APC/C subunits in the same study but single method","pmids":["34595750"],"is_preprint":false},{"year":2022,"finding":"Overexpression of ANAPC2 significantly reduces KRAS protein levels in colorectal cancer cells via the ubiquitin-proteasome pathway (reversed by MG132), and knockdown of ANAPC2 abolishes the KRAS-reducing effect of combined artesunate/WNT974 treatment, establishing ANAPC2 as an E3 ligase that can target KRAS for proteasomal degradation.","method":"ANAPC2 overexpression, shRNA knockdown, MG132 proteasome inhibitor rescue, Western blot, xenograft mouse model, IHC","journal":"Cell communication and signaling : CCS","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function and gain-of-function with defined molecular phenotype (KRAS protein level), proteasome inhibitor rescue confirming pathway, and in vivo validation; single lab","pmids":["35305671"],"is_preprint":false},{"year":2023,"finding":"In C. elegans, loss of function of apc-2 (ANAPC2 ortholog) produces a migratory detachment phenotype similar to loss of lnkn-1 (LINKIN/ITFG1), and ANAPC2 was identified as a potential interactor of ITFG1 by IP-MS in human HEK293T cells, suggesting ANAPC2 participates in LINKIN-dependent cell adhesion during collective cell migration.","method":"IP-MS in human HEK293T cells, C. elegans lnkn-1/apc-2 loss-of-function genetic analysis with migration phenotype readout","journal":"bioRxiv","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single IP-MS hit combined with C. elegans genetic phenotype in a preprint; no direct mechanistic validation of ANAPC2's role in the adhesion complex","pmids":["36798316"],"is_preprint":true},{"year":2024,"finding":"Overexpression of ANAPC2 in human subcutaneous pre-adipocytes inhibits adipogenesis, establishing a functional role for ANAPC2 in regulating adipocyte differentiation.","method":"Lentiviral overexpression of ANAPC2 in human subcutaneous pre-adipocytes with adipogenesis readout","journal":"Life science alliance","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single gain-of-function experiment in one cell type, single lab, no mechanistic pathway placement for adipogenesis inhibition","pmids":["38702075"],"is_preprint":false}],"current_model":"ANAPC2 is the cullin subunit of the APC/C E3 ubiquitin ligase complex that, together with ANAPC11 and the E2 UBE2C, catalyzes K11-linked ubiquitin chain synthesis on substrates including cell-cycle regulators (e.g., Dvl for cilia polarity), KRAS (for proteasomal degradation), and bacterial effectors (e.g., Mtb Rv0222 to suppress innate immune signaling via SHP1-TRAF6); ANAPC2 is essential for hematopoietic stem cell quiescence and is directly bound and inhibited by the APC/C inhibitor FBXO43/Emi2 through its post-ZBR region."},"narrative":{"mechanistic_narrative":"ANAPC2 is the cullin subunit of the anaphase-promoting complex/cyclosome (APC/C), a multi-subunit E3 ubiquitin ligase that controls cell-cycle progression and broader protein turnover [PMID:28968996]. Together with the RING subunit ANAPC11, ANAPC2 forms the catalytic cullin-RING module that drives ubiquitin chain elongation in conjunction with the E2 enzyme UBE2C/UbcH10 [PMID:25161877]. APC/C engages substrates through D-box recognition, as shown for Dishevelled, which it ubiquitylates on a conserved lysine to position ANAPC2/APC/C upstream of Dvl in the planar cell polarity pathway governing motile cilia orientation and directional fluid flow [PMID:19805045]. Beyond canonical cell-cycle control, ANAPC2 directs substrate ubiquitination in diverse contexts: it can target KRAS for proteasomal degradation in colorectal cancer cells [PMID:35305671], and is hijacked by the Mycobacterium tuberculosis effector Rv0222, attaching K11-linked ubiquitin chains to Rv0222 to recruit SHP1 to TRAF6 and suppress proinflammatory signaling [PMID:31942069]. Its catalytic activity is held in check by the inhibitor FBXO43/Emi2, which binds ANAPC2 directly through its post-ZBR region [PMID:25161877, PMID:34595750]. Physiologically, ANAPC2 is essential for hematopoietic stem and progenitor cell quiescence; its loss drives quiescent HSPCs into mitosis and apoptosis with dysregulation of Skp2, p27, Cdk2 and Cyclin E1, causing fatal bone marrow failure [PMID:28968996].","teleology":[{"year":2009,"claim":"Established that ANAPC2/APC/C acts upstream of Dishevelled in planar cell polarity, answering how APC/C activity links to a developmental patterning output beyond classical mitotic control.","evidence":"Morpholino knockdown in Xenopus embryos with D-box mapping and ubiquitylation assay of Dvl, plus live imaging of cilia polarity and fluid flow","pmids":["19805045"],"confidence":"High","gaps":["Does not resolve whether Dvl ubiquitylation triggers degradation or non-degradative regulation","Coactivator (Cdc20/Cdh1) dependence of Dvl recognition not defined"]},{"year":2014,"claim":"Defined the enzymatic core of APC/C catalysis and a mechanism of inhibition, showing the ANAPC2-ANAPC11 module elongates ubiquitin chains with UBE2C and is directly inhibited by Emi2's post-ZBR region binding ANAPC2.","evidence":"In vitro ubiquitin chain elongation assay with reconstituted ANAPC2-ANAPC11 subcomplex and Emi2 ZBR domain mutagenesis with interaction mapping","pmids":["25161877"],"confidence":"High","gaps":["Structural basis of the Emi2-ANAPC2 contact not resolved","Does not address physiological substrate selectivity within intact APC/C"]},{"year":2017,"claim":"Demonstrated a non-redundant physiological requirement for ANAPC2 in maintaining stem cell quiescence, answering whether APC/C is needed to keep HSPCs out of cycle.","evidence":"Inducible Anapc2 knockout mouse with BrdU label-retention, colony assays, and cell-cycle regulator profiling","pmids":["28968996"],"confidence":"High","gaps":["Direct APC/C substrates responsible for quiescence not pinpointed","Whether Skp2/p27/Cdk2/Cyclin E1 changes are direct or downstream is unresolved"]},{"year":2020,"claim":"Revealed that a bacterial pathogen exploits ANAPC2 for immune evasion, establishing that ANAPC2 can mediate K11-linked ubiquitination of a non-host substrate to dampen innate signaling.","evidence":"Reciprocal Co-IP, shRNA knockdown, mutagenesis of the Rv0222 ubiquitination site, and a mouse infection model","pmids":["31942069"],"confidence":"High","gaps":["Which APC/C coactivator/adaptor recognizes Rv0222 is unknown","Whether host substrates are co-regulated during infection not addressed"]},{"year":2021,"claim":"Corroborated FBXO43/Emi2 as a physical APC/C inhibitor in a native tissue context, extending the in vitro Emi2-ANAPC2 interaction to endogenous complexes.","evidence":"Co-immunoprecipitation from mouse testicular extracts across ANAPC2, ANAPC8 and ANAPC10, validated by Western blot","pmids":["34595750"],"confidence":"Medium","gaps":["Single Co-IP method without reciprocal functional assay in this system","Does not map which subunit contact is rate-limiting for inhibition in vivo"]},{"year":2022,"claim":"Identified KRAS as a target whose levels ANAPC2 controls via the ubiquitin-proteasome pathway, linking ANAPC2 to oncogene turnover in colorectal cancer.","evidence":"ANAPC2 overexpression and knockdown with MG132 rescue, Western blot, and xenograft/IHC validation","pmids":["35305671"],"confidence":"Medium","gaps":["Direct ubiquitination of KRAS by ANAPC2 not demonstrated in vitro","Degron/D-box on KRAS and coactivator requirement undefined"]},{"year":2023,"claim":"Raised the possibility that ANAPC2 participates in collective cell migration through a LINKIN/ITFG1-dependent adhesion process, linking it to a non-canonical adhesion function.","evidence":"IP-MS in HEK293T cells identifying ANAPC2 as an ITFG1 interactor plus C. elegans apc-2/lnkn-1 migration phenotype (preprint)","pmids":["36798316"],"confidence":"Low","gaps":["Single IP-MS hit without reciprocal or functional validation of the interaction","No demonstration that ANAPC2 ubiquitin ligase activity underlies the migration phenotype"]},{"year":2024,"claim":"Suggested a role for ANAPC2 in adipocyte differentiation, indicating a possible metabolic/developmental function.","evidence":"Lentiviral overexpression of ANAPC2 in human subcutaneous pre-adipocytes with adipogenesis readout","pmids":["38702075"],"confidence":"Low","gaps":["Single gain-of-function experiment in one cell type","No substrate or pathway mechanism for the anti-adipogenic effect identified"]},{"year":null,"claim":"How ANAPC2/APC/C achieves substrate selectivity across its disparate targets (cell-cycle regulators, Dvl, KRAS, bacterial effectors) and how coactivators direct these context-specific functions remain unresolved.","evidence":"","pmids":[],"confidence":"Low","gaps":["No unified structural model of substrate engagement across contexts","Coactivator (Cdc20/Cdh1) usage for non-canonical substrates undefined","Direct in vitro ubiquitination not shown for KRAS"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0016874","term_label":"ligase activity","supporting_discovery_ids":[0,1,5]},{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0,1,2]}],"localization":[],"pathway":[{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[3]},{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[0,1,5]}],"complexes":["APC/C"],"partners":["ANAPC11","UBE2C","FBXO43","ANAPC8","ANAPC10","ITFG1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9UJX6","full_name":"Anaphase-promoting complex subunit 2","aliases":["Cyclosome subunit 2"],"length_aa":822,"mass_kda":93.8,"function":"Together with the RING-H2 protein ANAPC11, constitutes the catalytic component of the anaphase promoting complex/cyclosome (APC/C), a cell cycle-regulated E3 ubiquitin ligase that controls progression through mitosis and the G1 phase of the cell cycle (PubMed:11739784, PubMed:18485873). The APC/C complex acts by mediating ubiquitination and subsequent degradation of target proteins: it mainly mediates the formation of 'Lys-11'-linked polyubiquitin chains and, to a lower extent, the formation of 'Lys-48'- and 'Lys-63'-linked polyubiquitin chains (PubMed:11739784, PubMed:18485873). The APC/C complex catalyzes assembly of branched 'Lys-11'-/'Lys-48'-linked branched ubiquitin chains on target proteins (PubMed:29033132). The CDC20-APC/C complex positively regulates the formation of synaptic vesicle clustering at active zone to the presynaptic membrane in postmitotic neurons (By similarity). CDC20-APC/C-induced degradation of NEUROD2 drives presynaptic differentiation (By similarity)","subcellular_location":"","url":"https://www.uniprot.org/uniprotkb/Q9UJX6/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/ANAPC2","classification":"Common Essential","n_dependent_lines":1202,"n_total_lines":1208,"dependency_fraction":0.9950331125827815},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000176248","cell_line_id":"CID000223","localizations":[{"compartment":"cytoplasmic","grade":3},{"compartment":"nucleoplasm","grade":3}],"interactors":[{"gene":"CDC23","stoichiometry":10.0},{"gene":"ANAPC4","stoichiometry":10.0},{"gene":"ANAPC13","stoichiometry":10.0},{"gene":"ANAPC10","stoichiometry":4.0},{"gene":"ANAPC5","stoichiometry":4.0},{"gene":"NEK2","stoichiometry":4.0},{"gene":"ANAPC16","stoichiometry":0.2},{"gene":"CDC16","stoichiometry":0.2},{"gene":"CDC26","stoichiometry":0.2},{"gene":"CDC20","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/target/CID000223","total_profiled":1310},"omim":[{"mim_id":"614534","title":"ANAPHASE-PROMOTING COMPLEX SUBUNIT 11; ANAPC11","url":"https://www.omim.org/entry/614534"},{"mim_id":"609110","title":"F-BOX ONLY PROTEIN 43; FBXO43","url":"https://www.omim.org/entry/609110"},{"mim_id":"606946","title":"ANAPHASE-PROMOTING COMPLEX, SUBUNIT 2; ANAPC2","url":"https://www.omim.org/entry/606946"},{"mim_id":"603850","title":"DYNAMIN 1-LIKE; DNM1L","url":"https://www.omim.org/entry/603850"},{"mim_id":"603619","title":"FIZZY AND CELL DIVISION CYCLE 20-RELATED PROTEIN 1; FZR1","url":"https://www.omim.org/entry/603619"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Nucleoplasm","reliability":"Supported"},{"location":"Mitochondria","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/ANAPC2"},"hgnc":{"alias_symbol":["APC2","KIAA1406"],"prev_symbol":[]},"alphafold":{"accession":"Q9UJX6","domains":[{"cath_id":"-","chopping":"2-39_49-165","consensus_level":"high","plddt":77.2821,"start":2,"end":165},{"cath_id":"3.30.230.130","chopping":"604-737","consensus_level":"high","plddt":79.5558,"start":604,"end":737},{"cath_id":"1.10.10.10","chopping":"744-817","consensus_level":"high","plddt":78.143,"start":744,"end":817}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9UJX6","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9UJX6-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9UJX6-F1-predicted_aligned_error_v6.png","plddt_mean":78.75},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=ANAPC2","jax_strain_url":"https://www.jax.org/strain/search?query=ANAPC2"},"sequence":{"accession":"Q9UJX6","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9UJX6.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9UJX6/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9UJX6"}},"corpus_meta":[{"pmid":"31942069","id":"PMC_31942069","title":"Host-mediated ubiquitination of a mycobacterial protein suppresses immunity.","date":"2020","source":"Nature","url":"https://pubmed.ncbi.nlm.nih.gov/31942069","citation_count":81,"is_preprint":false},{"pmid":"27476875","id":"PMC_27476875","title":"A genome-wide association study of heat stress-associated SNPs in catfish.","date":"2016","source":"Animal genetics","url":"https://pubmed.ncbi.nlm.nih.gov/27476875","citation_count":50,"is_preprint":false},{"pmid":"21042722","id":"PMC_21042722","title":"Novel medicinal mushroom blend suppresses growth and invasiveness of human breast cancer cells.","date":"2010","source":"International journal of oncology","url":"https://pubmed.ncbi.nlm.nih.gov/21042722","citation_count":49,"is_preprint":false},{"pmid":"19805045","id":"PMC_19805045","title":"Regulation of ciliary polarity by the APC/C.","date":"2009","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/19805045","citation_count":47,"is_preprint":false},{"pmid":"22262126","id":"PMC_22262126","title":"Inhibition of STAT1 sensitizes renal cell carcinoma cells to radiotherapy and chemotherapy.","date":"2012","source":"Cancer biology & therapy","url":"https://pubmed.ncbi.nlm.nih.gov/22262126","citation_count":37,"is_preprint":false},{"pmid":"28090815","id":"PMC_28090815","title":"Comparison of Genome-Wide and Gene-Specific DNA Methylation Profiling in First-Trimester Chorionic Villi From Pregnancies Conceived With Infertility Treatments.","date":"2016","source":"Reproductive sciences (Thousand Oaks, Calif.)","url":"https://pubmed.ncbi.nlm.nih.gov/28090815","citation_count":32,"is_preprint":false},{"pmid":"33190592","id":"PMC_33190592","title":"Post translational modifications in tuberculosis: ubiquitination paradox.","date":"2020","source":"Autophagy","url":"https://pubmed.ncbi.nlm.nih.gov/33190592","citation_count":21,"is_preprint":false},{"pmid":"35305671","id":"PMC_35305671","title":"Combination of artesunate and WNT974 induces KRAS protein degradation by upregulating E3 ligase ANACP2 and β-TrCP in the ubiquitin-proteasome pathway.","date":"2022","source":"Cell communication and signaling : CCS","url":"https://pubmed.ncbi.nlm.nih.gov/35305671","citation_count":20,"is_preprint":false},{"pmid":"25161877","id":"PMC_25161877","title":"The zinc-binding region (ZBR) fragment of Emi2 can inhibit APC/C by targeting its association with the coactivator Cdc20 and UBE2C-mediated ubiquitylation.","date":"2014","source":"FEBS open bio","url":"https://pubmed.ncbi.nlm.nih.gov/25161877","citation_count":20,"is_preprint":false},{"pmid":"24486740","id":"PMC_24486740","title":"Activated pregnane X receptor inhibits cervical cancer cell proliferation and tumorigenicity by inducing G2/M cell-cycle 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multiple orthogonal methods in a single rigorous study\",\n      \"pmids\": [\"31942069\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"The ANAPC2-ANAPC11 subcomplex (the cullin-RING ligase module of APC/C) catalyzes ubiquitin chain elongation in combination with the E2 UBE2C/UbcH10, and the post-ZBR region of Emi2 interacts directly with the cullin subunit ANAPC2 to inhibit this activity.\",\n      \"method\": \"In vitro ubiquitin chain elongation assay with reconstituted ANAPC2-ANAPC11 subcomplex, domain mutagenesis of Emi2 ZBR, pulldown/interaction mapping\",\n      \"journal\": \"FEBS open bio\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution of enzymatic activity with the ANAPC2-ANAPC11 subcomplex, mutagenesis identifying key residues, two orthogonal methods in a single study\",\n      \"pmids\": [\"25161877\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Knockdown of the ANAPC2 subunit in Xenopus disrupts the polarity of motile cilia and alters the directionality of fluid movement along the epidermis; mechanistically, the APC/C recognizes a D-box motif on Dishevelled (Dvl) and ubiquitylates Dvl on a conserved lysine residue, placing ANAPC2/APC/C upstream of Dvl in the planar cell polarity pathway required for cilia orientation.\",\n      \"method\": \"Morpholino knockdown of ANAPC2 in Xenopus embryos, D-box motif identification, ubiquitylation assay of Dvl, live imaging of cilia polarity and fluid flow\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic loss-of-function with defined ciliary polarity phenotype, substrate D-box recognition and ubiquitylation assay, multiple orthogonal methods\",\n      \"pmids\": [\"19805045\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Inducible knockout of Anapc2 in mice causes fatal bone marrow failure within 7 days, with rapid loss of hematopoietic stem and progenitor cells (HSPCs); cell cycle analysis showed that quiescent HSPCs shift from quiescence to mitosis followed by apoptosis, associated with dysregulation of Skp2, P27, Cdk2, and Cyclin E1, establishing ANAPC2/APC/C as essential for HSPC quiescence maintenance.\",\n      \"method\": \"Inducible Anapc2 knockout mouse model, BrdU label-retaining cell assay, colony formation assay, cell cycle regulator protein analysis, bone marrow failure phenotyping\",\n      \"journal\": \"Oncotarget\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — clean conditional knockout with defined hematopoietic phenotype, BrdU quiescence assay, and cell cycle molecular readouts; single lab with multiple orthogonal methods\",\n      \"pmids\": [\"28968996\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"ANAPC2, along with ANAPC8 and ANAPC10, directly interacts with FBXO43 (Emi2) in mouse testicular protein extracts, confirming that FBXO43 is a direct inhibitor of the APC/C complex through physical association with core APC/C subunits including ANAPC2.\",\n      \"method\": \"Co-immunoprecipitation from mouse testicular protein extracts, validated by Western blot\",\n      \"journal\": \"Clinical genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — single Co-IP experiment from tissue extracts, replicated across three APC/C subunits in the same study but single method\",\n      \"pmids\": [\"34595750\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Overexpression of ANAPC2 significantly reduces KRAS protein levels in colorectal cancer cells via the ubiquitin-proteasome pathway (reversed by MG132), and knockdown of ANAPC2 abolishes the KRAS-reducing effect of combined artesunate/WNT974 treatment, establishing ANAPC2 as an E3 ligase that can target KRAS for proteasomal degradation.\",\n      \"method\": \"ANAPC2 overexpression, shRNA knockdown, MG132 proteasome inhibitor rescue, Western blot, xenograft mouse model, IHC\",\n      \"journal\": \"Cell communication and signaling : CCS\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function and gain-of-function with defined molecular phenotype (KRAS protein level), proteasome inhibitor rescue confirming pathway, and in vivo validation; single lab\",\n      \"pmids\": [\"35305671\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"In C. elegans, loss of function of apc-2 (ANAPC2 ortholog) produces a migratory detachment phenotype similar to loss of lnkn-1 (LINKIN/ITFG1), and ANAPC2 was identified as a potential interactor of ITFG1 by IP-MS in human HEK293T cells, suggesting ANAPC2 participates in LINKIN-dependent cell adhesion during collective cell migration.\",\n      \"method\": \"IP-MS in human HEK293T cells, C. elegans lnkn-1/apc-2 loss-of-function genetic analysis with migration phenotype readout\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single IP-MS hit combined with C. elegans genetic phenotype in a preprint; no direct mechanistic validation of ANAPC2's role in the adhesion complex\",\n      \"pmids\": [\"36798316\"],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Overexpression of ANAPC2 in human subcutaneous pre-adipocytes inhibits adipogenesis, establishing a functional role for ANAPC2 in regulating adipocyte differentiation.\",\n      \"method\": \"Lentiviral overexpression of ANAPC2 in human subcutaneous pre-adipocytes with adipogenesis readout\",\n      \"journal\": \"Life science alliance\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single gain-of-function experiment in one cell type, single lab, no mechanistic pathway placement for adipogenesis inhibition\",\n      \"pmids\": [\"38702075\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"ANAPC2 is the cullin subunit of the APC/C E3 ubiquitin ligase complex that, together with ANAPC11 and the E2 UBE2C, catalyzes K11-linked ubiquitin chain synthesis on substrates including cell-cycle regulators (e.g., Dvl for cilia polarity), KRAS (for proteasomal degradation), and bacterial effectors (e.g., Mtb Rv0222 to suppress innate immune signaling via SHP1-TRAF6); ANAPC2 is essential for hematopoietic stem cell quiescence and is directly bound and inhibited by the APC/C inhibitor FBXO43/Emi2 through its post-ZBR region.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"ANAPC2 is the cullin subunit of the anaphase-promoting complex/cyclosome (APC/C), a multi-subunit E3 ubiquitin ligase that controls cell-cycle progression and broader protein turnover [#3]. Together with the RING subunit ANAPC11, ANAPC2 forms the catalytic cullin-RING module that drives ubiquitin chain elongation in conjunction with the E2 enzyme UBE2C/UbcH10 [#1]. APC/C engages substrates through D-box recognition, as shown for Dishevelled, which it ubiquitylates on a conserved lysine to position ANAPC2/APC/C upstream of Dvl in the planar cell polarity pathway governing motile cilia orientation and directional fluid flow [#2]. Beyond canonical cell-cycle control, ANAPC2 directs substrate ubiquitination in diverse contexts: it can target KRAS for proteasomal degradation in colorectal cancer cells [#5], and is hijacked by the Mycobacterium tuberculosis effector Rv0222, attaching K11-linked ubiquitin chains to Rv0222 to recruit SHP1 to TRAF6 and suppress proinflammatory signaling [#0]. Its catalytic activity is held in check by the inhibitor FBXO43/Emi2, which binds ANAPC2 directly through its post-ZBR region [#1, #4]. Physiologically, ANAPC2 is essential for hematopoietic stem and progenitor cell quiescence; its loss drives quiescent HSPCs into mitosis and apoptosis with dysregulation of Skp2, p27, Cdk2 and Cyclin E1, causing fatal bone marrow failure [#3].\",\n  \"teleology\": [\n    {\n      \"year\": 2009,\n      \"claim\": \"Established that ANAPC2/APC/C acts upstream of Dishevelled in planar cell polarity, answering how APC/C activity links to a developmental patterning output beyond classical mitotic control.\",\n      \"evidence\": \"Morpholino knockdown in Xenopus embryos with D-box mapping and ubiquitylation assay of Dvl, plus live imaging of cilia polarity and fluid flow\",\n      \"pmids\": [\"19805045\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Does not resolve whether Dvl ubiquitylation triggers degradation or non-degradative regulation\", \"Coactivator (Cdc20/Cdh1) dependence of Dvl recognition not defined\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Defined the enzymatic core of APC/C catalysis and a mechanism of inhibition, showing the ANAPC2-ANAPC11 module elongates ubiquitin chains with UBE2C and is directly inhibited by Emi2's post-ZBR region binding ANAPC2.\",\n      \"evidence\": \"In vitro ubiquitin chain elongation assay with reconstituted ANAPC2-ANAPC11 subcomplex and Emi2 ZBR domain mutagenesis with interaction mapping\",\n      \"pmids\": [\"25161877\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of the Emi2-ANAPC2 contact not resolved\", \"Does not address physiological substrate selectivity within intact APC/C\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Demonstrated a non-redundant physiological requirement for ANAPC2 in maintaining stem cell quiescence, answering whether APC/C is needed to keep HSPCs out of cycle.\",\n      \"evidence\": \"Inducible Anapc2 knockout mouse with BrdU label-retention, colony assays, and cell-cycle regulator profiling\",\n      \"pmids\": [\"28968996\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct APC/C substrates responsible for quiescence not pinpointed\", \"Whether Skp2/p27/Cdk2/Cyclin E1 changes are direct or downstream is unresolved\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Revealed that a bacterial pathogen exploits ANAPC2 for immune evasion, establishing that ANAPC2 can mediate K11-linked ubiquitination of a non-host substrate to dampen innate signaling.\",\n      \"evidence\": \"Reciprocal Co-IP, shRNA knockdown, mutagenesis of the Rv0222 ubiquitination site, and a mouse infection model\",\n      \"pmids\": [\"31942069\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Which APC/C coactivator/adaptor recognizes Rv0222 is unknown\", \"Whether host substrates are co-regulated during infection not addressed\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Corroborated FBXO43/Emi2 as a physical APC/C inhibitor in a native tissue context, extending the in vitro Emi2-ANAPC2 interaction to endogenous complexes.\",\n      \"evidence\": \"Co-immunoprecipitation from mouse testicular extracts across ANAPC2, ANAPC8 and ANAPC10, validated by Western blot\",\n      \"pmids\": [\"34595750\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single Co-IP method without reciprocal functional assay in this system\", \"Does not map which subunit contact is rate-limiting for inhibition in vivo\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Identified KRAS as a target whose levels ANAPC2 controls via the ubiquitin-proteasome pathway, linking ANAPC2 to oncogene turnover in colorectal cancer.\",\n      \"evidence\": \"ANAPC2 overexpression and knockdown with MG132 rescue, Western blot, and xenograft/IHC validation\",\n      \"pmids\": [\"35305671\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct ubiquitination of KRAS by ANAPC2 not demonstrated in vitro\", \"Degron/D-box on KRAS and coactivator requirement undefined\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Raised the possibility that ANAPC2 participates in collective cell migration through a LINKIN/ITFG1-dependent adhesion process, linking it to a non-canonical adhesion function.\",\n      \"evidence\": \"IP-MS in HEK293T cells identifying ANAPC2 as an ITFG1 interactor plus C. elegans apc-2/lnkn-1 migration phenotype (preprint)\",\n      \"pmids\": [\"36798316\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single IP-MS hit without reciprocal or functional validation of the interaction\", \"No demonstration that ANAPC2 ubiquitin ligase activity underlies the migration phenotype\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Suggested a role for ANAPC2 in adipocyte differentiation, indicating a possible metabolic/developmental function.\",\n      \"evidence\": \"Lentiviral overexpression of ANAPC2 in human subcutaneous pre-adipocytes with adipogenesis readout\",\n      \"pmids\": [\"38702075\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single gain-of-function experiment in one cell type\", \"No substrate or pathway mechanism for the anti-adipogenic effect identified\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How ANAPC2/APC/C achieves substrate selectivity across its disparate targets (cell-cycle regulators, Dvl, KRAS, bacterial effectors) and how coactivators direct these context-specific functions remain unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No unified structural model of substrate engagement across contexts\", \"Coactivator (Cdc20/Cdh1) usage for non-canonical substrates undefined\", \"Direct in vitro ubiquitination not shown for KRAS\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016874\", \"supporting_discovery_ids\": [0, 1, 5]},\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0, 1, 2]}\n    ],\n    \"localization\": [],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [3]},\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [0, 1, 5]}\n    ],\n    \"complexes\": [\"APC/C\"],\n    \"partners\": [\"ANAPC11\", \"UBE2C\", \"FBXO43\", \"ANAPC8\", \"ANAPC10\", \"ITFG1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}