{"gene":"BIRC7","run_date":"2026-06-09T22:02:44","timeline":{"discoveries":[{"year":2000,"finding":"ML-IAP (BIRC7) contains a single BIR domain and RING finger motif; deletion and mutational analysis demonstrated that integrity of the BIR domain is required for anti-apoptotic function, likely through direct inhibition of downstream effector caspases.","method":"Deletion and mutational analysis; overexpression in cell lines with apoptosis assays","journal":"Current biology : CB","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function mutagenesis with defined cellular phenotype, single lab, BIR domain necessity confirmed; caspase inhibition proposed but not fully reconstituted in vitro","pmids":["11084335"],"is_preprint":false},{"year":2002,"finding":"ML-IAP BIR domain physically interacts with SMAC/DIABLO through its BIR domain with low nanomolar affinity; single amino acid mutations in either SMAC N-terminus or ML-IAP BIR abolished this interaction; SMAC coexpression or SMAC peptides abrogated ML-IAP-mediated inhibition of cell death.","method":"Co-immunoprecipitation, binding assays with SMAC peptides, single amino acid mutagenesis, cell-based apoptosis assays","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — reciprocal binding assays with mutagenesis defining the interaction interface, functional cell-death rescue, replicated with peptide variants","pmids":["11801603"],"is_preprint":false},{"year":2003,"finding":"X-ray crystal structures of ML-IAP BIR domain in complex with Smac-derived and phage-derived peptides revealed the peptide-binding surface; structure-activity relationships showed that modifications at Pro3' of the Smac peptide confer 7-fold greater affinity for ML-IAP-BIR with ~100-fold selectivity over XIAP-BIR3.","method":"X-ray crystallography, phage display, synthetic peptide SAR, binding affinity measurements","journal":"Biochemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structures with functional validation by SAR and affinity measurements, multiple orthogonal methods in one study","pmids":["12846571"],"is_preprint":false},{"year":2005,"finding":"ML-IAP BIR domain is a weak direct inhibitor of caspases 3 and 9; the primary anti-apoptotic mechanism of ML-IAP is sequestration of Smac/DIABLO, preventing it from antagonizing XIAP-mediated caspase inhibition. Increased ML-IAP expression results in formation of a ML-IAP–Smac complex and disruption of the endogenous XIAP–Smac interaction.","method":"In vitro caspase inhibition assays, BIR domain engineering, co-immunoprecipitation of endogenous ML-IAP–Smac and XIAP–Smac complexes, mutagenesis","journal":"The Biochemical journal","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — in vitro caspase assays, mutagenesis of BIR domain, co-IP of endogenous complexes, multiple orthogonal methods establishing mechanism","pmids":["15485396"],"is_preprint":false},{"year":2006,"finding":"Livin (ML-IAP) is proteolytically cleaved during apoptosis at residue Asp52 (within DHVD52); mutation of Asp52 to Ala or deletion of the adjacent region abrogated cleavage. Cleavage was largely independent of Omi/HtrA2 and for some stimuli independent of caspases, suggesting a non-canonical caspase mediates cleavage.","method":"Site-directed mutagenesis of Asp52, deletion mapping, RNAi knockdown of Omi/HtrA2, pan-caspase inhibitor (zVAD-fmk) treatment, multiple apoptotic stimuli","journal":"Journal of dermatological science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — mutagenesis and deletion mapping with defined cleavage site, single lab, RNAi and pharmacological inhibition used orthogonally","pmids":["16806840"],"is_preprint":false},{"year":2009,"finding":"Livin (ML-IAP) is unique among IAP members in being specifically cleaved by caspases upon strong apoptotic stimulus to produce a truncated protein (tLivin) with paradoxical pro-apoptotic activity; NK cells induce Livin cleavage to tLivin which inhibits tumor growth in vivo.","method":"Point mutations to eliminate caspase cleavage site, tumor xenograft animal model with NK cell-depleted mice, in vivo tumor growth assays","journal":"Cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo epistasis with NK cells, point mutant rescue, single lab with two orthogonal approaches (mutagenesis + animal model)","pmids":["19549891"],"is_preprint":false},{"year":2012,"finding":"The crystal structure of the dimeric BIRC7 RING domain in complex with E2 UbcH5B covalently linked to ubiquitin (UbcH5B~Ub) revealed that RING dimerization stabilizes the donor Ub through extensive noncovalent interactions with UbcH5B and both RING subunits; mutations disrupting these interactions or RING dimerization reduced UbcH5B~Ub binding affinity and ubiquitination activity.","method":"X-ray crystallography, NMR peak-shift analysis, mutagenesis, in vitro ubiquitination assays, binding affinity measurements","journal":"Nature structural & molecular biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure combined with NMR, mutagenesis, and in vitro ubiquitination reconstitution, multiple orthogonal methods in one rigorous study","pmids":["22902369"],"is_preprint":false},{"year":2012,"finding":"ML-IAP directly binds C-RAF kinase; ML-IAP depletion leads to increased C-RAF protein levels, MAPK activation, and increased cell migration in melanoma cells, indicating ML-IAP controls C-RAF protein stability and cell migration.","method":"Co-immunoprecipitation (ML-IAP–C-RAF interaction), siRNA knockdown with Western blot for C-RAF levels, MAPK activation assays, migration assays","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — Co-IP plus loss-of-function with defined cellular phenotypes (C-RAF stability, MAPK, migration), single lab","pmids":["22711539"],"is_preprint":false},{"year":2012,"finding":"ML-IAP undergoes proteasomal degradation in response to bivalent (but not monovalent) IAP antagonists; this degradation requires ML-IAP's own ubiquitin ligase (RING domain) activity and is independent of c-IAP1 or c-IAP2.","method":"IAP antagonist treatment, proteasome inhibitor rescue, RING domain mutants, c-IAP1/c-IAP2 knockout/knockdown cell lines","journal":"The Biochemical journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — RING-domain mutants plus genetic depletion of c-IAP1/2, single lab, two orthogonal approaches","pmids":["22853455"],"is_preprint":false},{"year":2018,"finding":"MicroRNA-23a down-regulates KIAP (BIRC7) expression; inhibition of KIAP enhanced the pro-apoptotic effect of miR-23a, while overexpression of KIAP abrogated it, placing KIAP downstream of miR-23a in a pathway controlling hepatocarcinoma cell apoptosis and caspase-3 activation.","method":"miRNA transfection, siRNA knockdown, plasmid overexpression, Western blot, caspase-3 activity assay, flow cytometry","journal":"European review for medical and pharmacological sciences","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, single cell line, rescue experiment is indirect (miRNA target validation without direct binding confirmation), no orthogonal method","pmids":["30280770"],"is_preprint":false},{"year":2020,"finding":"BIRC7 suppresses autophagy through modulating expression of ATG5 and BECN1; this autophagy suppression drives epithelial-mesenchymal transition (EMT) and metastasis in papillary thyroid carcinoma cells; combined targeting of BIRC7 and mTOR enhances autophagy and achieves synergistic anti-metastatic effects in vitro and in vivo.","method":"BIRC7 overexpression and siRNA knockdown, transmission electron microscopy, Western blot for ATG5/BECN1/EMT markers, wound-healing and invasion assays, xenograft tumor model","journal":"American journal of cancer research","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — loss- and gain-of-function with defined molecular readouts (ATG5, BECN1) and in vivo validation, single lab","pmids":["32064154"],"is_preprint":false},{"year":2022,"finding":"BIRC7 is a transcriptional downstream target of HIF-1α in melanoma; its promoter contains HIF-1α binding sites confirmed by luciferase assay; BIRC7 knockdown reversed the pro-tumorigenic effects of HIF-1α under hypoxia.","method":"Luciferase reporter assay (HIF-1α binding to BIRC7 promoter), RNA-seq under hypoxia, qRT-PCR, Western blot, siRNA knockdown with cellular function assays","journal":"Clinical, cosmetic and investigational dermatology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — luciferase assay for transcriptional regulation plus knockdown phenotype, single lab, no ChIP or direct binding confirmation of HIF-1α to endogenous promoter","pmids":["35747741"],"is_preprint":false}],"current_model":"BIRC7 (ML-IAP/Livin) is a single-BIR/RING-domain IAP that inhibits apoptosis primarily by sequestering Smac/DIABLO via its BIR domain (preventing Smac from antagonizing XIAP), functions as a RING-type E3 ubiquitin ligase that dimerizes to optimally position donor ubiquitin on E2 UbcH5B for transfer, undergoes caspase-mediated cleavage at Asp52 to generate a paradoxically pro-apoptotic truncated form (tLivin), regulates C-RAF protein stability and cell migration in melanoma, and suppresses autophagy (via ATG5/BECN1) to promote EMT and metastasis; its expression is transcriptionally induced by HIF-1α under hypoxia."},"narrative":{"mechanistic_narrative":"BIRC7 (ML-IAP/Livin) is a single-BIR/RING-domain inhibitor of apoptosis that restrains effector caspase activity and couples apoptotic control to ubiquitin signaling [PMID:11084335, PMID:22902369]. Its anti-apoptotic function depends on the integrity of its BIR domain, which is only a weak direct inhibitor of caspases 3 and 9; the dominant mechanism is high-affinity sequestration of Smac/DIABLO, which displaces Smac from XIAP and thereby preserves XIAP-mediated caspase inhibition [PMID:11801603, PMID:15485396]. Crystal structures of the BIR domain bound to Smac-derived and phage-derived peptides defined the peptide-binding groove and enabled selective high-affinity peptide antagonists [PMID:12846571]. The RING domain confers E3 ubiquitin ligase activity: it dimerizes to lock the donor ubiquitin on the E2 UbcH5B~Ub conjugate in a transfer-competent orientation, and this same RING activity drives BIRC7 autodegradation in response to bivalent IAP antagonists [PMID:22902369, PMID:22853455]. BIRC7 is also processed during apoptosis by cleavage at Asp52 to a truncated form (tLivin) that paradoxically acquires pro-apoptotic activity, a conversion exploited by NK cells to limit tumor growth in vivo [PMID:16806840, PMID:19549891]. Beyond core apoptotic control, BIRC7 binds C-RAF and restrains its protein level, MAPK activation, and melanoma cell migration, and it suppresses autophagy through ATG5 and BECN1 to promote EMT and metastasis [PMID:22711539, PMID:32064154].","teleology":[{"year":2000,"claim":"Established BIRC7 as a domain-defined IAP and showed the BIR domain is necessary for anti-apoptotic function, framing the question of how a single-BIR protein blocks cell death.","evidence":"Deletion/mutational analysis with overexpression apoptosis assays in cell lines","pmids":["11084335"],"confidence":"Medium","gaps":["Direct caspase inhibition proposed but not reconstituted in vitro","RING-domain function not addressed","No structural basis for BIR activity"]},{"year":2002,"claim":"Identified Smac/DIABLO as a high-affinity BIR-domain ligand, shifting the mechanistic model away from direct caspase inhibition toward Smac sequestration.","evidence":"Co-IP, Smac-peptide binding assays, single-residue mutagenesis, cell-death rescue","pmids":["11801603"],"confidence":"High","gaps":["Did not quantify the effect on endogenous XIAP-Smac equilibrium","Structural interface not yet resolved"]},{"year":2003,"claim":"Resolved the atomic structure of the BIR–peptide interface, enabling design of antagonists selective for BIRC7 over XIAP.","evidence":"X-ray crystallography, phage display, synthetic peptide SAR, affinity measurements","pmids":["12846571"],"confidence":"High","gaps":["Structures use peptide surrogates rather than full-length Smac","No structure of the RING/E3 module"]},{"year":2005,"claim":"Directly tested the competing mechanisms, showing BIR is only a weak caspase inhibitor and that Smac sequestration disrupting the XIAP-Smac complex is the primary anti-apoptotic route.","evidence":"In vitro caspase assays, BIR engineering, co-IP of endogenous ML-IAP-Smac and XIAP-Smac complexes","pmids":["15485396"],"confidence":"High","gaps":["Quantitative contribution of residual direct caspase inhibition unresolved","Does not address RING-dependent functions"]},{"year":2006,"claim":"Mapped a defined apoptotic cleavage site at Asp52 and showed cleavage can occur by a non-canonical, partly caspase-independent protease, opening the question of how an IAP is itself processed during death.","evidence":"Asp52 mutagenesis, deletion mapping, Omi/HtrA2 RNAi, zVAD-fmk treatment across stimuli","pmids":["16806840"],"confidence":"Medium","gaps":["Identity of the responsible protease not established","Functional consequence of cleavage not defined here"]},{"year":2009,"claim":"Showed cleavage generates a truncated tLivin with paradoxical pro-apoptotic activity that NK cells exploit to suppress tumors, redefining BIRC7 as a context-dependent regulator of death.","evidence":"Caspase-site point mutants, NK-cell-depleted xenograft mouse models, in vivo growth assays","pmids":["19549891"],"confidence":"Medium","gaps":["Molecular basis of tLivin pro-apoptotic gain-of-function unclear","Single lab, limited tumor models"]},{"year":2012,"claim":"Defined the catalytic mechanism of the RING E3 activity, showing dimerization positions the UbcH5B~Ub conjugate for ubiquitin transfer.","evidence":"Crystal structure of dimeric RING–UbcH5B~Ub, NMR, mutagenesis, in vitro ubiquitination","pmids":["22902369"],"confidence":"High","gaps":["Physiological substrates of the ligase not identified in this study","Regulation of dimerization in cells unknown"]},{"year":2012,"claim":"Linked RING activity to drug-induced self-destruction, showing bivalent IAP antagonists trigger BIRC7 autodegradation independent of c-IAP1/2.","evidence":"IAP antagonist treatment, proteasome rescue, RING mutants, c-IAP1/2 knockout/knockdown lines","pmids":["22853455"],"confidence":"Medium","gaps":["Whether degradation is autoubiquitination vs trans-ubiquitination not fully resolved","E2 used in cells not pinpointed"]},{"year":2012,"claim":"Extended BIRC7 function beyond apoptosis by showing it binds C-RAF and limits its stability, MAPK signaling, and melanoma migration.","evidence":"Co-IP, siRNA knockdown with C-RAF Western blot, MAPK and migration assays","pmids":["22711539"],"confidence":"Medium","gaps":["Whether C-RAF is a direct ubiquitination substrate not shown","Single lab, reciprocal validation limited"]},{"year":2018,"claim":"Placed BIRC7 downstream of miR-23a in controlling hepatocarcinoma apoptosis, adding an upstream regulatory layer.","evidence":"miRNA/siRNA transfection, overexpression rescue, caspase-3 assay, flow cytometry","pmids":["30280770"],"confidence":"Low","gaps":["No direct miR-23a–BIRC7 binding confirmation","Single cell line, no orthogonal method"]},{"year":2020,"claim":"Connected BIRC7 to autophagy suppression via ATG5/BECN1 and to EMT/metastasis, with mTOR co-targeting as a synergistic vulnerability.","evidence":"Gain/loss-of-function, electron microscopy, ATG5/BECN1/EMT Westerns, invasion assays, xenografts","pmids":["32064154"],"confidence":"Medium","gaps":["Mechanism by which BIRC7 modulates ATG5/BECN1 expression undefined","Single tumor type"]},{"year":2022,"claim":"Identified BIRC7 as a HIF-1α transcriptional target, linking its expression to the hypoxic tumor microenvironment.","evidence":"Luciferase promoter reporter, hypoxia RNA-seq, qRT-PCR, siRNA knockdown phenotypes","pmids":["35747741"],"confidence":"Low","gaps":["No ChIP confirming HIF-1α binding to endogenous promoter","Single lab"]},{"year":null,"claim":"The endogenous substrate repertoire of the BIRC7 RING ligase and the protease that generates tLivin remain unidentified, leaving the link between its E3 activity and its non-apoptotic roles (C-RAF, autophagy) mechanistically open.","evidence":"","pmids":[],"confidence":"Low","gaps":["No validated physiological ubiquitination substrate","tLivin-generating protease unidentified","Whether C-RAF and autophagy effects are RING-dependent not established"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0016874","term_label":"ligase activity","supporting_discovery_ids":[6,8]},{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[6]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[1,3]},{"term_id":"GO:0140313","term_label":"molecular sequestering activity","supporting_discovery_ids":[3]}],"localization":[],"pathway":[{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[0,3,5]},{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[6,8]}],"complexes":[],"partners":["DIABLO","UBE2D2","RAF1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q96CA5","full_name":"Baculoviral IAP repeat-containing protein 7","aliases":["Kidney inhibitor of apoptosis protein","KIAP","Livin","Melanoma inhibitor of apoptosis protein","ML-IAP","RING finger protein 50","RING-type E3 ubiquitin transferase BIRC7"],"length_aa":298,"mass_kda":32.8,"function":"Apoptotic regulator capable of exerting proapoptotic and anti-apoptotic activities and plays crucial roles in apoptosis, cell proliferation, and cell cycle control (PubMed:11024045, PubMed:11084335, PubMed:11162435, PubMed:16729033, PubMed:17294084). Its anti-apoptotic activity is mediated through the inhibition of CASP3, CASP7 and CASP9, as well as by its E3 ubiquitin-protein ligase activity (PubMed:11024045, PubMed:16729033). As it is a weak caspase inhibitor, its anti-apoptotic activity is thought to be due to its ability to ubiquitinate DIABLO/SMAC targeting it for degradation thereby promoting cell survival (PubMed:16729033). May contribute to caspase inhibition, by blocking the ability of DIABLO/SMAC to disrupt XIAP/BIRC4-caspase interactions (PubMed:16729033). Protects against apoptosis induced by TNF or by chemical agents such as adriamycin, etoposide or staurosporine (PubMed:11084335, PubMed:11162435, PubMed:11865055). Suppression of apoptosis is mediated by activation of MAPK8/JNK1, and possibly also of MAPK9/JNK2 (PubMed:11865055). This activation depends on TAB1 and MAP3K7/TAK1 (PubMed:11865055). In vitro, inhibits CASP3 and proteolytic activation of pro-CASP9 (PubMed:11024045) Blocks staurosporine-induced apoptosis (PubMed:11322947). Promotes natural killer (NK) cell-mediated killing (PubMed:18034418) Blocks etoposide-induced apoptosis (PubMed:11162435, PubMed:11322947). Protects against natural killer (NK) cell-mediated killing (PubMed:18034418)","subcellular_location":"Nucleus; Cytoplasm; Golgi apparatus","url":"https://www.uniprot.org/uniprotkb/Q96CA5/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/BIRC7","classification":"Not Classified","n_dependent_lines":0,"n_total_lines":1208,"dependency_fraction":0.0},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/BIRC7","total_profiled":1310},"omim":[{"mim_id":"605737","title":"BACULOVIRAL IAP REPEAT-CONTAINING PROTEIN 7; BIRC7","url":"https://www.omim.org/entry/605737"},{"mim_id":"605547","title":"FOLLISTATIN-LIKE 1; FSTL1","url":"https://www.omim.org/entry/605547"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Enhanced","locations":[{"location":"Nucleoplasm","reliability":"Enhanced"},{"location":"Centrosome","reliability":"Additional"},{"location":"Cytosol","reliability":"Additional"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in some","driving_tissues":[{"tissue":"placenta","ntpm":6.1}],"url":"https://www.proteinatlas.org/search/BIRC7"},"hgnc":{"alias_symbol":["mliap","ML-IAP","KIAP","RNF50"],"prev_symbol":[]},"alphafold":{"accession":"Q96CA5","domains":[{"cath_id":"1.10.1170.10","chopping":"79-174","consensus_level":"medium","plddt":95.5527,"start":79,"end":174},{"cath_id":"3.30.40.10","chopping":"250-298","consensus_level":"medium","plddt":94.9235,"start":250,"end":298}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q96CA5","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q96CA5-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q96CA5-F1-predicted_aligned_error_v6.png","plddt_mean":71.69},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=BIRC7","jax_strain_url":"https://www.jax.org/strain/search?query=BIRC7"},"sequence":{"accession":"Q96CA5","fasta_url":"https://rest.uniprot.org/uniprotkb/Q96CA5.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q96CA5/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q96CA5"}},"corpus_meta":[{"pmid":"11084335","id":"PMC_11084335","title":"ML-IAP, a novel inhibitor of apoptosis that is preferentially expressed in human melanomas.","date":"2000","source":"Current biology : CB","url":"https://pubmed.ncbi.nlm.nih.gov/11084335","citation_count":336,"is_preprint":false},{"pmid":"22902369","id":"PMC_22902369","title":"BIRC7-E2 ubiquitin conjugate structure reveals the mechanism of ubiquitin transfer by a RING dimer.","date":"2012","source":"Nature structural & molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/22902369","citation_count":281,"is_preprint":false},{"pmid":"11801603","id":"PMC_11801603","title":"SMAC negatively regulates the anti-apoptotic activity of melanoma inhibitor of apoptosis (ML-IAP).","date":"2002","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/11801603","citation_count":119,"is_preprint":false},{"pmid":"15485396","id":"PMC_15485396","title":"Engineering ML-IAP to produce an extraordinarily potent caspase 9 inhibitor: implications for Smac-dependent anti-apoptotic 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dermatology","url":"https://pubmed.ncbi.nlm.nih.gov/19212346","citation_count":8,"is_preprint":false},{"pmid":"23906305","id":"PMC_23906305","title":"Comparative study of clinicopathological significance, BIRC7, and STC2 expression between squamous cell/adenosquamous carcinomas and adenocarcinoma of gallbladder.","date":"2013","source":"Neoplasma","url":"https://pubmed.ncbi.nlm.nih.gov/23906305","citation_count":6,"is_preprint":false},{"pmid":"22510211","id":"PMC_22510211","title":"BIRC7 gene in channel catfish (Ictalurus punctatus): identification and expression analysis in response to Edwardsiella tarda, Streptococcus iniae and Channel catfish Hemorrhage Reovirus.","date":"2012","source":"Fish & shellfish immunology","url":"https://pubmed.ncbi.nlm.nih.gov/22510211","citation_count":6,"is_preprint":false},{"pmid":"16840203","id":"PMC_16840203","title":"Expression of BIRC7 protein and mRNA in non-Hodgkin's lymphoma.","date":"2006","source":"Leukemia & lymphoma","url":"https://pubmed.ncbi.nlm.nih.gov/16840203","citation_count":5,"is_preprint":false},{"pmid":"35747741","id":"PMC_35747741","title":"BIRC7 is Beneficial for Melanoma Progression and Hypoxic Response.","date":"2022","source":"Clinical, cosmetic and investigational dermatology","url":"https://pubmed.ncbi.nlm.nih.gov/35747741","citation_count":2,"is_preprint":false},{"pmid":"30280770","id":"PMC_30280770","title":"MicroRNA-23a induces apoptosis of hepatocarcinoma cell line MHCC97H via down-regulating KIAP: a mechanism study.","date":"2018","source":"European review for medical and pharmacological sciences","url":"https://pubmed.ncbi.nlm.nih.gov/30280770","citation_count":0,"is_preprint":false},{"pmid":"41969613","id":"PMC_41969613","title":"Engineering of the Melanoma Inhibitor of Apoptosis (ML-IAP) Anticancer Peptide Through Comprehensive In Silico Approaches.","date":"2026","source":"Human mutation","url":"https://pubmed.ncbi.nlm.nih.gov/41969613","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":16476,"output_tokens":3128,"usd":0.048174,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":10605,"output_tokens":3562,"usd":0.071038,"stage2_stop_reason":"end_turn"},"total_usd":0.119212,"stage1_batch_id":"msgbatch_0179Z2pxRYBsXi2xADneafRw","stage2_batch_id":"msgbatch_01LaY9ZaVdgjWArcZo9eqQE5","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2000,\n      \"finding\": \"ML-IAP (BIRC7) contains a single BIR domain and RING finger motif; deletion and mutational analysis demonstrated that integrity of the BIR domain is required for anti-apoptotic function, likely through direct inhibition of downstream effector caspases.\",\n      \"method\": \"Deletion and mutational analysis; overexpression in cell lines with apoptosis assays\",\n      \"journal\": \"Current biology : CB\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function mutagenesis with defined cellular phenotype, single lab, BIR domain necessity confirmed; caspase inhibition proposed but not fully reconstituted in vitro\",\n      \"pmids\": [\"11084335\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"ML-IAP BIR domain physically interacts with SMAC/DIABLO through its BIR domain with low nanomolar affinity; single amino acid mutations in either SMAC N-terminus or ML-IAP BIR abolished this interaction; SMAC coexpression or SMAC peptides abrogated ML-IAP-mediated inhibition of cell death.\",\n      \"method\": \"Co-immunoprecipitation, binding assays with SMAC peptides, single amino acid mutagenesis, cell-based apoptosis assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — reciprocal binding assays with mutagenesis defining the interaction interface, functional cell-death rescue, replicated with peptide variants\",\n      \"pmids\": [\"11801603\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"X-ray crystal structures of ML-IAP BIR domain in complex with Smac-derived and phage-derived peptides revealed the peptide-binding surface; structure-activity relationships showed that modifications at Pro3' of the Smac peptide confer 7-fold greater affinity for ML-IAP-BIR with ~100-fold selectivity over XIAP-BIR3.\",\n      \"method\": \"X-ray crystallography, phage display, synthetic peptide SAR, binding affinity measurements\",\n      \"journal\": \"Biochemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structures with functional validation by SAR and affinity measurements, multiple orthogonal methods in one study\",\n      \"pmids\": [\"12846571\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"ML-IAP BIR domain is a weak direct inhibitor of caspases 3 and 9; the primary anti-apoptotic mechanism of ML-IAP is sequestration of Smac/DIABLO, preventing it from antagonizing XIAP-mediated caspase inhibition. Increased ML-IAP expression results in formation of a ML-IAP–Smac complex and disruption of the endogenous XIAP–Smac interaction.\",\n      \"method\": \"In vitro caspase inhibition assays, BIR domain engineering, co-immunoprecipitation of endogenous ML-IAP–Smac and XIAP–Smac complexes, mutagenesis\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — in vitro caspase assays, mutagenesis of BIR domain, co-IP of endogenous complexes, multiple orthogonal methods establishing mechanism\",\n      \"pmids\": [\"15485396\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Livin (ML-IAP) is proteolytically cleaved during apoptosis at residue Asp52 (within DHVD52); mutation of Asp52 to Ala or deletion of the adjacent region abrogated cleavage. Cleavage was largely independent of Omi/HtrA2 and for some stimuli independent of caspases, suggesting a non-canonical caspase mediates cleavage.\",\n      \"method\": \"Site-directed mutagenesis of Asp52, deletion mapping, RNAi knockdown of Omi/HtrA2, pan-caspase inhibitor (zVAD-fmk) treatment, multiple apoptotic stimuli\",\n      \"journal\": \"Journal of dermatological science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — mutagenesis and deletion mapping with defined cleavage site, single lab, RNAi and pharmacological inhibition used orthogonally\",\n      \"pmids\": [\"16806840\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Livin (ML-IAP) is unique among IAP members in being specifically cleaved by caspases upon strong apoptotic stimulus to produce a truncated protein (tLivin) with paradoxical pro-apoptotic activity; NK cells induce Livin cleavage to tLivin which inhibits tumor growth in vivo.\",\n      \"method\": \"Point mutations to eliminate caspase cleavage site, tumor xenograft animal model with NK cell-depleted mice, in vivo tumor growth assays\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo epistasis with NK cells, point mutant rescue, single lab with two orthogonal approaches (mutagenesis + animal model)\",\n      \"pmids\": [\"19549891\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"The crystal structure of the dimeric BIRC7 RING domain in complex with E2 UbcH5B covalently linked to ubiquitin (UbcH5B~Ub) revealed that RING dimerization stabilizes the donor Ub through extensive noncovalent interactions with UbcH5B and both RING subunits; mutations disrupting these interactions or RING dimerization reduced UbcH5B~Ub binding affinity and ubiquitination activity.\",\n      \"method\": \"X-ray crystallography, NMR peak-shift analysis, mutagenesis, in vitro ubiquitination assays, binding affinity measurements\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure combined with NMR, mutagenesis, and in vitro ubiquitination reconstitution, multiple orthogonal methods in one rigorous study\",\n      \"pmids\": [\"22902369\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"ML-IAP directly binds C-RAF kinase; ML-IAP depletion leads to increased C-RAF protein levels, MAPK activation, and increased cell migration in melanoma cells, indicating ML-IAP controls C-RAF protein stability and cell migration.\",\n      \"method\": \"Co-immunoprecipitation (ML-IAP–C-RAF interaction), siRNA knockdown with Western blot for C-RAF levels, MAPK activation assays, migration assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — Co-IP plus loss-of-function with defined cellular phenotypes (C-RAF stability, MAPK, migration), single lab\",\n      \"pmids\": [\"22711539\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"ML-IAP undergoes proteasomal degradation in response to bivalent (but not monovalent) IAP antagonists; this degradation requires ML-IAP's own ubiquitin ligase (RING domain) activity and is independent of c-IAP1 or c-IAP2.\",\n      \"method\": \"IAP antagonist treatment, proteasome inhibitor rescue, RING domain mutants, c-IAP1/c-IAP2 knockout/knockdown cell lines\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — RING-domain mutants plus genetic depletion of c-IAP1/2, single lab, two orthogonal approaches\",\n      \"pmids\": [\"22853455\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"MicroRNA-23a down-regulates KIAP (BIRC7) expression; inhibition of KIAP enhanced the pro-apoptotic effect of miR-23a, while overexpression of KIAP abrogated it, placing KIAP downstream of miR-23a in a pathway controlling hepatocarcinoma cell apoptosis and caspase-3 activation.\",\n      \"method\": \"miRNA transfection, siRNA knockdown, plasmid overexpression, Western blot, caspase-3 activity assay, flow cytometry\",\n      \"journal\": \"European review for medical and pharmacological sciences\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, single cell line, rescue experiment is indirect (miRNA target validation without direct binding confirmation), no orthogonal method\",\n      \"pmids\": [\"30280770\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"BIRC7 suppresses autophagy through modulating expression of ATG5 and BECN1; this autophagy suppression drives epithelial-mesenchymal transition (EMT) and metastasis in papillary thyroid carcinoma cells; combined targeting of BIRC7 and mTOR enhances autophagy and achieves synergistic anti-metastatic effects in vitro and in vivo.\",\n      \"method\": \"BIRC7 overexpression and siRNA knockdown, transmission electron microscopy, Western blot for ATG5/BECN1/EMT markers, wound-healing and invasion assays, xenograft tumor model\",\n      \"journal\": \"American journal of cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — loss- and gain-of-function with defined molecular readouts (ATG5, BECN1) and in vivo validation, single lab\",\n      \"pmids\": [\"32064154\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"BIRC7 is a transcriptional downstream target of HIF-1α in melanoma; its promoter contains HIF-1α binding sites confirmed by luciferase assay; BIRC7 knockdown reversed the pro-tumorigenic effects of HIF-1α under hypoxia.\",\n      \"method\": \"Luciferase reporter assay (HIF-1α binding to BIRC7 promoter), RNA-seq under hypoxia, qRT-PCR, Western blot, siRNA knockdown with cellular function assays\",\n      \"journal\": \"Clinical, cosmetic and investigational dermatology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — luciferase assay for transcriptional regulation plus knockdown phenotype, single lab, no ChIP or direct binding confirmation of HIF-1α to endogenous promoter\",\n      \"pmids\": [\"35747741\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"BIRC7 (ML-IAP/Livin) is a single-BIR/RING-domain IAP that inhibits apoptosis primarily by sequestering Smac/DIABLO via its BIR domain (preventing Smac from antagonizing XIAP), functions as a RING-type E3 ubiquitin ligase that dimerizes to optimally position donor ubiquitin on E2 UbcH5B for transfer, undergoes caspase-mediated cleavage at Asp52 to generate a paradoxically pro-apoptotic truncated form (tLivin), regulates C-RAF protein stability and cell migration in melanoma, and suppresses autophagy (via ATG5/BECN1) to promote EMT and metastasis; its expression is transcriptionally induced by HIF-1α under hypoxia.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"BIRC7 (ML-IAP/Livin) is a single-BIR/RING-domain inhibitor of apoptosis that restrains effector caspase activity and couples apoptotic control to ubiquitin signaling [#0, #6]. Its anti-apoptotic function depends on the integrity of its BIR domain, which is only a weak direct inhibitor of caspases 3 and 9; the dominant mechanism is high-affinity sequestration of Smac/DIABLO, which displaces Smac from XIAP and thereby preserves XIAP-mediated caspase inhibition [#1, #3]. Crystal structures of the BIR domain bound to Smac-derived and phage-derived peptides defined the peptide-binding groove and enabled selective high-affinity peptide antagonists [#2]. The RING domain confers E3 ubiquitin ligase activity: it dimerizes to lock the donor ubiquitin on the E2 UbcH5B~Ub conjugate in a transfer-competent orientation, and this same RING activity drives BIRC7 autodegradation in response to bivalent IAP antagonists [#6, #8]. BIRC7 is also processed during apoptosis by cleavage at Asp52 to a truncated form (tLivin) that paradoxically acquires pro-apoptotic activity, a conversion exploited by NK cells to limit tumor growth in vivo [#4, #5]. Beyond core apoptotic control, BIRC7 binds C-RAF and restrains its protein level, MAPK activation, and melanoma cell migration, and it suppresses autophagy through ATG5 and BECN1 to promote EMT and metastasis [#7, #10].\",\n  \"teleology\": [\n    {\n      \"year\": 2000,\n      \"claim\": \"Established BIRC7 as a domain-defined IAP and showed the BIR domain is necessary for anti-apoptotic function, framing the question of how a single-BIR protein blocks cell death.\",\n      \"evidence\": \"Deletion/mutational analysis with overexpression apoptosis assays in cell lines\",\n      \"pmids\": [\"11084335\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct caspase inhibition proposed but not reconstituted in vitro\", \"RING-domain function not addressed\", \"No structural basis for BIR activity\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Identified Smac/DIABLO as a high-affinity BIR-domain ligand, shifting the mechanistic model away from direct caspase inhibition toward Smac sequestration.\",\n      \"evidence\": \"Co-IP, Smac-peptide binding assays, single-residue mutagenesis, cell-death rescue\",\n      \"pmids\": [\"11801603\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not quantify the effect on endogenous XIAP-Smac equilibrium\", \"Structural interface not yet resolved\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Resolved the atomic structure of the BIR–peptide interface, enabling design of antagonists selective for BIRC7 over XIAP.\",\n      \"evidence\": \"X-ray crystallography, phage display, synthetic peptide SAR, affinity measurements\",\n      \"pmids\": [\"12846571\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structures use peptide surrogates rather than full-length Smac\", \"No structure of the RING/E3 module\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Directly tested the competing mechanisms, showing BIR is only a weak caspase inhibitor and that Smac sequestration disrupting the XIAP-Smac complex is the primary anti-apoptotic route.\",\n      \"evidence\": \"In vitro caspase assays, BIR engineering, co-IP of endogenous ML-IAP-Smac and XIAP-Smac complexes\",\n      \"pmids\": [\"15485396\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Quantitative contribution of residual direct caspase inhibition unresolved\", \"Does not address RING-dependent functions\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Mapped a defined apoptotic cleavage site at Asp52 and showed cleavage can occur by a non-canonical, partly caspase-independent protease, opening the question of how an IAP is itself processed during death.\",\n      \"evidence\": \"Asp52 mutagenesis, deletion mapping, Omi/HtrA2 RNAi, zVAD-fmk treatment across stimuli\",\n      \"pmids\": [\"16806840\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Identity of the responsible protease not established\", \"Functional consequence of cleavage not defined here\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Showed cleavage generates a truncated tLivin with paradoxical pro-apoptotic activity that NK cells exploit to suppress tumors, redefining BIRC7 as a context-dependent regulator of death.\",\n      \"evidence\": \"Caspase-site point mutants, NK-cell-depleted xenograft mouse models, in vivo growth assays\",\n      \"pmids\": [\"19549891\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular basis of tLivin pro-apoptotic gain-of-function unclear\", \"Single lab, limited tumor models\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Defined the catalytic mechanism of the RING E3 activity, showing dimerization positions the UbcH5B~Ub conjugate for ubiquitin transfer.\",\n      \"evidence\": \"Crystal structure of dimeric RING–UbcH5B~Ub, NMR, mutagenesis, in vitro ubiquitination\",\n      \"pmids\": [\"22902369\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Physiological substrates of the ligase not identified in this study\", \"Regulation of dimerization in cells unknown\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Linked RING activity to drug-induced self-destruction, showing bivalent IAP antagonists trigger BIRC7 autodegradation independent of c-IAP1/2.\",\n      \"evidence\": \"IAP antagonist treatment, proteasome rescue, RING mutants, c-IAP1/2 knockout/knockdown lines\",\n      \"pmids\": [\"22853455\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether degradation is autoubiquitination vs trans-ubiquitination not fully resolved\", \"E2 used in cells not pinpointed\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Extended BIRC7 function beyond apoptosis by showing it binds C-RAF and limits its stability, MAPK signaling, and melanoma migration.\",\n      \"evidence\": \"Co-IP, siRNA knockdown with C-RAF Western blot, MAPK and migration assays\",\n      \"pmids\": [\"22711539\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether C-RAF is a direct ubiquitination substrate not shown\", \"Single lab, reciprocal validation limited\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Placed BIRC7 downstream of miR-23a in controlling hepatocarcinoma apoptosis, adding an upstream regulatory layer.\",\n      \"evidence\": \"miRNA/siRNA transfection, overexpression rescue, caspase-3 assay, flow cytometry\",\n      \"pmids\": [\"30280770\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No direct miR-23a–BIRC7 binding confirmation\", \"Single cell line, no orthogonal method\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Connected BIRC7 to autophagy suppression via ATG5/BECN1 and to EMT/metastasis, with mTOR co-targeting as a synergistic vulnerability.\",\n      \"evidence\": \"Gain/loss-of-function, electron microscopy, ATG5/BECN1/EMT Westerns, invasion assays, xenografts\",\n      \"pmids\": [\"32064154\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism by which BIRC7 modulates ATG5/BECN1 expression undefined\", \"Single tumor type\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Identified BIRC7 as a HIF-1α transcriptional target, linking its expression to the hypoxic tumor microenvironment.\",\n      \"evidence\": \"Luciferase promoter reporter, hypoxia RNA-seq, qRT-PCR, siRNA knockdown phenotypes\",\n      \"pmids\": [\"35747741\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No ChIP confirming HIF-1α binding to endogenous promoter\", \"Single lab\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The endogenous substrate repertoire of the BIRC7 RING ligase and the protease that generates tLivin remain unidentified, leaving the link between its E3 activity and its non-apoptotic roles (C-RAF, autophagy) mechanistically open.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No validated physiological ubiquitination substrate\", \"tLivin-generating protease unidentified\", \"Whether C-RAF and autophagy effects are RING-dependent not established\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016874\", \"supporting_discovery_ids\": [6, 8]},\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [6]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [1, 3]},\n      {\"term_id\": \"GO:0140313\", \"supporting_discovery_ids\": [3]}\n    ],\n    \"localization\": [],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [0, 3, 5]},\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [6, 8]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"DIABLO\", \"UBE2D2\", \"RAF1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}