{"gene":"DPP9","run_date":"2026-04-28T17:46:03","timeline":{"discoveries":[{"year":2002,"finding":"DPP9 was identified as a novel cytosolic serine protease with a catalytic triad (Ser, Asp, His) and a GWSYG motif identical to DPP IV, placing it in the DPP IV gene family. It lacks transmembrane domains and a signal sequence, consistent with cytosolic localization, and in vitro translation produced a ~98 kDa protein.","method":"In silico identification, in vitro translation, SDS-PAGE, northern blot","journal":"Gene","confidence":"Medium","confidence_rationale":"Tier 2 — original identification with multiple biochemical methods in a single study","pmids":["12459266"],"is_preprint":false},{"year":2009,"finding":"DPP9 is rate-limiting for degradation of proline-containing peptides in the cytoplasm. DPP9 cleaves the natural substrate RU1(34-42) antigenic peptide in vitro, and DPP9 knockdown in intact cells results in increased antigen presentation of this peptide, demonstrating a role in antigen presentation via peptide turnover.","method":"In vitro peptidase assay, siRNA knockdown in intact cells, antigen presentation assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1-2 — in vitro assay with natural substrate plus functional cellular validation","pmids":["19667070"],"is_preprint":false},{"year":2009,"finding":"Bovine DPP9 purified from testes exists as the short-form isoform, is activated and stabilized by DTT, and loses activity upon alkylation (N-ethylmaleimide, iodoacetamide), indicating dependence on free cysteine residues for enzymatic activity. No evidence of glycosylation was found.","method":"Purification, MALDI-TOF/TOF MS, N-terminal sequencing, enzymatic activity assays with chemical modifiers","journal":"Biochimica et biophysica acta","confidence":"Medium","confidence_rationale":"Tier 1 — biochemical characterization of purified native enzyme, single lab","pmids":["20026260"],"is_preprint":false},{"year":2012,"finding":"DPP9 binds SUMO1 via a novel interaction site (not the canonical SIM motif) involving an extended arm structure flanking the substrate entry site. SUMO1 binding stimulates DPP9 enzymatic activity, and SUMO1 silencing reduces cytosolic prolyl-peptidase activity. Mutants in the SUMO1-binding arm are less catalytically active than wild-type DPP9.","method":"Co-IP, mutagenesis, in vitro activity assay, siRNA knockdown","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1-2 — multiple orthogonal methods including mutagenesis and activity assays in single study","pmids":["23152501"],"is_preprint":false},{"year":2016,"finding":"DPP8 and DPP9 inhibition by Val-boroPro (Talabostat) activates pro-caspase-1 independent of the inflammasome adaptor ASC in monocytes and macrophages. Activated pro-caspase-1 cleaves gasdermin D to induce pyroptosis without efficiently processing itself or IL-1β. Mice lacking caspase-1 do not show immune stimulation after Val-boroPro treatment.","method":"Pharmacological inhibition, caspase-1 knockout mice, gasdermin D cleavage assay, cell death assay","journal":"Nature chemical biology","confidence":"High","confidence_rationale":"Tier 1-2 — multiple orthogonal methods, genetic (KO mice) and biochemical validation, highly cited","pmids":["27820798"],"is_preprint":false},{"year":2016,"finding":"DPP9 cleaves Syk kinase to produce a neo N-terminus with serine at position 1, which destabilizes Syk via the N-end rule pathway. DPP9 interacts with Filamin A, which recruits DPP9 to Syk. DPP9 processing is a prerequisite for Cbl-mediated Syk ubiquitination, and DPP9 inhibition stabilizes Syk and modulates B-cell signaling.","method":"Co-IP, pulse-chase experiments, mutagenesis, siRNA knockdown, N-end rule pathway assays","journal":"eLife","confidence":"High","confidence_rationale":"Tier 1-2 — multiple orthogonal methods (Co-IP, mutagenesis, pulse-chase) establishing DPP9 as N-end rule pathway component","pmids":["27614019"],"is_preprint":false},{"year":2018,"finding":"DPP8/DPP9 inhibitors induce caspase-1-dependent pyroptosis in human myeloid cells via CARD8. CARD8 mediates DPP8/DPP9 inhibitor-induced pyroptosis in human myeloid cells, and these inhibitors inhibit AML progression in mouse xenograft models.","method":"Pharmacological inhibition, CARD8 knockdown/knockout, caspase-1 activity assays, AML mouse models","journal":"Nature medicine","confidence":"High","confidence_rationale":"Tier 2 — multiple cell types and genetic approaches, in vivo validation, highly cited","pmids":["29967349"],"is_preprint":false},{"year":2018,"finding":"DPP9 is a direct binding partner of NLRP1 and CARD8, interacting via the FIIND (Function to Find Domain). DPP9 acts as an endogenous inhibitor of the NLRP1 inflammasome. Both DPP9 catalytic activity and its scaffolding interaction with the NLRP1 FIIND act synergistically to maintain NLRP1 in its inactive state. A patient-derived NLRP1 FIIND missense mutation that abrogates DPP9 binding causes inflammasome hyperactivation.","method":"Proteomics screen, Co-IP, CRISPR/Cas9 knockout, small molecule inhibition, ASC speck formation assay, IL-1β secretion assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 — multiple orthogonal approaches (proteomics, Co-IP, genetic deletion, pharmacological), multiple cell types, patient mutation validation","pmids":["30291141"],"is_preprint":false},{"year":2018,"finding":"DPP9 enzyme activity in vivo controls survival of migratory tongue muscle progenitors. Knock-in mice expressing catalytically inactive DPP9 (S729A) display microglossia due to increased apoptosis of occipital somite-derived muscle progenitors, leading to suckling defect and neonatal lethality.","method":"Knock-in mouse model (S729A catalytic mutant), histology, apoptosis assays, developmental analysis","journal":"Developmental biology","confidence":"High","confidence_rationale":"Tier 2 — catalytic mutant knock-in with specific phenotypic readout demonstrating enzymatic activity is required in vivo","pmids":["28887018"],"is_preprint":false},{"year":2018,"finding":"DPP9 inhibition (by saxagliptin or TC-E5007) in cardiomyocytes impairs CaMKII-phospholamban signaling and PKC activity. Knockdown of DPP9 (but not DPP8) recapitulates these effects, implicating DPP9 specifically in cardiac CaMKII/PKC signaling. Saxagliptin impairs Ca2+ transient relaxation and prolongs action potential duration via DPP9 inhibition.","method":"Pharmacological inhibition, siRNA knockdown, CaMKII/PKC activity assays, Ca2+ transient measurements, action potential recordings","journal":"Frontiers in physiology","confidence":"Medium","confidence_rationale":"Tier 2 — siRNA knockdown combined with pharmacological approach and functional readouts, single lab","pmids":["30487758"],"is_preprint":false},{"year":2019,"finding":"DPP9's catalytic activity, not its binding to CARD8, restrains the CARD8 inflammasome. Wild-type but not catalytically inactive DPP9 rescues CARD8-mediated cell death in DPP9-knockout cells. The DPP9-CARD8 interaction is not disrupted by DPP9 inhibitors, unlike the DPP9-NLRP1 interaction.","method":"Activity-based probes, reconstituted inflammasome assays, mass spectrometry proteomics, DPP9 knockout cells, catalytic mutant rescue","journal":"ACS chemical biology","confidence":"High","confidence_rationale":"Tier 1-2 — reconstituted assays plus catalytic mutant rescue in KO cells, multiple orthogonal methods","pmids":["31525884"],"is_preprint":false},{"year":2020,"finding":"DPP8/DPP9 mediate N-terminal processing of adenylate kinase 2 (AK2), a mitochondrial protein lacking an N-terminal targeting sequence. DPP9-mediated processing of AK2 induces its rapid proteasomal degradation and prevents cytosolic accumulation of enzymatically active AK2, thus regulating the competition between mitochondrial import and proteasomal degradation.","method":"siRNA knockdown, proteasome inhibition, biochemical fractionation, pulse-chase analysis, proteomic identification of DPP8/9 substrates","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1-2 — multiple orthogonal methods establishing substrate processing and downstream fate, in vitro and cellular validation","pmids":["32815200"],"is_preprint":false},{"year":2021,"finding":"Cryo-EM structures of the human NLRP1-DPP9 complex reveal a ternary complex comprising DPP9, full-length NLRP1, and the NLRP1 C-terminal fragment (CT). The N-terminus of the NLRP1 CT inserts into the DPP9 active site. Val-boroPro (VbP) disrupts this interaction, weakening the NLRP1-DPP9 complex and accelerating degradation of the NLRP1 N-terminal fragment to activate the inflammasome. Full-length NLRP1 is required for NLRP1 CT binding to DPP9, suggesting inflammasome activation is regulated by the ratio of NLRP1 CT to full-length NLRP1.","method":"Cryo-EM structure determination, biochemical binding assays, ectopic expression rescue experiments, VbP inhibitor studies","journal":"Nature","confidence":"High","confidence_rationale":"Tier 1 — cryo-EM structure plus functional validation with mutagenesis and rescue experiments","pmids":["33731932"],"is_preprint":false},{"year":2021,"finding":"Structural and biochemical analysis of rat NLRP1-DPP9 reveals a 2:1 complex containing one autoinhibited full-length NLRP1 and one active UPA-CARD fragment of NLRP1 bound to DPP9. The NLRP1 ZU5 domain is required for both autoinhibition and 2:1 complex assembly. Complex formation prevents UPA-mediated higher-order oligomerization and strengthens ZU5-mediated autoinhibition. Both NLRP1 binding and enzymatic activity of DPP9 are required to suppress NLRP1 inflammasome in human cells.","method":"Cryo-EM, X-ray crystallography, biochemical reconstitution, mutagenesis, functional inflammasome assays","journal":"Nature","confidence":"High","confidence_rationale":"Tier 1 — structural determination combined with biochemical and functional validation, multiple orthogonal methods","pmids":["33731929"],"is_preprint":false},{"year":2022,"finding":"The NLRP1 disease variant M1184V stabilizes the FIIND domain in a monomeric conformation, promotes autoproteolysis, enhances DPP9 binding (as shown by surface plasmon resonance and immunoprecipitation), and leads to improved formation of an autoinhibited complex with DPP9.","method":"Size-exclusion chromatography, surface plasmon resonance, molecular dynamics simulation, immunoprecipitation, activity assays","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 — multiple biophysical and biochemical methods, single lab","pmids":["36309085"],"is_preprint":false},{"year":2023,"finding":"DPP9 binds to KEAP1 via a conserved ESGE motif and disrupts KEAP1-NRF2 binding by competing with NRF2 for KEAP1 in an enzyme-independent manner. DPP9 overexpression stabilizes NRF2, drives NRF2-dependent transcription, decreases cellular ROS, suppresses ferroptosis, and induces sorafenib resistance in ccRCC cells via upregulation of the NRF2 target SLC7A11.","method":"Protein affinity purification, Co-IP, mutagenesis (ESGE motif), cell viability assays, ROS measurement, ferroptosis assays","journal":"Cancer research","confidence":"Medium","confidence_rationale":"Tier 2 — Co-IP/pulldown with motif mutagenesis and multiple functional readouts, single lab","pmids":["37713596"],"is_preprint":false},{"year":2023,"finding":"DPP9 and DPP8 function as amino-dipeptidyl peptidases that remove N-terminal Xaa-Pro dipeptides. Confirmed substrates include Syk, AK2, and BRCA2; N-terminal processing of these proteins by DPP9 triggers their rapid proteasomal degradation via the N-degron pathway.","method":"Enzymatic assays, substrate identification, proteomics (review/methods chapter with original data on purification)","journal":"Methods in enzymology","confidence":"Medium","confidence_rationale":"Tier 2 — consolidates multiple substrate identifications with enzymatic and cellular validation across independent studies","pmids":["37230592"],"is_preprint":false},{"year":2023,"finding":"A de novo DPP9 mutation (p.Arg252Pro) destabilizes the DPP9 protein and abolishes its ability to restrain NLRP1 and CARD8 inflammasomes in HEK293T cells and patient-derived iPSCs, resulting in constitutive inflammasome activation and severe autoinflammation, demonstrating that DPP9 protein stability is required for inflammasome suppression.","method":"Heterologous expression in HEK293T, patient-derived iPSCs, inflammasome activation assays, protein stability assessment","journal":"The Journal of allergy and clinical immunology","confidence":"Medium","confidence_rationale":"Tier 2 — patient mutation functionally validated in multiple cell systems with direct inflammasome readouts","pmids":["37544411"],"is_preprint":false},{"year":2024,"finding":"KEAP1 binds DPP9 in an inactive (non-native) conformation and stabilizes this misfolded state. Reciprocally, this inactive form of DPP9 inhibits KEAP1-mediated NRF2 degradation by preventing KEAP1-NRF2 interaction, thereby inducing an antioxidant response. This reveals an endogenous mechanism for DPP9 inhibition linked to the intracellular redox state.","method":"Co-IP, biochemical binding assays, NRF2 stability assays, DPP9 enzymatic activity assays under various conditions","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 — reciprocal inhibition demonstrated biochemically, but key physiological trigger not yet identified, single lab","pmids":["39615677"],"is_preprint":false},{"year":2024,"finding":"DPP9 inhibition in liver cancer cells reduces NRF2 stability by releasing KEAP1 from DPP9 binding, thereby increasing KEAP1-mediated NRF2 ubiquitination. DPP9 overexpression up-regulates NQO1 via NRF2 and inhibits intracellular ROS, promoting chemoresistance; NQO1 inhibition by dicoumarol reverses this.","method":"siRNA knockdown, overexpression, ubiquitination assay, ROS measurement, cell viability assay with chemotherapy drugs","journal":"Redox biology","confidence":"Medium","confidence_rationale":"Tier 2 — mechanistic pathway established by KD/OE with multiple readouts, consistent with prior KEAP1-DPP9 finding","pmids":["39094401"],"is_preprint":false},{"year":2025,"finding":"SFTSV non-structural protein (NSs) activates NLRP1 and CARD8 inflammasomes by disrupting the DPP9-mediated inhibitory ternary complex: NSs interacts with NLRP1/CARD8 FIIND domains (competing with DPP8/9 binding) and promotes degradation of DPP8/DPP9, releasing activated C-terminal fragments.","method":"Primary keratinocyte and macrophage infection assays, Co-IP, inflammasome activation assays, CARD8 deletion studies","journal":"PLoS pathogens","confidence":"Medium","confidence_rationale":"Tier 2 — Co-IP with functional inflammasome readouts in primary cells and genetic validation, single lab","pmids":["40608794"],"is_preprint":false},{"year":2026,"finding":"Proximity labeling (TurboID) of DPP9 in DPP9-knockout cells identified novel DPP9 interactors including DPP8, the E3 ligase CBL, the deubiquitinase complex CYLD-SPATA2, and the BRISC complex (BRCC36/BRCC3 and ABRO1/ABRAXAS2). NanoBRET assays demonstrated that DPP9 disrupts BRCC36-ABRO1 binding and CYLD-SPATA2 binding, revealing non-catalytic scaffolding functions of DPP9 in the ubiquitin system.","method":"TurboID proximity labeling, mass spectrometry, Co-IP validation, NanoBRET assays in living cells","journal":"Cellular and molecular life sciences","confidence":"Medium","confidence_rationale":"Tier 2 — proximity labeling with orthogonal validation (Co-IP, NanoBRET), single lab","pmids":["41636814"],"is_preprint":false},{"year":2026,"finding":"DPP9 disrupts the BRISC-SHMT2 complex, enhancing BRISC-mediated deubiquitination of IFNAR1, which activates JAK/STAT signaling and drives PD-L1 transcription in ccRCC. DPP9 inhibition with 1G244 reverses this by reducing DPP9-SHMT2 interaction, promoting IFNAR1 ubiquitination and degradation, and restoring T cell cytotoxicity.","method":"Co-IP, pharmacological inhibition (1G244), IFNAR1 ubiquitination assays, JAK/STAT pathway assays, T cell cytotoxicity assays, in vivo mouse models","journal":"Cell death and differentiation","confidence":"Medium","confidence_rationale":"Tier 2 — mechanistic pathway established with Co-IP, ubiquitination assays, and functional readouts in vitro and in vivo","pmids":["41826729"],"is_preprint":false}],"current_model":"DPP9 is a cytosolic serine dipeptidyl peptidase that removes N-terminal Xaa-Pro dipeptides from substrates (including Syk, AK2, BRCA2) to regulate their stability via the N-degron/proteasomal pathway, acts as a critical checkpoint for NLRP1 and CARD8 inflammasome activation by forming inhibitory ternary complexes in which the NLRP1/CARD8 C-terminal fragment's N-terminus inserts into the DPP9 active site, and additionally exerts non-catalytic scaffolding functions by interacting with partners such as KEAP1 (modulating NRF2-driven antioxidant responses), the BRISC deubiquitinase complex (regulating IFNAR1 stability and PD-L1 expression), and ubiquitin E3/deubiquitinase complexes (CBL, CYLD-SPATA2), with both its enzymatic activity and protein-binding functions being required for full physiological function."},"narrative":{"teleology":[{"year":2002,"claim":"Establishing DPP9 as a new member of the DPP IV serine protease family with cytosolic localization resolved its gene family membership and predicted intracellular rather than extracellular substrates.","evidence":"In silico identification, in vitro translation, SDS-PAGE, and northern blot showing ~98 kDa protein lacking transmembrane domains","pmids":["12459266"],"confidence":"Medium","gaps":["No endogenous substrates identified","Catalytic activity not demonstrated against peptide substrates","Cellular localization not confirmed by imaging"]},{"year":2009,"claim":"Demonstrating that DPP9 is rate-limiting for cytoplasmic proline-containing peptide degradation and influences MHC-I antigen presentation established its first defined cellular function.","evidence":"In vitro cleavage of RU1(34-42) peptide plus siRNA knockdown showing enhanced antigen presentation in intact cells","pmids":["19667070"],"confidence":"High","gaps":["Protein substrates not yet identified","In vivo relevance to immune surveillance not tested"]},{"year":2012,"claim":"Discovery that SUMO1 binds DPP9 via a non-canonical site on an extended arm near the substrate entry region and stimulates its enzymatic activity revealed the first allosteric regulator of DPP9.","evidence":"Co-IP, mutagenesis of SUMO1-binding arm, in vitro activity assays, and SUMO1 siRNA knockdown reducing cytosolic prolyl-peptidase activity","pmids":["23152501"],"confidence":"High","gaps":["Structural basis of SUMO1-DPP9 interaction not resolved at atomic level","Physiological conditions modulating SUMO1-DPP9 interaction unknown"]},{"year":2016,"claim":"Linking DPP8/DPP9 inhibition to caspase-1-dependent pyroptosis and gasdermin D cleavage in monocytes identified the inflammasome axis as a major DPP9-regulated pathway, and demonstrating that DPP9 cleaves Syk to generate an N-degron established its role in targeted protein degradation.","evidence":"Val-boroPro treatment of monocytes/macrophages and caspase-1 KO mice for inflammasome; Co-IP, pulse-chase, and mutagenesis for Syk N-terminal processing and Cbl-mediated ubiquitination","pmids":["27820798","27614019"],"confidence":"High","gaps":["DPP8 vs DPP9 individual contributions to pyroptosis not resolved","Inflammasome sensor upstream of caspase-1 not yet identified","Full scope of N-degron substrates unknown"]},{"year":2018,"claim":"Identifying CARD8 as the sensor mediating DPP8/9 inhibitor-induced pyroptosis in human myeloid cells, and demonstrating DPP9 as a direct binding partner and endogenous inhibitor of both NLRP1 and CARD8 inflammasomes via the FIIND domain, resolved the upstream sensor question and established DPP9 as a dual inflammasome checkpoint.","evidence":"CARD8 KO/knockdown with pharmacological inhibition in myeloid cells; proteomics screen, Co-IP, CRISPR KO, and patient NLRP1 FIIND mutation abrogating DPP9 binding","pmids":["29967349","30291141"],"confidence":"High","gaps":["Structural basis of DPP9-FIIND interaction not yet resolved","Relative contributions of DPP9 catalytic vs scaffolding functions not disentangled"]},{"year":2018,"claim":"Knock-in mice expressing catalytically inactive DPP9 (S729A) demonstrated that DPP9 enzymatic activity is essential in vivo for survival of migratory tongue muscle progenitors, establishing a non-redundant developmental role.","evidence":"S729A knock-in mice showing microglossia, increased apoptosis of occipital somite progenitors, suckling defect, and neonatal lethality","pmids":["28887018"],"confidence":"High","gaps":["Substrate(s) responsible for muscle progenitor phenotype not identified","Whether inflammasome dysregulation contributes to neonatal lethality not tested"]},{"year":2020,"claim":"Identification of AK2 as a DPP9 substrate showed that DPP9-mediated N-terminal processing diverts proteins from mitochondrial import to proteasomal degradation, broadening DPP9's role beyond signaling kinases to organellar protein quality control.","evidence":"siRNA knockdown, proteasome inhibition, pulse-chase, and biochemical fractionation demonstrating DPP9 processing triggers AK2 degradation and prevents cytosolic accumulation","pmids":["32815200"],"confidence":"High","gaps":["Full inventory of DPP9 N-degron substrates still incomplete","Whether DPP9 regulates import of other mitochondrial proteins unknown"]},{"year":2021,"claim":"Cryo-EM structures of the NLRP1-DPP9 ternary complex revealed the atomic mechanism of inflammasome suppression: the NLRP1 C-terminal fragment N-terminus inserts into the DPP9 active site, and full-length NLRP1 is required to scaffold this interaction, explaining how VbP disrupts the complex to activate inflammasome signaling.","evidence":"Cryo-EM structures (human and rat), biochemical reconstitution, mutagenesis, and functional inflammasome assays","pmids":["33731932","33731929"],"confidence":"High","gaps":["High-resolution structure of CARD8-DPP9 complex not determined","Endogenous signals that shift the NLRP1 CT/full-length ratio in vivo remain elusive"]},{"year":2023,"claim":"Discovery that DPP9 binds KEAP1 via an ESGE motif to competitively displace NRF2, stabilizing the antioxidant response independently of DPP9 catalytic activity, revealed a major enzyme-independent scaffolding function linking DPP9 to redox homeostasis and chemoresistance.","evidence":"Co-IP with ESGE motif mutagenesis, NRF2 stability and transcription assays, ROS measurement, and ferroptosis/sorafenib resistance assays in ccRCC cells","pmids":["37713596"],"confidence":"Medium","gaps":["Physiological trigger switching DPP9 between active and KEAP1-bound inactive conformations not defined","In vivo validation of DPP9-KEAP1 axis in animal models lacking","Whether DPP9-KEAP1 interaction is tissue-specific unknown"]},{"year":2023,"claim":"A de novo DPP9 mutation (p.Arg252Pro) causing severe autoinflammation in a patient demonstrated that DPP9 loss of function is sufficient to constitutively activate NLRP1 and CARD8 inflammasomes in humans, establishing DPP9 deficiency as a Mendelian autoinflammatory condition.","evidence":"Functional validation in HEK293T cells and patient-derived iPSCs showing protein destabilization and constitutive inflammasome activation","pmids":["37544411"],"confidence":"Medium","gaps":["Single patient reported; additional patients needed to define genotype-phenotype spectrum","Whether partial loss of DPP9 activity confers intermediate inflammatory phenotypes unknown"]},{"year":2024,"claim":"Reciprocal biochemical analysis showed that KEAP1 preferentially binds inactive/misfolded DPP9, stabilizing this non-native form, while inactive DPP9 in turn blocks KEAP1-NRF2 interaction—establishing a redox-sensitive feedback loop between DPP9 conformational state and antioxidant signaling.","evidence":"Co-IP, DPP9 activity assays under various conditions, NRF2 stability assays; DPP9 KD/OE with ubiquitination and ROS readouts in liver cancer cells","pmids":["39615677","39094401"],"confidence":"Medium","gaps":["Identity of the cellular signal driving DPP9 misfolding/inactivation not established","Structural basis of KEAP1 recognizing inactive DPP9 not resolved"]},{"year":2025,"claim":"Demonstration that SFTSV NSs protein activates NLRP1/CARD8 inflammasomes by competing with DPP9 for FIIND binding and promoting DPP9 degradation established viral hijacking of the DPP9 inflammasome checkpoint as a pathogenic mechanism.","evidence":"Primary keratinocyte and macrophage infection, Co-IP, inflammasome activation assays, CARD8 deletion studies","pmids":["40608794"],"confidence":"Medium","gaps":["Whether other viruses exploit the same DPP9-displacement strategy unknown","Structural basis of NSs-FIIND interaction not resolved"]},{"year":2026,"claim":"Proximity labeling and NanoBRET identified DPP9 as a non-catalytic disruptor of BRISC (BRCC36-ABRO1) and CYLD-SPATA2 deubiquitinase complexes, and functional studies showed DPP9 enhances BRISC-mediated IFNAR1 deubiquitination to drive JAK/STAT-dependent PD-L1 expression, revealing a scaffolding role in immune checkpoint regulation.","evidence":"TurboID proximity labeling, Co-IP, NanoBRET in living cells; IFNAR1 ubiquitination assays, JAK/STAT pathway assays, T cell cytotoxicity assays, and in vivo mouse models with DPP9 inhibitor 1G244","pmids":["41636814","41826729"],"confidence":"Medium","gaps":["Direct structural interface between DPP9 and BRISC components not mapped","Whether DPP9-BRISC interaction is catalytic-activity-dependent or independent not fully resolved","In vivo immunotherapy efficacy of DPP9 inhibition not tested in clinical setting"]},{"year":null,"claim":"Major open questions include the complete inventory of physiological DPP9 N-degron substrates, the structural basis of the CARD8-DPP9 complex, the endogenous signals that toggle DPP9 between active and inactive/KEAP1-bound states, and whether DPP9's scaffolding roles in ubiquitin signaling and inflammasome suppression are coordinated or independent pathways.","evidence":"","pmids":[],"confidence":"Low","gaps":["Full substrate repertoire via unbiased proteomics not yet reported","CARD8-DPP9 complex structure not determined","Physiological trigger for DPP9 conformational switching unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[1,2,5,11,16]},{"term_id":"GO:0016787","term_label":"hydrolase activity","supporting_discovery_ids":[0,1,2,5,11,16]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[7,10,12,13,15,18,22]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[15,21,22]}],"localization":[{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[0,1,11]}],"pathway":[{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[1,4,6,7,12,13,17,20,22]},{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[4,6,8]},{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[5,11,16]},{"term_id":"R-HSA-8953897","term_label":"Cellular responses to stimuli","supporting_discovery_ids":[15,18,19]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[9,15,22]}],"complexes":["NLRP1-DPP9 ternary complex","CARD8-DPP9 complex"],"partners":["NLRP1","CARD8","KEAP1","SUMO1","FLNA","CBL","BRCC3","ABRO1"],"other_free_text":[]},"mechanistic_narrative":"DPP9 is a cytosolic serine dipeptidyl peptidase that cleaves N-terminal Xaa-Pro dipeptides from substrates including Syk, AK2, and BRCA2, thereby targeting them for proteasomal degradation via the N-degron pathway, and additionally functions as a non-catalytic scaffold that modulates inflammasome activation, redox homeostasis, and ubiquitin signaling [PMID:27614019, PMID:32815200, PMID:37230592]. Cryo-EM structures reveal that DPP9 forms an inhibitory ternary complex with full-length NLRP1 and the NLRP1 C-terminal fragment—whose N-terminus inserts into the DPP9 active site—and both catalytic activity and scaffolding binding are required to suppress NLRP1 and CARD8 inflammasome activation; disruption of this complex by pharmacological inhibitors or viral proteins triggers pyroptosis [PMID:33731932, PMID:33731929, PMID:30291141, PMID:40608794]. Independent of its peptidase activity, DPP9 competes with NRF2 for KEAP1 binding via a conserved ESGE motif, stabilizing NRF2 to drive antioxidant gene expression and suppress ferroptosis, and disrupts the BRISC deubiquitinase complex to enhance IFNAR1-mediated JAK/STAT signaling and PD-L1 expression [PMID:37713596, PMID:39615677, PMID:41826729]. De novo loss-of-function DPP9 mutations cause constitutive NLRP1/CARD8 inflammasome activation and severe autoinflammation in humans [PMID:37544411]."},"prefetch_data":{"uniprot":{"accession":"Q86TI2","full_name":"Dipeptidyl peptidase 9","aliases":["Dipeptidyl peptidase IV-related protein 2","DPRP-2","Dipeptidyl peptidase IX","DPP IX","Dipeptidyl peptidase-like protein 9","DPLP9"],"length_aa":863,"mass_kda":98.3,"function":"Dipeptidyl peptidase that cleaves off N-terminal dipeptides from proteins having a Pro or Ala residue at position 2 (PubMed:12662155, PubMed:16475979, PubMed:19667070, PubMed:29382749, PubMed:30291141, PubMed:33731929, PubMed:36112693). Acts as a key inhibitor of caspase-1-dependent monocyte and macrophage pyroptosis in resting cells by preventing activation of NLRP1 and CARD8 (PubMed:27820798, PubMed:29967349, PubMed:30291141, PubMed:31525884, PubMed:32796818, PubMed:36112693, PubMed:36357533). Sequesters the cleaved C-terminal part of NLRP1 and CARD8, which respectively constitute the active part of the NLRP1 and CARD8 inflammasomes, in a ternary complex, thereby preventing their oligomerization and activation (PubMed:33731929, PubMed:33731932, PubMed:34019797). The dipeptidyl peptidase activity is required to suppress NLRP1 and CARD8; however, neither NLRP1 nor CARD8 are bona fide substrates of DPP9, suggesting the existence of substrate(s) required for NLRP1 and CARD8 inhibition (PubMed:33731929)","subcellular_location":"Nucleus","url":"https://www.uniprot.org/uniprotkb/Q86TI2/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/DPP9","classification":"Not Classified","n_dependent_lines":4,"n_total_lines":1208,"dependency_fraction":0.0033112582781456954},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/DPP9","total_profiled":1310},"omim":[{"mim_id":"620331","title":"HATIPOGLU IMMUNODEFICIENCY SYNDROME; HATIS","url":"https://www.omim.org/entry/620331"},{"mim_id":"609051","title":"CASPASE RECRUITMENT DOMAIN-CONTAINING PROTEIN 8; CARD8","url":"https://www.omim.org/entry/609051"},{"mim_id":"608258","title":"DIPEPTIDYL PEPTIDASE IX; DPP9","url":"https://www.omim.org/entry/608258"},{"mim_id":"606819","title":"DIPEPTIDYL PEPTIDASE VIII; DPP8","url":"https://www.omim.org/entry/606819"},{"mim_id":"606636","title":"NLR FAMILY, PYRIN DOMAIN-CONTAINING 1; NLRP1","url":"https://www.omim.org/entry/606636"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Cytosol","reliability":"Supported"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in 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It lacks transmembrane domains and a signal sequence, consistent with cytosolic localization, and in vitro translation produced a ~98 kDa protein.\",\n      \"method\": \"In silico identification, in vitro translation, SDS-PAGE, northern blot\",\n      \"journal\": \"Gene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — original identification with multiple biochemical methods in a single study\",\n      \"pmids\": [\"12459266\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"DPP9 is rate-limiting for degradation of proline-containing peptides in the cytoplasm. DPP9 cleaves the natural substrate RU1(34-42) antigenic peptide in vitro, and DPP9 knockdown in intact cells results in increased antigen presentation of this peptide, demonstrating a role in antigen presentation via peptide turnover.\",\n      \"method\": \"In vitro peptidase assay, siRNA knockdown in intact cells, antigen presentation assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — in vitro assay with natural substrate plus functional cellular validation\",\n      \"pmids\": [\"19667070\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Bovine DPP9 purified from testes exists as the short-form isoform, is activated and stabilized by DTT, and loses activity upon alkylation (N-ethylmaleimide, iodoacetamide), indicating dependence on free cysteine residues for enzymatic activity. No evidence of glycosylation was found.\",\n      \"method\": \"Purification, MALDI-TOF/TOF MS, N-terminal sequencing, enzymatic activity assays with chemical modifiers\",\n      \"journal\": \"Biochimica et biophysica acta\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 — biochemical characterization of purified native enzyme, single lab\",\n      \"pmids\": [\"20026260\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"DPP9 binds SUMO1 via a novel interaction site (not the canonical SIM motif) involving an extended arm structure flanking the substrate entry site. SUMO1 binding stimulates DPP9 enzymatic activity, and SUMO1 silencing reduces cytosolic prolyl-peptidase activity. Mutants in the SUMO1-binding arm are less catalytically active than wild-type DPP9.\",\n      \"method\": \"Co-IP, mutagenesis, in vitro activity assay, siRNA knockdown\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — multiple orthogonal methods including mutagenesis and activity assays in single study\",\n      \"pmids\": [\"23152501\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"DPP8 and DPP9 inhibition by Val-boroPro (Talabostat) activates pro-caspase-1 independent of the inflammasome adaptor ASC in monocytes and macrophages. Activated pro-caspase-1 cleaves gasdermin D to induce pyroptosis without efficiently processing itself or IL-1β. Mice lacking caspase-1 do not show immune stimulation after Val-boroPro treatment.\",\n      \"method\": \"Pharmacological inhibition, caspase-1 knockout mice, gasdermin D cleavage assay, cell death assay\",\n      \"journal\": \"Nature chemical biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — multiple orthogonal methods, genetic (KO mice) and biochemical validation, highly cited\",\n      \"pmids\": [\"27820798\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"DPP9 cleaves Syk kinase to produce a neo N-terminus with serine at position 1, which destabilizes Syk via the N-end rule pathway. DPP9 interacts with Filamin A, which recruits DPP9 to Syk. DPP9 processing is a prerequisite for Cbl-mediated Syk ubiquitination, and DPP9 inhibition stabilizes Syk and modulates B-cell signaling.\",\n      \"method\": \"Co-IP, pulse-chase experiments, mutagenesis, siRNA knockdown, N-end rule pathway assays\",\n      \"journal\": \"eLife\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — multiple orthogonal methods (Co-IP, mutagenesis, pulse-chase) establishing DPP9 as N-end rule pathway component\",\n      \"pmids\": [\"27614019\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"DPP8/DPP9 inhibitors induce caspase-1-dependent pyroptosis in human myeloid cells via CARD8. CARD8 mediates DPP8/DPP9 inhibitor-induced pyroptosis in human myeloid cells, and these inhibitors inhibit AML progression in mouse xenograft models.\",\n      \"method\": \"Pharmacological inhibition, CARD8 knockdown/knockout, caspase-1 activity assays, AML mouse models\",\n      \"journal\": \"Nature medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — multiple cell types and genetic approaches, in vivo validation, highly cited\",\n      \"pmids\": [\"29967349\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"DPP9 is a direct binding partner of NLRP1 and CARD8, interacting via the FIIND (Function to Find Domain). DPP9 acts as an endogenous inhibitor of the NLRP1 inflammasome. Both DPP9 catalytic activity and its scaffolding interaction with the NLRP1 FIIND act synergistically to maintain NLRP1 in its inactive state. A patient-derived NLRP1 FIIND missense mutation that abrogates DPP9 binding causes inflammasome hyperactivation.\",\n      \"method\": \"Proteomics screen, Co-IP, CRISPR/Cas9 knockout, small molecule inhibition, ASC speck formation assay, IL-1β secretion assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — multiple orthogonal approaches (proteomics, Co-IP, genetic deletion, pharmacological), multiple cell types, patient mutation validation\",\n      \"pmids\": [\"30291141\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"DPP9 enzyme activity in vivo controls survival of migratory tongue muscle progenitors. Knock-in mice expressing catalytically inactive DPP9 (S729A) display microglossia due to increased apoptosis of occipital somite-derived muscle progenitors, leading to suckling defect and neonatal lethality.\",\n      \"method\": \"Knock-in mouse model (S729A catalytic mutant), histology, apoptosis assays, developmental analysis\",\n      \"journal\": \"Developmental biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — catalytic mutant knock-in with specific phenotypic readout demonstrating enzymatic activity is required in vivo\",\n      \"pmids\": [\"28887018\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"DPP9 inhibition (by saxagliptin or TC-E5007) in cardiomyocytes impairs CaMKII-phospholamban signaling and PKC activity. Knockdown of DPP9 (but not DPP8) recapitulates these effects, implicating DPP9 specifically in cardiac CaMKII/PKC signaling. Saxagliptin impairs Ca2+ transient relaxation and prolongs action potential duration via DPP9 inhibition.\",\n      \"method\": \"Pharmacological inhibition, siRNA knockdown, CaMKII/PKC activity assays, Ca2+ transient measurements, action potential recordings\",\n      \"journal\": \"Frontiers in physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — siRNA knockdown combined with pharmacological approach and functional readouts, single lab\",\n      \"pmids\": [\"30487758\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"DPP9's catalytic activity, not its binding to CARD8, restrains the CARD8 inflammasome. Wild-type but not catalytically inactive DPP9 rescues CARD8-mediated cell death in DPP9-knockout cells. The DPP9-CARD8 interaction is not disrupted by DPP9 inhibitors, unlike the DPP9-NLRP1 interaction.\",\n      \"method\": \"Activity-based probes, reconstituted inflammasome assays, mass spectrometry proteomics, DPP9 knockout cells, catalytic mutant rescue\",\n      \"journal\": \"ACS chemical biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — reconstituted assays plus catalytic mutant rescue in KO cells, multiple orthogonal methods\",\n      \"pmids\": [\"31525884\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"DPP8/DPP9 mediate N-terminal processing of adenylate kinase 2 (AK2), a mitochondrial protein lacking an N-terminal targeting sequence. DPP9-mediated processing of AK2 induces its rapid proteasomal degradation and prevents cytosolic accumulation of enzymatically active AK2, thus regulating the competition between mitochondrial import and proteasomal degradation.\",\n      \"method\": \"siRNA knockdown, proteasome inhibition, biochemical fractionation, pulse-chase analysis, proteomic identification of DPP8/9 substrates\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — multiple orthogonal methods establishing substrate processing and downstream fate, in vitro and cellular validation\",\n      \"pmids\": [\"32815200\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Cryo-EM structures of the human NLRP1-DPP9 complex reveal a ternary complex comprising DPP9, full-length NLRP1, and the NLRP1 C-terminal fragment (CT). The N-terminus of the NLRP1 CT inserts into the DPP9 active site. Val-boroPro (VbP) disrupts this interaction, weakening the NLRP1-DPP9 complex and accelerating degradation of the NLRP1 N-terminal fragment to activate the inflammasome. Full-length NLRP1 is required for NLRP1 CT binding to DPP9, suggesting inflammasome activation is regulated by the ratio of NLRP1 CT to full-length NLRP1.\",\n      \"method\": \"Cryo-EM structure determination, biochemical binding assays, ectopic expression rescue experiments, VbP inhibitor studies\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — cryo-EM structure plus functional validation with mutagenesis and rescue experiments\",\n      \"pmids\": [\"33731932\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Structural and biochemical analysis of rat NLRP1-DPP9 reveals a 2:1 complex containing one autoinhibited full-length NLRP1 and one active UPA-CARD fragment of NLRP1 bound to DPP9. The NLRP1 ZU5 domain is required for both autoinhibition and 2:1 complex assembly. Complex formation prevents UPA-mediated higher-order oligomerization and strengthens ZU5-mediated autoinhibition. Both NLRP1 binding and enzymatic activity of DPP9 are required to suppress NLRP1 inflammasome in human cells.\",\n      \"method\": \"Cryo-EM, X-ray crystallography, biochemical reconstitution, mutagenesis, functional inflammasome assays\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — structural determination combined with biochemical and functional validation, multiple orthogonal methods\",\n      \"pmids\": [\"33731929\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"The NLRP1 disease variant M1184V stabilizes the FIIND domain in a monomeric conformation, promotes autoproteolysis, enhances DPP9 binding (as shown by surface plasmon resonance and immunoprecipitation), and leads to improved formation of an autoinhibited complex with DPP9.\",\n      \"method\": \"Size-exclusion chromatography, surface plasmon resonance, molecular dynamics simulation, immunoprecipitation, activity assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — multiple biophysical and biochemical methods, single lab\",\n      \"pmids\": [\"36309085\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"DPP9 binds to KEAP1 via a conserved ESGE motif and disrupts KEAP1-NRF2 binding by competing with NRF2 for KEAP1 in an enzyme-independent manner. DPP9 overexpression stabilizes NRF2, drives NRF2-dependent transcription, decreases cellular ROS, suppresses ferroptosis, and induces sorafenib resistance in ccRCC cells via upregulation of the NRF2 target SLC7A11.\",\n      \"method\": \"Protein affinity purification, Co-IP, mutagenesis (ESGE motif), cell viability assays, ROS measurement, ferroptosis assays\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — Co-IP/pulldown with motif mutagenesis and multiple functional readouts, single lab\",\n      \"pmids\": [\"37713596\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"DPP9 and DPP8 function as amino-dipeptidyl peptidases that remove N-terminal Xaa-Pro dipeptides. Confirmed substrates include Syk, AK2, and BRCA2; N-terminal processing of these proteins by DPP9 triggers their rapid proteasomal degradation via the N-degron pathway.\",\n      \"method\": \"Enzymatic assays, substrate identification, proteomics (review/methods chapter with original data on purification)\",\n      \"journal\": \"Methods in enzymology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — consolidates multiple substrate identifications with enzymatic and cellular validation across independent studies\",\n      \"pmids\": [\"37230592\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"A de novo DPP9 mutation (p.Arg252Pro) destabilizes the DPP9 protein and abolishes its ability to restrain NLRP1 and CARD8 inflammasomes in HEK293T cells and patient-derived iPSCs, resulting in constitutive inflammasome activation and severe autoinflammation, demonstrating that DPP9 protein stability is required for inflammasome suppression.\",\n      \"method\": \"Heterologous expression in HEK293T, patient-derived iPSCs, inflammasome activation assays, protein stability assessment\",\n      \"journal\": \"The Journal of allergy and clinical immunology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — patient mutation functionally validated in multiple cell systems with direct inflammasome readouts\",\n      \"pmids\": [\"37544411\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"KEAP1 binds DPP9 in an inactive (non-native) conformation and stabilizes this misfolded state. Reciprocally, this inactive form of DPP9 inhibits KEAP1-mediated NRF2 degradation by preventing KEAP1-NRF2 interaction, thereby inducing an antioxidant response. This reveals an endogenous mechanism for DPP9 inhibition linked to the intracellular redox state.\",\n      \"method\": \"Co-IP, biochemical binding assays, NRF2 stability assays, DPP9 enzymatic activity assays under various conditions\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — reciprocal inhibition demonstrated biochemically, but key physiological trigger not yet identified, single lab\",\n      \"pmids\": [\"39615677\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"DPP9 inhibition in liver cancer cells reduces NRF2 stability by releasing KEAP1 from DPP9 binding, thereby increasing KEAP1-mediated NRF2 ubiquitination. DPP9 overexpression up-regulates NQO1 via NRF2 and inhibits intracellular ROS, promoting chemoresistance; NQO1 inhibition by dicoumarol reverses this.\",\n      \"method\": \"siRNA knockdown, overexpression, ubiquitination assay, ROS measurement, cell viability assay with chemotherapy drugs\",\n      \"journal\": \"Redox biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — mechanistic pathway established by KD/OE with multiple readouts, consistent with prior KEAP1-DPP9 finding\",\n      \"pmids\": [\"39094401\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"SFTSV non-structural protein (NSs) activates NLRP1 and CARD8 inflammasomes by disrupting the DPP9-mediated inhibitory ternary complex: NSs interacts with NLRP1/CARD8 FIIND domains (competing with DPP8/9 binding) and promotes degradation of DPP8/DPP9, releasing activated C-terminal fragments.\",\n      \"method\": \"Primary keratinocyte and macrophage infection assays, Co-IP, inflammasome activation assays, CARD8 deletion studies\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — Co-IP with functional inflammasome readouts in primary cells and genetic validation, single lab\",\n      \"pmids\": [\"40608794\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"Proximity labeling (TurboID) of DPP9 in DPP9-knockout cells identified novel DPP9 interactors including DPP8, the E3 ligase CBL, the deubiquitinase complex CYLD-SPATA2, and the BRISC complex (BRCC36/BRCC3 and ABRO1/ABRAXAS2). NanoBRET assays demonstrated that DPP9 disrupts BRCC36-ABRO1 binding and CYLD-SPATA2 binding, revealing non-catalytic scaffolding functions of DPP9 in the ubiquitin system.\",\n      \"method\": \"TurboID proximity labeling, mass spectrometry, Co-IP validation, NanoBRET assays in living cells\",\n      \"journal\": \"Cellular and molecular life sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — proximity labeling with orthogonal validation (Co-IP, NanoBRET), single lab\",\n      \"pmids\": [\"41636814\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"DPP9 disrupts the BRISC-SHMT2 complex, enhancing BRISC-mediated deubiquitination of IFNAR1, which activates JAK/STAT signaling and drives PD-L1 transcription in ccRCC. DPP9 inhibition with 1G244 reverses this by reducing DPP9-SHMT2 interaction, promoting IFNAR1 ubiquitination and degradation, and restoring T cell cytotoxicity.\",\n      \"method\": \"Co-IP, pharmacological inhibition (1G244), IFNAR1 ubiquitination assays, JAK/STAT pathway assays, T cell cytotoxicity assays, in vivo mouse models\",\n      \"journal\": \"Cell death and differentiation\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — mechanistic pathway established with Co-IP, ubiquitination assays, and functional readouts in vitro and in vivo\",\n      \"pmids\": [\"41826729\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"DPP9 is a cytosolic serine dipeptidyl peptidase that removes N-terminal Xaa-Pro dipeptides from substrates (including Syk, AK2, BRCA2) to regulate their stability via the N-degron/proteasomal pathway, acts as a critical checkpoint for NLRP1 and CARD8 inflammasome activation by forming inhibitory ternary complexes in which the NLRP1/CARD8 C-terminal fragment's N-terminus inserts into the DPP9 active site, and additionally exerts non-catalytic scaffolding functions by interacting with partners such as KEAP1 (modulating NRF2-driven antioxidant responses), the BRISC deubiquitinase complex (regulating IFNAR1 stability and PD-L1 expression), and ubiquitin E3/deubiquitinase complexes (CBL, CYLD-SPATA2), with both its enzymatic activity and protein-binding functions being required for full physiological function.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"DPP9 is a cytosolic serine dipeptidyl peptidase that cleaves N-terminal Xaa-Pro dipeptides from substrates including Syk, AK2, and BRCA2, thereby targeting them for proteasomal degradation via the N-degron pathway, and additionally functions as a non-catalytic scaffold that modulates inflammasome activation, redox homeostasis, and ubiquitin signaling [PMID:27614019, PMID:32815200, PMID:37230592]. Cryo-EM structures reveal that DPP9 forms an inhibitory ternary complex with full-length NLRP1 and the NLRP1 C-terminal fragment—whose N-terminus inserts into the DPP9 active site—and both catalytic activity and scaffolding binding are required to suppress NLRP1 and CARD8 inflammasome activation; disruption of this complex by pharmacological inhibitors or viral proteins triggers pyroptosis [PMID:33731932, PMID:33731929, PMID:30291141, PMID:40608794]. Independent of its peptidase activity, DPP9 competes with NRF2 for KEAP1 binding via a conserved ESGE motif, stabilizing NRF2 to drive antioxidant gene expression and suppress ferroptosis, and disrupts the BRISC deubiquitinase complex to enhance IFNAR1-mediated JAK/STAT signaling and PD-L1 expression [PMID:37713596, PMID:39615677, PMID:41826729]. De novo loss-of-function DPP9 mutations cause constitutive NLRP1/CARD8 inflammasome activation and severe autoinflammation in humans [PMID:37544411].\",\n  \"teleology\": [\n    {\n      \"year\": 2002,\n      \"claim\": \"Establishing DPP9 as a new member of the DPP IV serine protease family with cytosolic localization resolved its gene family membership and predicted intracellular rather than extracellular substrates.\",\n      \"evidence\": \"In silico identification, in vitro translation, SDS-PAGE, and northern blot showing ~98 kDa protein lacking transmembrane domains\",\n      \"pmids\": [\"12459266\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No endogenous substrates identified\", \"Catalytic activity not demonstrated against peptide substrates\", \"Cellular localization not confirmed by imaging\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Demonstrating that DPP9 is rate-limiting for cytoplasmic proline-containing peptide degradation and influences MHC-I antigen presentation established its first defined cellular function.\",\n      \"evidence\": \"In vitro cleavage of RU1(34-42) peptide plus siRNA knockdown showing enhanced antigen presentation in intact cells\",\n      \"pmids\": [\"19667070\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Protein substrates not yet identified\", \"In vivo relevance to immune surveillance not tested\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Discovery that SUMO1 binds DPP9 via a non-canonical site on an extended arm near the substrate entry region and stimulates its enzymatic activity revealed the first allosteric regulator of DPP9.\",\n      \"evidence\": \"Co-IP, mutagenesis of SUMO1-binding arm, in vitro activity assays, and SUMO1 siRNA knockdown reducing cytosolic prolyl-peptidase activity\",\n      \"pmids\": [\"23152501\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of SUMO1-DPP9 interaction not resolved at atomic level\", \"Physiological conditions modulating SUMO1-DPP9 interaction unknown\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Linking DPP8/DPP9 inhibition to caspase-1-dependent pyroptosis and gasdermin D cleavage in monocytes identified the inflammasome axis as a major DPP9-regulated pathway, and demonstrating that DPP9 cleaves Syk to generate an N-degron established its role in targeted protein degradation.\",\n      \"evidence\": \"Val-boroPro treatment of monocytes/macrophages and caspase-1 KO mice for inflammasome; Co-IP, pulse-chase, and mutagenesis for Syk N-terminal processing and Cbl-mediated ubiquitination\",\n      \"pmids\": [\"27820798\", \"27614019\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"DPP8 vs DPP9 individual contributions to pyroptosis not resolved\", \"Inflammasome sensor upstream of caspase-1 not yet identified\", \"Full scope of N-degron substrates unknown\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Identifying CARD8 as the sensor mediating DPP8/9 inhibitor-induced pyroptosis in human myeloid cells, and demonstrating DPP9 as a direct binding partner and endogenous inhibitor of both NLRP1 and CARD8 inflammasomes via the FIIND domain, resolved the upstream sensor question and established DPP9 as a dual inflammasome checkpoint.\",\n      \"evidence\": \"CARD8 KO/knockdown with pharmacological inhibition in myeloid cells; proteomics screen, Co-IP, CRISPR KO, and patient NLRP1 FIIND mutation abrogating DPP9 binding\",\n      \"pmids\": [\"29967349\", \"30291141\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of DPP9-FIIND interaction not yet resolved\", \"Relative contributions of DPP9 catalytic vs scaffolding functions not disentangled\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Knock-in mice expressing catalytically inactive DPP9 (S729A) demonstrated that DPP9 enzymatic activity is essential in vivo for survival of migratory tongue muscle progenitors, establishing a non-redundant developmental role.\",\n      \"evidence\": \"S729A knock-in mice showing microglossia, increased apoptosis of occipital somite progenitors, suckling defect, and neonatal lethality\",\n      \"pmids\": [\"28887018\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Substrate(s) responsible for muscle progenitor phenotype not identified\", \"Whether inflammasome dysregulation contributes to neonatal lethality not tested\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Identification of AK2 as a DPP9 substrate showed that DPP9-mediated N-terminal processing diverts proteins from mitochondrial import to proteasomal degradation, broadening DPP9's role beyond signaling kinases to organellar protein quality control.\",\n      \"evidence\": \"siRNA knockdown, proteasome inhibition, pulse-chase, and biochemical fractionation demonstrating DPP9 processing triggers AK2 degradation and prevents cytosolic accumulation\",\n      \"pmids\": [\"32815200\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Full inventory of DPP9 N-degron substrates still incomplete\", \"Whether DPP9 regulates import of other mitochondrial proteins unknown\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Cryo-EM structures of the NLRP1-DPP9 ternary complex revealed the atomic mechanism of inflammasome suppression: the NLRP1 C-terminal fragment N-terminus inserts into the DPP9 active site, and full-length NLRP1 is required to scaffold this interaction, explaining how VbP disrupts the complex to activate inflammasome signaling.\",\n      \"evidence\": \"Cryo-EM structures (human and rat), biochemical reconstitution, mutagenesis, and functional inflammasome assays\",\n      \"pmids\": [\"33731932\", \"33731929\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"High-resolution structure of CARD8-DPP9 complex not determined\", \"Endogenous signals that shift the NLRP1 CT/full-length ratio in vivo remain elusive\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Discovery that DPP9 binds KEAP1 via an ESGE motif to competitively displace NRF2, stabilizing the antioxidant response independently of DPP9 catalytic activity, revealed a major enzyme-independent scaffolding function linking DPP9 to redox homeostasis and chemoresistance.\",\n      \"evidence\": \"Co-IP with ESGE motif mutagenesis, NRF2 stability and transcription assays, ROS measurement, and ferroptosis/sorafenib resistance assays in ccRCC cells\",\n      \"pmids\": [\"37713596\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Physiological trigger switching DPP9 between active and KEAP1-bound inactive conformations not defined\", \"In vivo validation of DPP9-KEAP1 axis in animal models lacking\", \"Whether DPP9-KEAP1 interaction is tissue-specific unknown\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"A de novo DPP9 mutation (p.Arg252Pro) causing severe autoinflammation in a patient demonstrated that DPP9 loss of function is sufficient to constitutively activate NLRP1 and CARD8 inflammasomes in humans, establishing DPP9 deficiency as a Mendelian autoinflammatory condition.\",\n      \"evidence\": \"Functional validation in HEK293T cells and patient-derived iPSCs showing protein destabilization and constitutive inflammasome activation\",\n      \"pmids\": [\"37544411\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single patient reported; additional patients needed to define genotype-phenotype spectrum\", \"Whether partial loss of DPP9 activity confers intermediate inflammatory phenotypes unknown\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Reciprocal biochemical analysis showed that KEAP1 preferentially binds inactive/misfolded DPP9, stabilizing this non-native form, while inactive DPP9 in turn blocks KEAP1-NRF2 interaction—establishing a redox-sensitive feedback loop between DPP9 conformational state and antioxidant signaling.\",\n      \"evidence\": \"Co-IP, DPP9 activity assays under various conditions, NRF2 stability assays; DPP9 KD/OE with ubiquitination and ROS readouts in liver cancer cells\",\n      \"pmids\": [\"39615677\", \"39094401\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Identity of the cellular signal driving DPP9 misfolding/inactivation not established\", \"Structural basis of KEAP1 recognizing inactive DPP9 not resolved\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Demonstration that SFTSV NSs protein activates NLRP1/CARD8 inflammasomes by competing with DPP9 for FIIND binding and promoting DPP9 degradation established viral hijacking of the DPP9 inflammasome checkpoint as a pathogenic mechanism.\",\n      \"evidence\": \"Primary keratinocyte and macrophage infection, Co-IP, inflammasome activation assays, CARD8 deletion studies\",\n      \"pmids\": [\"40608794\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether other viruses exploit the same DPP9-displacement strategy unknown\", \"Structural basis of NSs-FIIND interaction not resolved\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Proximity labeling and NanoBRET identified DPP9 as a non-catalytic disruptor of BRISC (BRCC36-ABRO1) and CYLD-SPATA2 deubiquitinase complexes, and functional studies showed DPP9 enhances BRISC-mediated IFNAR1 deubiquitination to drive JAK/STAT-dependent PD-L1 expression, revealing a scaffolding role in immune checkpoint regulation.\",\n      \"evidence\": \"TurboID proximity labeling, Co-IP, NanoBRET in living cells; IFNAR1 ubiquitination assays, JAK/STAT pathway assays, T cell cytotoxicity assays, and in vivo mouse models with DPP9 inhibitor 1G244\",\n      \"pmids\": [\"41636814\", \"41826729\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct structural interface between DPP9 and BRISC components not mapped\", \"Whether DPP9-BRISC interaction is catalytic-activity-dependent or independent not fully resolved\", \"In vivo immunotherapy efficacy of DPP9 inhibition not tested in clinical setting\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"Major open questions include the complete inventory of physiological DPP9 N-degron substrates, the structural basis of the CARD8-DPP9 complex, the endogenous signals that toggle DPP9 between active and inactive/KEAP1-bound states, and whether DPP9's scaffolding roles in ubiquitin signaling and inflammasome suppression are coordinated or independent pathways.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Full substrate repertoire via unbiased proteomics not yet reported\", \"CARD8-DPP9 complex structure not determined\", \"Physiological trigger for DPP9 conformational switching unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [1, 2, 5, 11, 16]},\n      {\"term_id\": \"GO:0016787\", \"supporting_discovery_ids\": [0, 1, 2, 5, 11, 16]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [7, 10, 12, 13, 15, 18, 22]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [15, 21, 22]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [0, 1, 11]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"GO:0016787\", \"supporting_discovery_ids\": []},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [1, 4, 6, 7, 12, 13, 17, 20, 22]},\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [4, 6, 8]},\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [5, 11, 16]},\n      {\"term_id\": \"R-HSA-8953897\", \"supporting_discovery_ids\": [15, 18, 19]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [9, 15, 22]}\n    ],\n    \"complexes\": [\n      \"NLRP1-DPP9 ternary complex\",\n      \"CARD8-DPP9 complex\"\n    ],\n    \"partners\": [\n      \"NLRP1\",\n      \"CARD8\",\n      \"KEAP1\",\n      \"SUMO1\",\n      \"FLNA\",\n      \"CBL\",\n      \"BRCC3\",\n      \"ABRO1\"\n    ],\n    \"other_free_text\": []\n  }\n}\n```"}