{"gene":"TSPAN15","run_date":"2026-06-10T10:51:56","timeline":{"discoveries":[{"year":2012,"finding":"TSPAN15 directly interacts with ADAM10 (identified by split-ubiquitin yeast two-hybrid screen and co-immunoprecipitation in mammalian cells), accelerates ER exit of the ADAM10-TSPAN15 complex (shown by pulse-chase experiments with an ER-retention mutant), stabilizes the active/mature form of ADAM10 at the cell surface, and increases ADAM10-mediated shedding of N-cadherin and amyloid precursor protein.","method":"Split-ubiquitin yeast two-hybrid, co-immunoprecipitation, ER-retention mutant overexpression, RNAi knockdown, pulse-chase experiments, N-cadherin/APP shedding assays","journal":"Cellular and molecular life sciences : CMLS","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — multiple orthogonal methods (Y2H, Co-IP, pulse-chase, functional shedding assay) in a single focused study; independently replicated by other groups","pmids":["22446748"],"is_preprint":false},{"year":2012,"finding":"TSPAN15 (along with all TspanC8 members: Tspan5, Tspan10, Tspan14, Tspan17, Tspan33) co-immunoprecipitates with ADAM10 and promotes ADAM10 maturation (prodomain processing) and trafficking to the cell surface, establishing TspanC8 tetraspanins as essential regulators of ADAM10 maturation.","method":"Co-immunoprecipitation, cell surface expression assays, maturation assays in multiple cell lines","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP and functional maturation assays, replicated across multiple labs and cell types","pmids":["23035126"],"is_preprint":false},{"year":2012,"finding":"TspanC8 tetraspanins including TSPAN15 directly interact with ADAM10, regulate its exit from the endoplasmic reticulum, and promote Notch activation; two TspanC8 genes were identified as Notch regulators in an independent Drosophila RNAi screen, and human Tspan5 and Tspan14 positively regulated ligand-induced ADAM10-dependent Notch1 signaling (TSPAN15 interaction with ADAM10 confirmed but its specific effect on Notch was not separately quantified in this paper).","method":"Co-immunoprecipitation, RNAi screen in Drosophila, Notch reporter assays, overexpression in mammalian cells","journal":"The Journal of cell biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP plus functional Notch assays plus independent RNAi screen replication in Drosophila","pmids":["23091066"],"is_preprint":false},{"year":2015,"finding":"TSPAN15 (and Tspan5, Tspan14, Tspan33) positively regulates ADAM10 surface expression levels and differentially impacts ADAM10-dependent cleavage of APP, N-cadherin, and CD44, as well as Notch activation, by differentially regulating ADAM10 membrane compartmentalization; sucrose gradient fractionation, single molecule tracking, and quantitative mass-spectrometry showed that Tspan5 and Tspan15 place ADAM10 in distinct membrane compartments with different molecular environments.","method":"Sucrose gradient fractionation, single molecule tracking, quantitative mass-spectrometry co-immunoprecipitation, Notch reporter assays, substrate shedding assays","journal":"Cellular and molecular life sciences : CMLS","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — three orthogonal methods (fractionation, single-molecule tracking, quantitative MS) plus functional shedding assays in a single study","pmids":["26686862"],"is_preprint":false},{"year":2015,"finding":"The large extracellular loop (LEL) of Tspan14 mediates co-immunoprecipitation with ADAM10 and promotes ADAM10 maturation and trafficking; chimeric ADAM10 constructs showed that the membrane-proximal stalk, cysteine-rich, and disintegrin domains of ADAM10 mediate its interaction with TspanC8s including Tspan15, and this region is required for ADAM10 ER exit. Tspan15 was the only TspanC8 to promote cleavage of N-cadherin, whereas Tspan14 uniquely reduced cleavage of GPVI, indicating distinct substrate selectivity depending on the associated TspanC8.","method":"Chimeric protein constructs, co-immunoprecipitation, substrate cleavage assays, antibody development to endogenous Tspan14","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — chimeric domain-mapping constructs combined with functional cleavage assays and antibody-validated endogenous interaction","pmids":["26668317"],"is_preprint":false},{"year":2018,"finding":"In Tspan15 knockout mice, the active/mature form of ADAM10 is substantially decreased in brain tissue, and this is accompanied by age-dependent reduced shedding of N-cadherin and cellular prion protein, but not APP alpha-secretase cleavage or Notch-dependent gene expression, demonstrating that Tspan15 in vivo preferentially directs ADAM10 toward specific substrates (N-cadherin, PrP) rather than others (APP, Notch targets).","method":"Tspan15 knockout mouse model, western blotting for ADAM10 maturation, substrate shedding assays (N-cadherin, PrP, APP), Notch target gene expression analysis","journal":"Cellular and molecular life sciences : CMLS","confidence":"High","confidence_rationale":"Tier 2 / Strong — clean genetic knockout with multiple substrate-specific readouts in vivo, consistent with in vitro data from other labs","pmids":["29520422"],"is_preprint":false},{"year":2019,"finding":"ADAM10 undergoes faster endocytosis in the presence of Tspan5 than Tspan15; Tspan15 stabilizes ADAM10 at the cell surface yielding high surface expression levels, and reciprocally ADAM10 stabilizes Tspan15 at the cell surface (the Tspan15/ADAM10 complex is retained at the plasma membrane). The cytoplasmic domains of Tspan5 and Tspan15 contribute to their opposite effects on ADAM10 trafficking and Notch signaling, while an unusual C-terminal palmitoylation site of Tspan15 is dispensable for these functions.","method":"Endocytosis assays, cell surface biotinylation, flow cytometry, chimeric cytoplasmic domain constructs, palmitoylation site mutagenesis","journal":"Life science alliance","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — functional domain mapping with chimeric constructs, endocytosis kinetics, and reciprocal stabilization demonstrated by multiple methods","pmids":["31792032"],"is_preprint":false},{"year":2020,"finding":"Endogenous Tspan15 and ADAM10 co-localize on the cell surface; ADAM10 is the principal Tspan15-interacting protein; endogenous Tspan15 expression requires ADAM10 in cell lines and primary cells (Tspan15 is unstable without ADAM10); a synthetic ADAM10/Tspan15 fusion protein is a functional scissor complex; two of four anti-Tspan15 monoclonal antibodies impaired ADAM10/Tspan15 activity.","method":"Monoclonal antibody generation, co-localization by immunofluorescence, co-immunoprecipitation of endogenous proteins, ADAM10-knockout cell lines, synthetic fusion protein assay, substrate cleavage inhibition assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — endogenous protein co-localization and co-IP, ADAM10-KO dependency, functional fusion protein, antibody inhibition — multiple orthogonal methods","pmids":["32111735"],"is_preprint":false},{"year":2021,"finding":"Crystal structure of the Tspan15 large extracellular loop (LEL) was solved, revealing a core helical framework with a variable region; a site on the Tspan15 LEL required for both ADAM10 binding and promoting N-cadherin substrate cleavage was identified by co-immunoprecipitation and cellular cleavage assay.","method":"X-ray crystallography, co-immunoprecipitation, N-cadherin cellular cleavage assay, mutagenesis of LEL binding site","journal":"Structure (London, England : 1993)","confidence":"High","confidence_rationale":"Tier 1 / Moderate — crystal structure plus mutagenesis plus functional cleavage assay in a single study; single lab","pmids":["34739841"],"is_preprint":false},{"year":2023,"finding":"Cryo-EM structure of a vFab-ADAM10-Tspan15 complex shows that Tspan15 binding relieves ADAM10 autoinhibition and acts as a molecular measuring stick, positioning the ADAM10 enzyme active site approximately 20 Å from the plasma membrane for membrane-proximal substrate cleavage. Cell-based N-cadherin shedding assays confirmed that the positioning of the active site by the ADAM10-Tspan15 interface influences preferred cleavage site selection.","method":"Cryo-EM structure determination, cell-based N-cadherin shedding assays, interface mutagenesis","journal":"Cell","confidence":"High","confidence_rationale":"Tier 1 / Strong — high-resolution cryo-EM structure combined with functional cell-based mutagenesis validating the structural mechanism","pmids":["37516108"],"is_preprint":false},{"year":2018,"finding":"TSPAN15 specifically interacts with BTRC (beta-TrCP) E3 ubiquitin ligase to promote ubiquitination and proteasomal degradation of phospho-IκBα, thereby triggering NF-κB nuclear translocation and activation of metastasis-related genes (ICAM1, VCAM1, uPA, MMP9, TNFα, CCL2) in oesophageal squamous cell carcinoma cells.","method":"Co-immunoprecipitation, ubiquitination assay, NF-κB reporter assay, western blotting for IκBα degradation and p65 nuclear translocation, target gene expression analysis","journal":"Nature communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP plus functional ubiquitination and NF-κB assays; single lab with multiple methods but no independent replication","pmids":["29650964"],"is_preprint":false},{"year":2022,"finding":"CRISPR/Cas9 knockout of Tspan15 (and Tspan33) in human cell lines demonstrated that Tspan15 and Tspan33 play compensatory roles in GPVI cleavage by ADAM10, with Tspan15 bearing the more important role. The Tspan15 extracellular region was found critical for promoting GPVI cleavage, enabling ADAM10 access to the cleavage site at a particular distance above the membrane.","method":"CRISPR/Cas9 knockout cell lines, Tspan15 and GPVI mutant expression constructs, GPVI cleavage assays","journal":"International journal of molecular sciences","confidence":"High","confidence_rationale":"Tier 2 / Moderate — CRISPR KO plus domain-mapping constructs with functional cleavage assays; single lab but multiple orthogonal approaches","pmids":["35269584"],"is_preprint":false},{"year":2023,"finding":"Tspan15 is a component of TSG101- and CD81-positive extracellular vesicle (EV) fractions in neurons. Tspan15 is dispensable at target neuron plasma membranes but is required at the EV surface to promote EV docking at target neurons, as EVs from Tspan15 knockout cortical neurons showed significantly impaired association with target cells compared to wild-type EVs.","method":"EV fractionation and marker analysis, fluorescent fusion protein tracking, Tspan15 knockout mouse cortical neuron EVs, target cell docking assays","journal":"Journal of extracellular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic KO with functional EV docking readout and EV fractionation; single lab, novel function outside established ADAM10 axis","pmids":["38938373"],"is_preprint":false},{"year":2019,"finding":"In oral squamous cell carcinoma cells, knockdown of Tspan15 reduced ADAM10 expression, decreased soluble N-cadherin shedding, reduced nuclear β-catenin immunoreactivity, and suppressed tumor invasion and migration, placing Tspan15 upstream of ADAM10-mediated N-cadherin shedding and downstream Wnt/β-catenin signaling in OSCC metastasis.","method":"siRNA knockdown, N-cadherin shedding ELISA, immunofluorescence for β-catenin localization, Transwell invasion/migration assays","journal":"Experimental cell research","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — siRNA KD with multiple mechanistic readouts (shedding, localization, invasion) but single lab and no rescue experiment","pmids":["31518558"],"is_preprint":false},{"year":2019,"finding":"Tspan15 overexpression in the hepatoma cell line HepG2 increases ERK1/2 phosphorylation, leading to increased CTGF expression and secretion; proteomic profiling of Tspan15 complexes identified multiple membrane proteins including growth factor receptors as interaction partners.","method":"Overexpression, western blotting for p-ERK1/2, quantitative secretome proteomics (MS), Tspan15 complex immunoprecipitation/MS","journal":"Proteomics","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — overexpression with western blot and MS-based proteomics; single lab, no loss-of-function rescue","pmids":["31390680"],"is_preprint":false},{"year":2025,"finding":"TSPAN15 directly interacts with integrin-β1 (ITGB1) and maintains ITGB1 stability by inhibiting its ubiquitination; this interaction activates downstream p-FAK/p-AKT/p-mTOR signaling and promotes GPX4 expression, thereby attenuating gemcitabine-induced ferroptosis in pancreatic ductal adenocarcinoma cells.","method":"Co-immunoprecipitation, ubiquitination assay, western blotting for FAK/AKT/mTOR phosphorylation and GPX4, TSPAN15 knockdown in vitro and in vivo xenograft, ferroptosis assays","journal":"Redox biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP plus functional pathway assays and in vivo xenograft; single lab, novel ITGB1 interaction outside established ADAM10 biology","pmids":["40505345"],"is_preprint":false},{"year":2025,"finding":"TSPAN15 physically interacts with BTRC to promote proteasomal degradation of tumor suppressor PDCD4 via ubiquitination, thereby activating autophagy and autophagy-mediated EMT and metastasis in hepatocellular carcinoma cells.","method":"Co-immunoprecipitation, ubiquitination assay, autophagic flux analysis, TSPAN15 silencing, PDCD4 rescue experiments, xenograft mouse model","journal":"Molecular immunology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP plus functional ubiquitination assay, autophagic flux, and in vivo model; single lab; mechanistically consistent with prior BTRC interaction data","pmids":["40398082"],"is_preprint":false},{"year":2023,"finding":"In intrahepatic cholangiocarcinoma (ICC) cells, Tspan15 mediates translocation of activated mature ADAM10 from the cytoplasm to the cell membrane surface, which cleaves the Notch1 intracellular domain from its extracellular domain, activating Notch1 signaling and enhancing cancer stem cell-like properties, EMT, and chemoresistance against gemcitabine and cisplatin.","method":"Western blotting, flow cytometry, immunohistochemistry, RT-PCR, Tspan15/ADAM10 knockdown, Notch1 activation assays, chemoresistance functional assays","journal":"Liver international : official journal of the International Association for the Study of the Liver","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — knockdown with multiple functional readouts (Notch activation, CSC markers, chemoresistance); mechanistically consistent with established Tspan15-ADAM10-Notch axis; single lab","pmids":["37545390"],"is_preprint":false},{"year":2024,"finding":"In invasive bladder cancer cells, Tspan15 is required for ADAM10-mediated selective cleavage of N-cadherin; the PPARβ/δ agonist GW501516 decreases Tspan15 expression and prevents N-cadherin cleavage (NTF generation) without modifying ADAM10 expression levels, demonstrating that pharmacological targeting of Tspan15 can selectively block ADAM10 substrate cleavage.","method":"siRNA knockdown of Tspan15, western blotting for N-cadherin fragments (NTF/CTF1), GW501516 treatment, ADAM10 expression analysis","journal":"Cells","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — siRNA KD plus pharmacological intervention with specific mechanistic readout; single lab; consistent with established Tspan15-ADAM10-N-cadherin mechanism","pmids":["38667323"],"is_preprint":false},{"year":2025,"finding":"A tricomponent complex of THSD7A/ADAM10/Tspan15 was found in podocytes; Tspan15 regulates ADAM10 substrate usage and the stability of podocyte cell surface proteins (THSD7A, PLA2R1, β-dystroglycan), where THSD7A acts as both an ADAM10 substrate and regulator of the complex; Tspan15 is present at podocyte foot processes.","method":"Co-localization and Co-IP, ADAM10-deficient mice, ADAM10-inhibited pig glomeruli, in vitro shedding assays, biochemical fractionation","journal":"Kidney international","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP plus genetic/pharmacological ADAM10 inhibition across multiple experimental systems; tricomponent complex is a novel finding from single lab","pmids":["40339751"],"is_preprint":false},{"year":2025,"finding":"Silencing of Tspan15 (and Tspan10) in astrocytoma cells reduced Venezuelan equine encephalitis virus (VEEV) genome replication without affecting viral entry; silencing of the ADAM10 substrate N-cadherin also reduced VEEV infectivity, suggesting Tspan15/ADAM10 and downstream substrates modulate VEEV replication.","method":"siRNA silencing, pharmacological ADAM10 inhibition, VEEV infection assays, viral entry vs. replication discrimination assays","journal":"Molecular biology of the cell","confidence":"Low","confidence_rationale":"Tier 3 / Weak — siRNA KD with infectivity readout; single lab, single method per conclusion; indirect mechanism","pmids":["39878649"],"is_preprint":false},{"year":2000,"finding":"TSPAN15 (identified as NET-7) was identified and sequenced as a new tetraspanin family member with the characteristic structure of four transmembrane domains, two extracellular regions, and conserved tetraspanin amino acid residues; it shows differential expression across human cell lines.","method":"EST sequencing, sequence analysis, RT-PCR expression profiling","journal":"Biochimica et biophysica acta","confidence":"Low","confidence_rationale":"Tier 3 / Weak — sequence/expression characterization only; no functional mechanism established","pmids":["10719184"],"is_preprint":false}],"current_model":"TSPAN15 is a TspanC8-subfamily tetraspanin that forms a direct, stoichiometric complex with the transmembrane metalloprotease ADAM10; structural and biochemical studies show that Tspan15 binds ADAM10 via its large extracellular loop (LEL) interacting with the stalk/cysteine-rich/disintegrin domains of ADAM10, relieves ADAM10 autoinhibition, and positions the catalytic active site ~20 Å above the plasma membrane as a 'molecular measuring stick' for membrane-proximal substrate cleavage. TSPAN15 promotes ADAM10 exit from the endoplasmic reticulum, stabilizes the mature active form of ADAM10 at the cell surface by slowing its endocytosis, and confers substrate selectivity on the complex—particularly for N-cadherin, PrP, and GPVI cleavage—distinct from other TspanC8/ADAM10 pairs. In cancer contexts, TSPAN15 also interacts with BTRC to promote IκBα degradation and NF-κB activation, and interacts with integrin-β1 to activate FAK/AKT/mTOR/GPX4 signaling and suppress ferroptosis; additionally, Tspan15 is required at the surface of neuronal extracellular vesicles to promote their docking at target neurons."},"narrative":{"mechanistic_narrative":"TSPAN15 is a TspanC8-subfamily tetraspanin that functions as a dedicated regulatory partner of the transmembrane metalloprotease ADAM10, controlling its maturation, surface stability, and substrate selectivity [PMID:22446748, PMID:23035126]. It directly binds ADAM10 and accelerates exit of the complex from the endoplasmic reticulum, stabilizing the active/mature enzyme at the plasma membrane by slowing its endocytosis; ADAM10 reciprocally stabilizes TSPAN15, and endogenous TSPAN15 is unstable in the absence of ADAM10 [PMID:22446748, PMID:31792032, PMID:32111735]. The interaction is mediated by the TSPAN15 large extracellular loop (LEL) contacting the membrane-proximal stalk, cysteine-rich, and disintegrin domains of ADAM10; crystallographic and cryo-EM structures show that TSPAN15 binding relieves ADAM10 autoinhibition and acts as a 'molecular measuring stick', positioning the catalytic active site approximately 20 Å above the membrane to dictate membrane-proximal cleavage-site selection [PMID:26668317, PMID:34739841, PMID:37516108]. Through this mechanism TSPAN15 confers substrate specificity distinct from other TspanC8/ADAM10 pairs, preferentially directing ADAM10 toward N-cadherin, cellular prion protein, and GPVI rather than APP or Notch targets, as confirmed in knockout mice [PMID:29520422, PMID:35269584]. Beyond the canonical protease axis, TSPAN15 has been implicated in cancer signaling—interacting with the BTRC E3 ligase to promote IκBα and PDCD4 degradation [PMID:29650964, PMID:40398082] and with integrin-β1 to sustain FAK/AKT/mTOR signaling [PMID:40505345]—and is required at the surface of neuronal extracellular vesicles for their docking at target neurons [PMID:38938373].","teleology":[{"year":2000,"claim":"Before any functional role was known, TSPAN15 had to be defined as a distinct gene; its identification as the tetraspanin NET-7 established the four-transmembrane architecture and conserved residues that place it in the tetraspanin family.","evidence":"EST sequencing, sequence analysis, and RT-PCR expression profiling across human cell lines","pmids":["10719184"],"confidence":"Low","gaps":["No functional mechanism established","Differential expression not linked to any pathway"]},{"year":2012,"claim":"The central question of what TSPAN15 does was answered by showing it directly binds ADAM10, accelerates ER exit of the complex, stabilizes mature surface ADAM10, and increases shedding of N-cadherin and APP—establishing TSPAN15 as an ADAM10 trafficking and maturation regulator.","evidence":"Split-ubiquitin Y2H, co-IP, ER-retention mutant pulse-chase, RNAi, and substrate shedding assays in mammalian cells; reciprocal Co-IP and maturation assays establishing the broader TspanC8 family role and Notch regulation","pmids":["22446748","23035126","23091066"],"confidence":"High","gaps":["Did not resolve whether substrate preference differs among TspanC8 members","Structural basis of the interaction not defined"]},{"year":2015,"claim":"To explain why different TspanC8s yield different ADAM10 outputs, studies mapped the interaction interface and demonstrated compartment-specific and substrate-specific effects, showing TSPAN15 uniquely promotes N-cadherin cleavage and places ADAM10 in a distinct membrane environment.","evidence":"Chimeric ADAM10/TspanC8 constructs, sucrose gradient fractionation, single-molecule tracking, quantitative MS co-IP, and substrate cleavage/Notch reporter assays","pmids":["26686862","26668317"],"confidence":"High","gaps":["Mechanism of how compartmentalization selects substrates not structurally defined","In vivo relevance not yet tested"]},{"year":2018,"claim":"Genetic knockout in mice resolved whether TSPAN15 substrate selectivity operates in vivo, showing loss of mature ADAM10 in brain and age-dependent loss of N-cadherin and PrP shedding but not APP or Notch readouts—proving physiological substrate steering.","evidence":"Tspan15 knockout mouse brain analyzed by western blotting for ADAM10 maturation and substrate-specific shedding and Notch target gene expression","pmids":["29520422"],"confidence":"High","gaps":["Molecular basis for in vivo substrate discrimination not addressed","Phenotypic consequences for brain physiology not characterized"]},{"year":2019,"claim":"The mechanism of surface stabilization was clarified by showing TSPAN15 slows ADAM10 endocytosis (in contrast to Tspan5), with the cytoplasmic domain—not the C-terminal palmitoylation site—governing trafficking and Notch outcomes.","evidence":"Endocytosis assays, cell surface biotinylation, flow cytometry, chimeric cytoplasmic-domain constructs, and palmitoylation-site mutagenesis","pmids":["31792032"],"confidence":"High","gaps":["Endocytic adaptor recognizing the cytoplasmic domain not identified","Quantitative contribution of trafficking versus stability to net activity unresolved"]},{"year":2020,"claim":"To confirm physiological relevance and therapeutic tractability, work on endogenous proteins showed ADAM10 is the principal TSPAN15 partner, TSPAN15 is unstable without ADAM10, a synthetic fusion is a functional scissor, and antibodies can inhibit the complex.","evidence":"Monoclonal antibody generation, endogenous co-localization and co-IP, ADAM10-KO cell lines, synthetic fusion protein, and antibody inhibition of substrate cleavage","pmids":["32111735"],"confidence":"High","gaps":["Epitopes of inhibitory antibodies not mapped","Therapeutic specificity against other TspanC8/ADAM10 complexes not established"]},{"year":2023,"claim":"The structural logic of substrate selection was resolved when crystal and cryo-EM structures showed the TSPAN15 LEL binding site relieves ADAM10 autoinhibition and acts as a molecular measuring stick positioning the active site ~20 Å above the membrane to dictate cleavage-site choice.","evidence":"X-ray crystallography of the TSPAN15 LEL with mutagenesis and N-cadherin cleavage assays; cryo-EM of a vFab-ADAM10-Tspan15 complex with interface mutagenesis and cell-based shedding assays","pmids":["34739841","37516108"],"confidence":"High","gaps":["Structures of complexes with other TspanC8s for comparison not available","Dynamics of substrate engagement not captured"]},{"year":2022,"claim":"Substrate-selectivity rules were extended by showing TSPAN15 and Tspan33 act compensatorily in ADAM10-mediated GPVI cleavage, with TSPAN15 dominant and its extracellular region setting the cleavage distance.","evidence":"CRISPR/Cas9 knockout cell lines with TSPAN15 and GPVI mutant constructs and GPVI cleavage assays","pmids":["35269584"],"confidence":"High","gaps":["Mechanism of TSPAN15/Tspan33 compensation not defined","Platelet-physiological consequences not tested here"]},{"year":2018,"claim":"A non-protease cancer role was opened by showing TSPAN15 binds the BTRC E3 ligase to drive phospho-IκBα ubiquitination and NF-κB activation of metastasis genes, later paralleled by BTRC-dependent PDCD4 degradation driving autophagy and EMT.","evidence":"Co-IP, ubiquitination assays, NF-κB reporter, and target gene analysis in oesophageal carcinoma; co-IP, ubiquitination, autophagic flux, PDCD4 rescue, and xenograft in hepatocellular carcinoma","pmids":["29650964","40398082"],"confidence":"Medium","gaps":["Direct versus ADAM10-dependent nature of these effects not separated","How a tetraspanin engages a cytosolic E3 ligase mechanistically unclear","Independent replication of BTRC axis limited"]},{"year":2023,"claim":"TSPAN15 was assigned a function distinct from ADAM10 in neuronal extracellular vesicle biology, where it is required at the EV surface for docking at target neurons rather than at the recipient membrane.","evidence":"EV fractionation and marker analysis, fluorescent fusion tracking, and Tspan15-knockout cortical neuron EV target-cell docking assays","pmids":["38938373"],"confidence":"Medium","gaps":["Docking receptor/ligand on target neurons not identified","Whether ADAM10 is involved not resolved","Single lab"]},{"year":2025,"claim":"An ADAM10-independent oncogenic axis was proposed in which TSPAN15 binds integrin-β1, blocks its ubiquitination, and sustains FAK/AKT/mTOR/GPX4 signaling to suppress ferroptosis and confer chemoresistance.","evidence":"Co-IP, ubiquitination assay, phospho-signaling and GPX4 westerns, knockdown in vitro and in vivo xenograft, and ferroptosis assays in pancreatic cancer cells","pmids":["40505345"],"confidence":"Medium","gaps":["Direct binding interface with ITGB1 not mapped","Independence from ADAM10 not formally tested","Single lab, novel interaction"]},{"year":2025,"claim":"Disease-tissue context was extended by identifying a THSD7A/ADAM10/TSPAN15 tricomponent complex at podocyte foot processes that regulates ADAM10 substrate usage and surface protein stability.","evidence":"Co-localization and Co-IP, ADAM10-deficient mice, ADAM10-inhibited pig glomeruli, in vitro shedding assays, and biochemical fractionation","pmids":["40339751"],"confidence":"Medium","gaps":["Whether THSD7A binds TSPAN15 directly not resolved","Pathophysiological link to membranous nephropathy not established"]},{"year":null,"claim":"It remains unresolved how the canonical ADAM10-scaffolding role of TSPAN15 mechanistically relates to its reported ADAM10-independent activities (BTRC/E3-ligase coupling, integrin-β1 stabilization, EV docking), and whether these reflect distinct molecular interfaces of the same protein.","evidence":"No single study reconciles the protease-scaffold and cytosolic-signaling functions","pmids":[],"confidence":"Low","gaps":["No structural or interface data for the cytosolic/cancer interactions","No demonstration that protease-independent roles occur without ADAM10 present","In vivo physiological weighting of the multiple roles unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[0,5,9]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[0,7,9]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[6,7,9]},{"term_id":"GO:0005783","term_label":"endoplasmic reticulum","supporting_discovery_ids":[0,4]}],"pathway":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0,5,9]}],"complexes":["ADAM10-TSPAN15 sheddase complex","THSD7A/ADAM10/TSPAN15 complex"],"partners":["ADAM10","BTRC","ITGB1","THSD7A"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"O95858","full_name":"Tetraspanin-15","aliases":["Tetraspan NET-7","Transmembrane 4 superfamily member 15"],"length_aa":294,"mass_kda":33.2,"function":"Part of TspanC8 subgroup, composed of 6 members that interact with the transmembrane metalloprotease ADAM10. This interaction is required for ADAM10 exit from the endoplasmic reticulum and for enzymatic maturation and trafficking to the cell surface as well as substrate specificity. Different TspanC8/ADAM10 complexes have distinct substrates (PubMed:26686862, PubMed:30463011, PubMed:31792032, PubMed:34739841). Promotes ADAM10-mediated cleavage of CDH2 (PubMed:34739841). Negatively regulates ligand-induced Notch activity probably by regulating ADAM10 activity (PubMed:26686862, PubMed:31792032)","subcellular_location":"Cell membrane; Late endosome membrane","url":"https://www.uniprot.org/uniprotkb/O95858/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/TSPAN15","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/TSPAN15","total_profiled":1310},"omim":[{"mim_id":"613140","title":"TETRASPANIN 15; TSPAN15","url":"https://www.omim.org/entry/613140"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Nuclear bodies","reliability":"Approved"},{"location":"Plasma membrane","reliability":"Approved"},{"location":"Vesicles","reliability":"Additional"},{"location":"Cytosol","reliability":"Additional"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in many","driving_tissues":[{"tissue":"brain","ntpm":98.0}],"url":"https://www.proteinatlas.org/search/TSPAN15"},"hgnc":{"alias_symbol":["NET-7"],"prev_symbol":["TM4SF15"]},"alphafold":{"accession":"O95858","domains":[{"cath_id":"-","chopping":"2-136_235-267","consensus_level":"high","plddt":90.4445,"start":2,"end":267}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/O95858","model_url":"https://alphafold.ebi.ac.uk/files/AF-O95858-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-O95858-F1-predicted_aligned_error_v6.png","plddt_mean":87.56},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=TSPAN15","jax_strain_url":"https://www.jax.org/strain/search?query=TSPAN15"},"sequence":{"accession":"O95858","fasta_url":"https://rest.uniprot.org/uniprotkb/O95858.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/O95858/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/O95858"}},"corpus_meta":[{"pmid":"23091066","id":"PMC_23091066","title":"TspanC8 tetraspanins regulate ADAM10/Kuzbanian trafficking and promote Notch activation in flies and mammals.","date":"2012","source":"The Journal of cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/23091066","citation_count":157,"is_preprint":false},{"pmid":"23035126","id":"PMC_23035126","title":"The TspanC8 subgroup of tetraspanins interacts with A disintegrin and metalloprotease 10 (ADAM10) and regulates its maturation and cell surface expression.","date":"2012","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/23035126","citation_count":136,"is_preprint":false},{"pmid":"26686862","id":"PMC_26686862","title":"TspanC8 tetraspanins differentially regulate the cleavage of ADAM10 substrates, Notch activation and ADAM10 membrane compartmentalization.","date":"2015","source":"Cellular and molecular life sciences : 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independently replicated by other groups\",\n      \"pmids\": [\"22446748\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"TSPAN15 (along with all TspanC8 members: Tspan5, Tspan10, Tspan14, Tspan17, Tspan33) co-immunoprecipitates with ADAM10 and promotes ADAM10 maturation (prodomain processing) and trafficking to the cell surface, establishing TspanC8 tetraspanins as essential regulators of ADAM10 maturation.\",\n      \"method\": \"Co-immunoprecipitation, cell surface expression assays, maturation assays in multiple cell lines\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP and functional maturation assays, replicated across multiple labs and cell types\",\n      \"pmids\": [\"23035126\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"TspanC8 tetraspanins including TSPAN15 directly interact with ADAM10, regulate its exit from the endoplasmic reticulum, and promote Notch activation; two TspanC8 genes were identified as Notch regulators in an independent Drosophila RNAi screen, and human Tspan5 and Tspan14 positively regulated ligand-induced ADAM10-dependent Notch1 signaling (TSPAN15 interaction with ADAM10 confirmed but its specific effect on Notch was not separately quantified in this paper).\",\n      \"method\": \"Co-immunoprecipitation, RNAi screen in Drosophila, Notch reporter assays, overexpression in mammalian cells\",\n      \"journal\": \"The Journal of cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP plus functional Notch assays plus independent RNAi screen replication in Drosophila\",\n      \"pmids\": [\"23091066\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"TSPAN15 (and Tspan5, Tspan14, Tspan33) positively regulates ADAM10 surface expression levels and differentially impacts ADAM10-dependent cleavage of APP, N-cadherin, and CD44, as well as Notch activation, by differentially regulating ADAM10 membrane compartmentalization; sucrose gradient fractionation, single molecule tracking, and quantitative mass-spectrometry showed that Tspan5 and Tspan15 place ADAM10 in distinct membrane compartments with different molecular environments.\",\n      \"method\": \"Sucrose gradient fractionation, single molecule tracking, quantitative mass-spectrometry co-immunoprecipitation, Notch reporter assays, substrate shedding assays\",\n      \"journal\": \"Cellular and molecular life sciences : CMLS\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — three orthogonal methods (fractionation, single-molecule tracking, quantitative MS) plus functional shedding assays in a single study\",\n      \"pmids\": [\"26686862\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"The large extracellular loop (LEL) of Tspan14 mediates co-immunoprecipitation with ADAM10 and promotes ADAM10 maturation and trafficking; chimeric ADAM10 constructs showed that the membrane-proximal stalk, cysteine-rich, and disintegrin domains of ADAM10 mediate its interaction with TspanC8s including Tspan15, and this region is required for ADAM10 ER exit. Tspan15 was the only TspanC8 to promote cleavage of N-cadherin, whereas Tspan14 uniquely reduced cleavage of GPVI, indicating distinct substrate selectivity depending on the associated TspanC8.\",\n      \"method\": \"Chimeric protein constructs, co-immunoprecipitation, substrate cleavage assays, antibody development to endogenous Tspan14\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — chimeric domain-mapping constructs combined with functional cleavage assays and antibody-validated endogenous interaction\",\n      \"pmids\": [\"26668317\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"In Tspan15 knockout mice, the active/mature form of ADAM10 is substantially decreased in brain tissue, and this is accompanied by age-dependent reduced shedding of N-cadherin and cellular prion protein, but not APP alpha-secretase cleavage or Notch-dependent gene expression, demonstrating that Tspan15 in vivo preferentially directs ADAM10 toward specific substrates (N-cadherin, PrP) rather than others (APP, Notch targets).\",\n      \"method\": \"Tspan15 knockout mouse model, western blotting for ADAM10 maturation, substrate shedding assays (N-cadherin, PrP, APP), Notch target gene expression analysis\",\n      \"journal\": \"Cellular and molecular life sciences : CMLS\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — clean genetic knockout with multiple substrate-specific readouts in vivo, consistent with in vitro data from other labs\",\n      \"pmids\": [\"29520422\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"ADAM10 undergoes faster endocytosis in the presence of Tspan5 than Tspan15; Tspan15 stabilizes ADAM10 at the cell surface yielding high surface expression levels, and reciprocally ADAM10 stabilizes Tspan15 at the cell surface (the Tspan15/ADAM10 complex is retained at the plasma membrane). The cytoplasmic domains of Tspan5 and Tspan15 contribute to their opposite effects on ADAM10 trafficking and Notch signaling, while an unusual C-terminal palmitoylation site of Tspan15 is dispensable for these functions.\",\n      \"method\": \"Endocytosis assays, cell surface biotinylation, flow cytometry, chimeric cytoplasmic domain constructs, palmitoylation site mutagenesis\",\n      \"journal\": \"Life science alliance\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — functional domain mapping with chimeric constructs, endocytosis kinetics, and reciprocal stabilization demonstrated by multiple methods\",\n      \"pmids\": [\"31792032\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Endogenous Tspan15 and ADAM10 co-localize on the cell surface; ADAM10 is the principal Tspan15-interacting protein; endogenous Tspan15 expression requires ADAM10 in cell lines and primary cells (Tspan15 is unstable without ADAM10); a synthetic ADAM10/Tspan15 fusion protein is a functional scissor complex; two of four anti-Tspan15 monoclonal antibodies impaired ADAM10/Tspan15 activity.\",\n      \"method\": \"Monoclonal antibody generation, co-localization by immunofluorescence, co-immunoprecipitation of endogenous proteins, ADAM10-knockout cell lines, synthetic fusion protein assay, substrate cleavage inhibition assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — endogenous protein co-localization and co-IP, ADAM10-KO dependency, functional fusion protein, antibody inhibition — multiple orthogonal methods\",\n      \"pmids\": [\"32111735\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Crystal structure of the Tspan15 large extracellular loop (LEL) was solved, revealing a core helical framework with a variable region; a site on the Tspan15 LEL required for both ADAM10 binding and promoting N-cadherin substrate cleavage was identified by co-immunoprecipitation and cellular cleavage assay.\",\n      \"method\": \"X-ray crystallography, co-immunoprecipitation, N-cadherin cellular cleavage assay, mutagenesis of LEL binding site\",\n      \"journal\": \"Structure (London, England : 1993)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — crystal structure plus mutagenesis plus functional cleavage assay in a single study; single lab\",\n      \"pmids\": [\"34739841\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Cryo-EM structure of a vFab-ADAM10-Tspan15 complex shows that Tspan15 binding relieves ADAM10 autoinhibition and acts as a molecular measuring stick, positioning the ADAM10 enzyme active site approximately 20 Å from the plasma membrane for membrane-proximal substrate cleavage. Cell-based N-cadherin shedding assays confirmed that the positioning of the active site by the ADAM10-Tspan15 interface influences preferred cleavage site selection.\",\n      \"method\": \"Cryo-EM structure determination, cell-based N-cadherin shedding assays, interface mutagenesis\",\n      \"journal\": \"Cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — high-resolution cryo-EM structure combined with functional cell-based mutagenesis validating the structural mechanism\",\n      \"pmids\": [\"37516108\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"TSPAN15 specifically interacts with BTRC (beta-TrCP) E3 ubiquitin ligase to promote ubiquitination and proteasomal degradation of phospho-IκBα, thereby triggering NF-κB nuclear translocation and activation of metastasis-related genes (ICAM1, VCAM1, uPA, MMP9, TNFα, CCL2) in oesophageal squamous cell carcinoma cells.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, NF-κB reporter assay, western blotting for IκBα degradation and p65 nuclear translocation, target gene expression analysis\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP plus functional ubiquitination and NF-κB assays; single lab with multiple methods but no independent replication\",\n      \"pmids\": [\"29650964\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"CRISPR/Cas9 knockout of Tspan15 (and Tspan33) in human cell lines demonstrated that Tspan15 and Tspan33 play compensatory roles in GPVI cleavage by ADAM10, with Tspan15 bearing the more important role. The Tspan15 extracellular region was found critical for promoting GPVI cleavage, enabling ADAM10 access to the cleavage site at a particular distance above the membrane.\",\n      \"method\": \"CRISPR/Cas9 knockout cell lines, Tspan15 and GPVI mutant expression constructs, GPVI cleavage assays\",\n      \"journal\": \"International journal of molecular sciences\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — CRISPR KO plus domain-mapping constructs with functional cleavage assays; single lab but multiple orthogonal approaches\",\n      \"pmids\": [\"35269584\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Tspan15 is a component of TSG101- and CD81-positive extracellular vesicle (EV) fractions in neurons. Tspan15 is dispensable at target neuron plasma membranes but is required at the EV surface to promote EV docking at target neurons, as EVs from Tspan15 knockout cortical neurons showed significantly impaired association with target cells compared to wild-type EVs.\",\n      \"method\": \"EV fractionation and marker analysis, fluorescent fusion protein tracking, Tspan15 knockout mouse cortical neuron EVs, target cell docking assays\",\n      \"journal\": \"Journal of extracellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic KO with functional EV docking readout and EV fractionation; single lab, novel function outside established ADAM10 axis\",\n      \"pmids\": [\"38938373\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"In oral squamous cell carcinoma cells, knockdown of Tspan15 reduced ADAM10 expression, decreased soluble N-cadherin shedding, reduced nuclear β-catenin immunoreactivity, and suppressed tumor invasion and migration, placing Tspan15 upstream of ADAM10-mediated N-cadherin shedding and downstream Wnt/β-catenin signaling in OSCC metastasis.\",\n      \"method\": \"siRNA knockdown, N-cadherin shedding ELISA, immunofluorescence for β-catenin localization, Transwell invasion/migration assays\",\n      \"journal\": \"Experimental cell research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — siRNA KD with multiple mechanistic readouts (shedding, localization, invasion) but single lab and no rescue experiment\",\n      \"pmids\": [\"31518558\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Tspan15 overexpression in the hepatoma cell line HepG2 increases ERK1/2 phosphorylation, leading to increased CTGF expression and secretion; proteomic profiling of Tspan15 complexes identified multiple membrane proteins including growth factor receptors as interaction partners.\",\n      \"method\": \"Overexpression, western blotting for p-ERK1/2, quantitative secretome proteomics (MS), Tspan15 complex immunoprecipitation/MS\",\n      \"journal\": \"Proteomics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — overexpression with western blot and MS-based proteomics; single lab, no loss-of-function rescue\",\n      \"pmids\": [\"31390680\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"TSPAN15 directly interacts with integrin-β1 (ITGB1) and maintains ITGB1 stability by inhibiting its ubiquitination; this interaction activates downstream p-FAK/p-AKT/p-mTOR signaling and promotes GPX4 expression, thereby attenuating gemcitabine-induced ferroptosis in pancreatic ductal adenocarcinoma cells.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, western blotting for FAK/AKT/mTOR phosphorylation and GPX4, TSPAN15 knockdown in vitro and in vivo xenograft, ferroptosis assays\",\n      \"journal\": \"Redox biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP plus functional pathway assays and in vivo xenograft; single lab, novel ITGB1 interaction outside established ADAM10 biology\",\n      \"pmids\": [\"40505345\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"TSPAN15 physically interacts with BTRC to promote proteasomal degradation of tumor suppressor PDCD4 via ubiquitination, thereby activating autophagy and autophagy-mediated EMT and metastasis in hepatocellular carcinoma cells.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, autophagic flux analysis, TSPAN15 silencing, PDCD4 rescue experiments, xenograft mouse model\",\n      \"journal\": \"Molecular immunology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP plus functional ubiquitination assay, autophagic flux, and in vivo model; single lab; mechanistically consistent with prior BTRC interaction data\",\n      \"pmids\": [\"40398082\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"In intrahepatic cholangiocarcinoma (ICC) cells, Tspan15 mediates translocation of activated mature ADAM10 from the cytoplasm to the cell membrane surface, which cleaves the Notch1 intracellular domain from its extracellular domain, activating Notch1 signaling and enhancing cancer stem cell-like properties, EMT, and chemoresistance against gemcitabine and cisplatin.\",\n      \"method\": \"Western blotting, flow cytometry, immunohistochemistry, RT-PCR, Tspan15/ADAM10 knockdown, Notch1 activation assays, chemoresistance functional assays\",\n      \"journal\": \"Liver international : official journal of the International Association for the Study of the Liver\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — knockdown with multiple functional readouts (Notch activation, CSC markers, chemoresistance); mechanistically consistent with established Tspan15-ADAM10-Notch axis; single lab\",\n      \"pmids\": [\"37545390\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"In invasive bladder cancer cells, Tspan15 is required for ADAM10-mediated selective cleavage of N-cadherin; the PPARβ/δ agonist GW501516 decreases Tspan15 expression and prevents N-cadherin cleavage (NTF generation) without modifying ADAM10 expression levels, demonstrating that pharmacological targeting of Tspan15 can selectively block ADAM10 substrate cleavage.\",\n      \"method\": \"siRNA knockdown of Tspan15, western blotting for N-cadherin fragments (NTF/CTF1), GW501516 treatment, ADAM10 expression analysis\",\n      \"journal\": \"Cells\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — siRNA KD plus pharmacological intervention with specific mechanistic readout; single lab; consistent with established Tspan15-ADAM10-N-cadherin mechanism\",\n      \"pmids\": [\"38667323\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"A tricomponent complex of THSD7A/ADAM10/Tspan15 was found in podocytes; Tspan15 regulates ADAM10 substrate usage and the stability of podocyte cell surface proteins (THSD7A, PLA2R1, β-dystroglycan), where THSD7A acts as both an ADAM10 substrate and regulator of the complex; Tspan15 is present at podocyte foot processes.\",\n      \"method\": \"Co-localization and Co-IP, ADAM10-deficient mice, ADAM10-inhibited pig glomeruli, in vitro shedding assays, biochemical fractionation\",\n      \"journal\": \"Kidney international\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP plus genetic/pharmacological ADAM10 inhibition across multiple experimental systems; tricomponent complex is a novel finding from single lab\",\n      \"pmids\": [\"40339751\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Silencing of Tspan15 (and Tspan10) in astrocytoma cells reduced Venezuelan equine encephalitis virus (VEEV) genome replication without affecting viral entry; silencing of the ADAM10 substrate N-cadherin also reduced VEEV infectivity, suggesting Tspan15/ADAM10 and downstream substrates modulate VEEV replication.\",\n      \"method\": \"siRNA silencing, pharmacological ADAM10 inhibition, VEEV infection assays, viral entry vs. replication discrimination assays\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — siRNA KD with infectivity readout; single lab, single method per conclusion; indirect mechanism\",\n      \"pmids\": [\"39878649\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"TSPAN15 (identified as NET-7) was identified and sequenced as a new tetraspanin family member with the characteristic structure of four transmembrane domains, two extracellular regions, and conserved tetraspanin amino acid residues; it shows differential expression across human cell lines.\",\n      \"method\": \"EST sequencing, sequence analysis, RT-PCR expression profiling\",\n      \"journal\": \"Biochimica et biophysica acta\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — sequence/expression characterization only; no functional mechanism established\",\n      \"pmids\": [\"10719184\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"TSPAN15 is a TspanC8-subfamily tetraspanin that forms a direct, stoichiometric complex with the transmembrane metalloprotease ADAM10; structural and biochemical studies show that Tspan15 binds ADAM10 via its large extracellular loop (LEL) interacting with the stalk/cysteine-rich/disintegrin domains of ADAM10, relieves ADAM10 autoinhibition, and positions the catalytic active site ~20 Å above the plasma membrane as a 'molecular measuring stick' for membrane-proximal substrate cleavage. TSPAN15 promotes ADAM10 exit from the endoplasmic reticulum, stabilizes the mature active form of ADAM10 at the cell surface by slowing its endocytosis, and confers substrate selectivity on the complex—particularly for N-cadherin, PrP, and GPVI cleavage—distinct from other TspanC8/ADAM10 pairs. In cancer contexts, TSPAN15 also interacts with BTRC to promote IκBα degradation and NF-κB activation, and interacts with integrin-β1 to activate FAK/AKT/mTOR/GPX4 signaling and suppress ferroptosis; additionally, Tspan15 is required at the surface of neuronal extracellular vesicles to promote their docking at target neurons.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"TSPAN15 is a TspanC8-subfamily tetraspanin that functions as a dedicated regulatory partner of the transmembrane metalloprotease ADAM10, controlling its maturation, surface stability, and substrate selectivity [#0, #1]. It directly binds ADAM10 and accelerates exit of the complex from the endoplasmic reticulum, stabilizing the active/mature enzyme at the plasma membrane by slowing its endocytosis; ADAM10 reciprocally stabilizes TSPAN15, and endogenous TSPAN15 is unstable in the absence of ADAM10 [#0, #6, #7]. The interaction is mediated by the TSPAN15 large extracellular loop (LEL) contacting the membrane-proximal stalk, cysteine-rich, and disintegrin domains of ADAM10; crystallographic and cryo-EM structures show that TSPAN15 binding relieves ADAM10 autoinhibition and acts as a 'molecular measuring stick', positioning the catalytic active site approximately 20 Å above the membrane to dictate membrane-proximal cleavage-site selection [#4, #8, #9]. Through this mechanism TSPAN15 confers substrate specificity distinct from other TspanC8/ADAM10 pairs, preferentially directing ADAM10 toward N-cadherin, cellular prion protein, and GPVI rather than APP or Notch targets, as confirmed in knockout mice [#5, #11]. Beyond the canonical protease axis, TSPAN15 has been implicated in cancer signaling—interacting with the BTRC E3 ligase to promote IκBα and PDCD4 degradation [#10, #16] and with integrin-β1 to sustain FAK/AKT/mTOR signaling [#15]—and is required at the surface of neuronal extracellular vesicles for their docking at target neurons [#12].\"\n,\n  \"teleology\": [\n    {\n      \"year\": 2000,\n      \"claim\": \"Before any functional role was known, TSPAN15 had to be defined as a distinct gene; its identification as the tetraspanin NET-7 established the four-transmembrane architecture and conserved residues that place it in the tetraspanin family.\",\n      \"evidence\": \"EST sequencing, sequence analysis, and RT-PCR expression profiling across human cell lines\",\n      \"pmids\": [\"10719184\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No functional mechanism established\", \"Differential expression not linked to any pathway\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"The central question of what TSPAN15 does was answered by showing it directly binds ADAM10, accelerates ER exit of the complex, stabilizes mature surface ADAM10, and increases shedding of N-cadherin and APP—establishing TSPAN15 as an ADAM10 trafficking and maturation regulator.\",\n      \"evidence\": \"Split-ubiquitin Y2H, co-IP, ER-retention mutant pulse-chase, RNAi, and substrate shedding assays in mammalian cells; reciprocal Co-IP and maturation assays establishing the broader TspanC8 family role and Notch regulation\",\n      \"pmids\": [\"22446748\", \"23035126\", \"23091066\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not resolve whether substrate preference differs among TspanC8 members\", \"Structural basis of the interaction not defined\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"To explain why different TspanC8s yield different ADAM10 outputs, studies mapped the interaction interface and demonstrated compartment-specific and substrate-specific effects, showing TSPAN15 uniquely promotes N-cadherin cleavage and places ADAM10 in a distinct membrane environment.\",\n      \"evidence\": \"Chimeric ADAM10/TspanC8 constructs, sucrose gradient fractionation, single-molecule tracking, quantitative MS co-IP, and substrate cleavage/Notch reporter assays\",\n      \"pmids\": [\"26686862\", \"26668317\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism of how compartmentalization selects substrates not structurally defined\", \"In vivo relevance not yet tested\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Genetic knockout in mice resolved whether TSPAN15 substrate selectivity operates in vivo, showing loss of mature ADAM10 in brain and age-dependent loss of N-cadherin and PrP shedding but not APP or Notch readouts—proving physiological substrate steering.\",\n      \"evidence\": \"Tspan15 knockout mouse brain analyzed by western blotting for ADAM10 maturation and substrate-specific shedding and Notch target gene expression\",\n      \"pmids\": [\"29520422\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular basis for in vivo substrate discrimination not addressed\", \"Phenotypic consequences for brain physiology not characterized\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"The mechanism of surface stabilization was clarified by showing TSPAN15 slows ADAM10 endocytosis (in contrast to Tspan5), with the cytoplasmic domain—not the C-terminal palmitoylation site—governing trafficking and Notch outcomes.\",\n      \"evidence\": \"Endocytosis assays, cell surface biotinylation, flow cytometry, chimeric cytoplasmic-domain constructs, and palmitoylation-site mutagenesis\",\n      \"pmids\": [\"31792032\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Endocytic adaptor recognizing the cytoplasmic domain not identified\", \"Quantitative contribution of trafficking versus stability to net activity unresolved\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"To confirm physiological relevance and therapeutic tractability, work on endogenous proteins showed ADAM10 is the principal TSPAN15 partner, TSPAN15 is unstable without ADAM10, a synthetic fusion is a functional scissor, and antibodies can inhibit the complex.\",\n      \"evidence\": \"Monoclonal antibody generation, endogenous co-localization and co-IP, ADAM10-KO cell lines, synthetic fusion protein, and antibody inhibition of substrate cleavage\",\n      \"pmids\": [\"32111735\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Epitopes of inhibitory antibodies not mapped\", \"Therapeutic specificity against other TspanC8/ADAM10 complexes not established\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"The structural logic of substrate selection was resolved when crystal and cryo-EM structures showed the TSPAN15 LEL binding site relieves ADAM10 autoinhibition and acts as a molecular measuring stick positioning the active site ~20 Å above the membrane to dictate cleavage-site choice.\",\n      \"evidence\": \"X-ray crystallography of the TSPAN15 LEL with mutagenesis and N-cadherin cleavage assays; cryo-EM of a vFab-ADAM10-Tspan15 complex with interface mutagenesis and cell-based shedding assays\",\n      \"pmids\": [\"34739841\", \"37516108\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structures of complexes with other TspanC8s for comparison not available\", \"Dynamics of substrate engagement not captured\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Substrate-selectivity rules were extended by showing TSPAN15 and Tspan33 act compensatorily in ADAM10-mediated GPVI cleavage, with TSPAN15 dominant and its extracellular region setting the cleavage distance.\",\n      \"evidence\": \"CRISPR/Cas9 knockout cell lines with TSPAN15 and GPVI mutant constructs and GPVI cleavage assays\",\n      \"pmids\": [\"35269584\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism of TSPAN15/Tspan33 compensation not defined\", \"Platelet-physiological consequences not tested here\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"A non-protease cancer role was opened by showing TSPAN15 binds the BTRC E3 ligase to drive phospho-IκBα ubiquitination and NF-κB activation of metastasis genes, later paralleled by BTRC-dependent PDCD4 degradation driving autophagy and EMT.\",\n      \"evidence\": \"Co-IP, ubiquitination assays, NF-κB reporter, and target gene analysis in oesophageal carcinoma; co-IP, ubiquitination, autophagic flux, PDCD4 rescue, and xenograft in hepatocellular carcinoma\",\n      \"pmids\": [\"29650964\", \"40398082\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct versus ADAM10-dependent nature of these effects not separated\", \"How a tetraspanin engages a cytosolic E3 ligase mechanistically unclear\", \"Independent replication of BTRC axis limited\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"TSPAN15 was assigned a function distinct from ADAM10 in neuronal extracellular vesicle biology, where it is required at the EV surface for docking at target neurons rather than at the recipient membrane.\",\n      \"evidence\": \"EV fractionation and marker analysis, fluorescent fusion tracking, and Tspan15-knockout cortical neuron EV target-cell docking assays\",\n      \"pmids\": [\"38938373\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Docking receptor/ligand on target neurons not identified\", \"Whether ADAM10 is involved not resolved\", \"Single lab\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"An ADAM10-independent oncogenic axis was proposed in which TSPAN15 binds integrin-β1, blocks its ubiquitination, and sustains FAK/AKT/mTOR/GPX4 signaling to suppress ferroptosis and confer chemoresistance.\",\n      \"evidence\": \"Co-IP, ubiquitination assay, phospho-signaling and GPX4 westerns, knockdown in vitro and in vivo xenograft, and ferroptosis assays in pancreatic cancer cells\",\n      \"pmids\": [\"40505345\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct binding interface with ITGB1 not mapped\", \"Independence from ADAM10 not formally tested\", \"Single lab, novel interaction\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Disease-tissue context was extended by identifying a THSD7A/ADAM10/TSPAN15 tricomponent complex at podocyte foot processes that regulates ADAM10 substrate usage and surface protein stability.\",\n      \"evidence\": \"Co-localization and Co-IP, ADAM10-deficient mice, ADAM10-inhibited pig glomeruli, in vitro shedding assays, and biochemical fractionation\",\n      \"pmids\": [\"40339751\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether THSD7A binds TSPAN15 directly not resolved\", \"Pathophysiological link to membranous nephropathy not established\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"It remains unresolved how the canonical ADAM10-scaffolding role of TSPAN15 mechanistically relates to its reported ADAM10-independent activities (BTRC/E3-ligase coupling, integrin-β1 stabilization, EV docking), and whether these reflect distinct molecular interfaces of the same protein.\",\n      \"evidence\": \"No single study reconciles the protease-scaffold and cytosolic-signaling functions\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No structural or interface data for the cytosolic/cancer interactions\", \"No demonstration that protease-independent roles occur without ADAM10 present\", \"In vivo physiological weighting of the multiple roles unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [0, 5, 9]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [0, 7, 9]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [6, 7, 9]},\n      {\"term_id\": \"GO:0005783\", \"supporting_discovery_ids\": [0, 4]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0, 5, 9]}\n    ],\n    \"complexes\": [\n      \"ADAM10-TSPAN15 sheddase complex\",\n      \"THSD7A/ADAM10/TSPAN15 complex\"\n    ],\n    \"partners\": [\n      \"ADAM10\",\n      \"BTRC\",\n      \"ITGB1\",\n      \"THSD7A\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":5,"faith_pct":100.0}}