{"gene":"GP6","run_date":"2026-06-10T01:55:21","timeline":{"discoveries":[{"year":1997,"finding":"Convulxin, a C-type lectin from snake venom, activates platelets through the p62/GPVI collagen receptor (not GPIb), inducing tyrosine phosphorylation of FcRγ chain, PLCγ2, Syk (p72), c-Cbl, and p36-38 in a pattern similar to collagen. Convulxin binds directly to GPVI immunoprecipitated from platelet lysates, and convulxin subunits inhibit both aggregation and tyrosine phosphorylation in response to collagen. Src family kinases phosphorylate FcRγ chain upon GPVI clustering, activating Syk downstream.","method":"125I-convulxin binding, immunoprecipitation, tyrosine phosphorylation western blot, platelet aggregation assay, tyrosine kinase inhibitor (piceatannol)","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal binding assay, Co-IP, phosphorylation cascade mapping, inhibitor studies, replicated across multiple methods in single study","pmids":["9153205"],"is_preprint":false},{"year":2001,"finding":"GPVI expression in RBL-2H3 cells (which express FcRγ chain) confers adhesive and signaling responses to convulxin via the Fc receptor γ-chain. Both the transmembrane arginine of GPVI and its intracellular C-terminal tail are required for coupling to FcRγ and signal transduction. GPVI expression was sufficient for convulxin responses but not for full collagen signaling, suggesting an additional collagen-binding receptor is required.","method":"Transfection of wild-type and mutant GPVI into RBL-2H3 cells, adhesion assays, signaling readouts, mutagenesis of transmembrane domain and C-tail","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — direct mutagenesis of functional domains with multiple orthogonal readouts (adhesion, signaling), reconstitution in heterologous cells","pmids":["11152687"],"is_preprint":false},{"year":2001,"finding":"GPVI is expressed early during megakaryocyte differentiation (CD41+ stage) and associates with the FcRγ chain in megakaryocytes as in platelets. Functional GPVI-mediated signaling (tyrosine phosphorylation of Syk, FcRγ, PLCγ2) is activated by convulxin in mature megakaryocytes; adhesion to immobilized convulxin occurs only at late stages of maturation when GPVI expression increases.","method":"Flow cytometry, RT-PCR, immunoblotting, ligand blotting, adhesion assays, tyrosine phosphorylation analysis","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (flow cytometry, biochemistry, functional adhesion), clear functional-expression correlation","pmids":["11278467"],"is_preprint":false},{"year":2001,"finding":"Alborhagin, a viper venom metalloproteinase, activates platelets through GPVI (not through a distinct receptor), inducing a tyrosine phosphorylation pattern similar to convulxin including PLCγ2 phosphorylation. Alborhagin-induced aggregation in mouse platelets is inhibited by anti-GPVI monoclonal antibody JAQ1, and alborhagin induces GPVI-dependent responses in GPVI-transfected K562 and Jurkat cells.","method":"Platelet aggregation, tyrosine phosphorylation western blot, transfected cell signaling, antibody inhibition (JAQ1), Src kinase inhibitor PP1","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — GPVI-transfected cell confirmation, antibody blockade, multiple orthogonal readouts","pmids":["11344165"],"is_preprint":false},{"year":2001,"finding":"GPVI mediates both adhesive and signaling responses to collagen in a receptor density-dependent manner. GPVI-FcRγ expression alone is sufficient to confer collagen adhesion and signaling in RBL-2H3 cells when GPVI density matches that on human platelets; collagen responses are proportional to receptor density. GPVI receptor density on human platelets is tightly regulated and independent of α2β1 expression.","method":"Novel anti-GPVI monoclonal antibody quantification, GPVI-expressing RBL-2H3 cell lines at varying receptor densities, adhesion and signaling assays","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — reconstitution in heterologous cells with defined receptor densities, multiple orthogonal readouts, resolves conflicting prior data","pmids":["11723134"],"is_preprint":false},{"year":2002,"finding":"Dimeric GPVI (GPVI-Fc fusion) but not monomeric GPVI (GPVIex) exhibits high affinity binding to fibrous collagen (KD ~576 nM by SPR), inhibits collagen-induced platelet aggregation, and inhibits collagen-related peptide-induced aggregation. Monomeric GPVI inhibited convulxin- but not collagen-induced aggregation. The convulxin binding site on GPVI is distinct from the collagen-binding site. GPVI is proposed to exist as a dimeric form on platelets for high-affinity collagen interaction.","method":"ELISA, surface plasmon resonance (SPR), platelet aggregation inhibition assays with soluble monomeric and dimeric recombinant GPVI ectodomains","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro binding reconstitution with SPR kinetics, multiple functional assays, rigorous controls distinguishing monomer vs. dimer","pmids":["12356768"],"is_preprint":false},{"year":1999,"finding":"LAT (linker for activation of T cells) is required for tyrosine phosphorylation of PLCγ2 downstream of the GPVI collagen receptor. LAT is tyrosine-phosphorylated downstream of Syk (not SLP-76 or Btk) following GPVI activation. In LAT-deficient platelets, PLCγ2 phosphorylation is substantially reduced, leading to marked inhibition of phosphatidic acid formation, pleckstrin phosphorylation, P-selectin expression, and reduced αIIbβ3 activation in response to CRP. A minor LAT-independent pathway of PLCγ2 phosphorylation exists.","method":"LAT-deficient mouse platelets, Syk-deficient mouse platelets, XLA human platelets (Btk-deficient), SLP-76-deficient mouse platelets, tyrosine phosphorylation western blot, metabolite measurement, flow cytometry","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic knockout models with multiple orthogonal biochemical and functional readouts, epistasis established","pmids":["10567557"],"is_preprint":false},{"year":2002,"finding":"SLP-76 plays a greater role than LAT in platelet activation downstream of GPVI. LAT-deficient platelets aggregate normally to high concentrations of collagen and convulxin, while SLP-76-deficient platelets do not. LAT is not tyrosine-phosphorylated after fibrinogen binding to integrin αIIbβ3; collagen-stimulated LAT-deficient platelets spread normally on fibrinogen. These findings establish differential requirements for LAT and SLP-76 in GPVI versus TCR signaling.","method":"LAT-/- and SLP-76-/- mouse platelets, platelet aggregation, P-selectin expression, spreading assays, phosphorylation western blot","journal":"The Journal of experimental medicine","confidence":"High","confidence_rationale":"Tier 2 / Strong — comparative genetic knockout models, multiple orthogonal functional readouts, epistasis between adaptor proteins","pmids":["11901197"],"is_preprint":false},{"year":2003,"finding":"Tec family kinase Tec compensates for Btk downstream of GPVI in regulating PLCγ2 phosphorylation. Btk/Tec double-deficient platelets fail to undergo Ca2+ increase, aggregation, secretion, and spreading in response to collagen or CRP, while single knockouts show partial defects. A residual GPVI signal persists in Btk/Tec double-deficient platelets as CRP synergizes with ADP to mediate aggregation.","method":"Btk-/-, Tec-/-, and Btk/Tec double-knockout mouse platelets, aggregation, Ca2+ measurement, secretion, spreading, PLCγ2 phosphorylation western blot","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 / Strong — double-knockout epistasis, multiple functional readouts, establishes Tec as redundant kinase for PLCγ2 activation downstream of GPVI","pmids":["12842985"],"is_preprint":false},{"year":2003,"finding":"Activation of GPVI by collagen is regulated by α2β1 integrin and secondary mediators (ADP, thromboxanes). α2β1 blockade has a greater inhibitory effect on collagen-induced PLC activation, Ca2+ elevation, and dense granule secretion than ADP receptor antagonists plus COX inhibitor. The major role of α2β1 is proposed to increase avidity of collagen for the platelet surface, thereby enhancing GPVI activation. Secondary mediators indirectly regulate GPVI signaling via activation of α2β1.","method":"Platelet aggregation, tyrosine phosphorylation, Ca2+ measurement, dense granule secretion with pharmacological inhibitors of α2β1, ADP receptors, and COX; integrin αIIbβ3 antagonist to isolate signaling from aggregation","journal":"Journal of thrombosis and haemostasis","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — pharmacological approach with multiple signaling readouts, single lab","pmids":["12871331"],"is_preprint":false},{"year":2004,"finding":"GPVI undergoes metalloproteinase-dependent ectodomain shedding generating a ~55 kDa soluble fragment released from platelets. This shedding is induced by mitochondrial injury (CCCP) and is inhibited by the broad-spectrum metalloproteinase inhibitor GM6001. CRP or thrombin stimulation induces shedding of GPIbα but not GPVI, suggesting different metalloproteinases regulate the two receptors or an additional signal is needed to render GPVI susceptible to cleavage.","method":"Mouse platelet stimulation with CCCP, GM6001 inhibition, immunoblotting of supernatant and lysate, platelet aggregation and CRP/thrombin response assays","journal":"Thrombosis and haemostasis","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct biochemical detection of ectodomain in supernatant, metalloproteinase inhibitor, functional readouts, single lab","pmids":["15116256"],"is_preprint":false},{"year":2005,"finding":"GPVI is a receptor for laminin. Lamellipodia (but not filopodia) formation on laminin requires GPVI, as demonstrated in GPVI-knockout mouse platelets. Integrin α6β1 is essential for adhesion to laminin and brings laminin to GPVI. GPVI-laminin interaction was confirmed by surface plasmon resonance spectroscopy and by lamellipodia formation on laminin in the presence of collagenase. Syk kinase is required for signaling downstream of both GPVI and α6β1.","method":"GPVI-/- mouse platelets, α6-blocking antibody, Syk-/- mouse platelets, surface plasmon resonance, collagenase treatment, platelet spreading microscopy","journal":"Blood","confidence":"High","confidence_rationale":"Tier 1 / Strong — SPR binding plus genetic knockout functional validation, collagenase control, multiple orthogonal approaches in single study","pmids":["16219796"],"is_preprint":false},{"year":2005,"finding":"Both GPVI and integrin α2β1 play independent critical roles in platelet adhesion to collagen under flow; loss of both receptors completely ablates this response. Intracellular signaling mediated by SLP-76 (but not LAT) is critical for platelet adhesion to collagen under flow. Reduced GPVI receptor density results in severe defects in platelet adhesion to collagen under flow.","method":"Pharmacological and genetic approaches with human and mouse platelets under flow conditions; SLP-76-/- and LAT-/- mice; GPVI density variation; tail bleeding times","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic knockout plus pharmacological inhibition, flow assays, epistasis between adaptor proteins and receptor density","pmids":["15886326"],"is_preprint":false},{"year":2005,"finding":"Syk undergoes rapid ubiquitination upon GPVI activation by collagen, CRP, and convulxin; this process is downstream of Src family kinases. Ubiquitinated Syk is ~5-fold more kinase-active than non-ubiquitinated Syk and represents 60-75% of active (pTyr525/526) Syk. In c-Cbl-deficient mice, Syk is not ubiquitinated and fails to be dephosphorylated, implicating c-Cbl as the E3 ubiquitin ligase for Syk in GPVI signaling.","method":"c-Cbl-/- mouse platelets, Src kinase inhibitor, ubiquitin immunoprecipitation, in vitro kinase assay on separated ubiquitinated/non-ubiquitinated Syk, phosphospecific antibody to Syk","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic knockout (c-Cbl-/-), in vitro kinase assay, multiple orthogonal methods identifying c-Cbl as E3 ligase","pmids":["15701717"],"is_preprint":false},{"year":2005,"finding":"Thrombopoietin (TPO) initiates CpG demethylation of the GP6 promoter, which drives GPVI transcription during megakaryocyte differentiation. The GP6 promoter CpG island is fully methylated in GPVI-non-expressive cell lines and completely unmethylated in GPVI-expressive lines. TPO treatment of UT-7/EPO-Mpl cells induces GP6 promoter demethylation correlated with increased mRNA.","method":"Sodium bisulfite genomic sequencing of CpG island, RT-PCR for mRNA quantification, TPO treatment of primary cord blood cells and cell lines","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct bisulfite sequencing of promoter methylation, multiple cell lines and primary cells, TPO treatment experiments","pmids":["15701720"],"is_preprint":false},{"year":2004,"finding":"Collagen type I-induced stress fiber formation in megakaryocytes requires α2β1 integrin (essential) and GPVI (triggers rapid and sustained ERK1/2 MAPK activation). Both pathways converge via synergistic MAPK/ERK1/2 and Rho/ROCK activation. After longer adhesion, proplatelet formation is inhibited by α2β1 but not by GPVI, through the Rho/ROCK pathway. This demonstrates differential regulation of megakaryocyte actin dynamics by the two collagen receptors.","method":"Function-blocking antibodies against α2β1 and GPVI-specific ligands in primary human megakaryocytes, kinase inhibitors (ROCK, ERK1/2), F-actin staining, proplatelet formation assay","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 / Strong — receptor-specific ligands and blocking antibodies, kinase inhibitors, multiple functional readouts, primary human cells","pmids":["15265786"],"is_preprint":false},{"year":2007,"finding":"ADAM10, but not ADAM17, cleaves a GPVI-based synthetic peptide within the extracellular membrane-proximal sequence (PAR;Q243YY) as identified by MALDI-TOF-MS. GPVI and GPIbα are shed by distinct ADAM family members (ADAM10 for GPVI, ADAM17 for GPIbα) in response to distinct stimuli (CCCP, PMA, W7, NEM). A significant fraction of GPIbα but not GPVI exists in cleaved state on resting platelets.","method":"Immunoblotting with affinity-purified cytoplasmic domain antibodies, MALDI-TOF-MS of synthetic peptide cleavage products, ADAM10/ADAM17 recombinant enzyme assays, metalloproteinase inhibitors, mutagenesis of cleavage sequences in transfected cells","journal":"Journal of thrombosis and haemostasis","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro peptide cleavage with mass spectrometry, mutagenesis validation, biochemical fractionation","pmids":["17445093"],"is_preprint":false},{"year":2007,"finding":"GPVI down-regulation occurs through two distinct pathways downstream of the FcRγ-ITAM: ectodomain shedding (via metalloproteinases, requiring LAT and PLCγ2 signaling) or internalization/intracellular clearing (independent of shedding). Both pathways are abrogated in mice with a point mutation in the FcRγ ITAM. In LAT-/- or PLCγ2-/- mice, GPVI shedding is abolished but receptor is down-regulated through internalization without thrombocytopenia or altered thrombin responses.","method":"FcRγ ITAM point mutant mice, LAT-/- mice, PLCγ2-/- mice, anti-GPVI antibody (JAQ1) treatment, flow cytometry of GPVI surface levels, platelet count monitoring, thrombin response assays","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple genetic models (point mutant + knockouts), epistasis establishing two distinct signaling branches, multiple functional readouts","pmids":["17374738"],"is_preprint":false},{"year":2009,"finding":"GPVI ectodomain shedding in vitro can be mediated by either ADAM10 or ADAM17 in response to distinct stimuli. In vivo antibody (JAQ1)-induced GPVI shedding occurs even in mice lacking both ADAM10 and ADAM17 in platelets, establishing the existence of a third GPVI-cleaving platelet enzyme.","method":"Megakaryocyte-specific ADAM10-deficient mice, Adam17ex/ex mice, ADAM10/ADAM17 double-deficient platelets in vitro and in vivo, anti-GPVI antibody JAQ1 treatment, flow cytometry","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 / Strong — two genetic models plus double deficiency, in vitro and in vivo shedding experiments","pmids":["20644114"],"is_preprint":false},{"year":2009,"finding":"GPVI constitutively associates in resting platelets with the Src-family kinase Lyn via a unique intracellular proline-rich domain (PRD), which directly activates Lyn presumably through SH3 displacement. Lyn is bound to GPVI in an activated state. Platelets lacking Lyn exhibit defective collagen adhesion under flow similar to platelets with GPVI receptors lacking the PRD. This PRD-Lyn interaction constitutes a molecular priming mechanism enabling rapid hemostatic response.","method":"Co-immunoprecipitation of Lyn with GPVI, PRD deletion mutant GPVI, Lyn-/- mouse platelets, platelet adhesion to collagen under flow, in vitro binding assays","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, PRD mutant analysis, Lyn-/- genetic model, flow adhesion assay, multiple orthogonal methods","pmids":["19940238"],"is_preprint":false},{"year":2009,"finding":"GPVI is a component of tetraspanin microdomains on platelets, specifically associating with Tspan9 and integrin α6β1, but not with GPIbα or integrins αIIbβ3 or α2β1. This selective tetraspanin microdomain association was established by Co-IP and co-localization.","method":"Co-immunoprecipitation, western blotting, flow cytometry with novel Tspan9 antibodies, megakaryocyte SAGE and DNA microarrays","journal":"The Biochemical journal","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Co-IP evidence of GPVI in specific tetraspanin microdomain, single lab","pmids":["18795891"],"is_preprint":false},{"year":2009,"finding":"GPVI functions as a receptor for EMMPRIN (CD147/basigin) and mediates platelet rolling on immobilized EMMPRIN under arterial shear. Anti-GPVI blocking antibodies reduce platelet rolling on EMMPRIN-Fc. CHO cells transfected with GPVI show enhanced rolling on EMMPRIN-Fc; CHO cells transfected with EMMPRIN show enhanced rolling on GPVI-Fc. Direct GPVI-EMMPRIN binding was confirmed by ELISA and SPR with KD = 88 nM.","method":"Flow-based platelet rolling assay, blocking monoclonal antibodies (anti-EMMPRIN, anti-GPVI), CHO cell transfection, modified ELISA, surface plasmon resonance","journal":"Thrombosis and haemostasis","confidence":"High","confidence_rationale":"Tier 1 / Strong — SPR with KD measurement, transfected cell assays, blocking antibodies, multiple orthogonal methods","pmids":["19350111"],"is_preprint":false},{"year":2009,"finding":"Lyn, PKC-δ, and SHIP-1 form a complex that specifically regulates GPVI-mediated (but not PAR-mediated) dense granule secretion. GPVI stimulation induces rapid SHIP-1 phosphorylation at Y1020 and association of SHIP-1 and Lyn with PKC-δ. In Lyn-/- platelets, GPVI-mediated phosphorylations of SHIP-1 (Y1020) and PKC-δ (Y311) are inhibited. In Lyn-/- or SHIP-1-/- platelets, GPVI-mediated dense granule secretion is potentiated (negative regulation), while PAR-mediated secretion is inhibited. Lyn-mediated phosphorylation of PKC-δ and SHIP-1 negatively regulates GPVI-specific dense granule secretion.","method":"PKC-δ-/-, Lyn-/-, SHIP-1-/- mouse platelets, Co-IP, phosphospecific antibodies, dense granule secretion assays (convulxin vs PAR agonists)","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple genetic knockout models, reciprocal Co-IP, phosphospecific antibodies, comparing GPVI vs. PAR pathways","pmids":["19587372"],"is_preprint":false},{"year":2009,"finding":"The crystal structure of LAIR-1 (a collagen receptor homologous to GPVI's collagen-binding domain) was determined. NMR titrations and mutagenesis mapped the collagen-binding site: residues R59, E61, and W109 are key for collagen interaction. These residues are strictly conserved in both LAIR-1 and GPVI but are located outside the previously proposed GPVI collagen-binding site, revealing an unanticipated common mechanism of collagen recognition.","method":"X-ray crystallography (LAIR-1 structure), NMR titrations with collagen peptides, mutagenesis of collagen-contact residues","journal":"Blood","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure plus NMR plus mutagenesis, directly informing GPVI collagen-binding mechanism through conserved residues","pmids":["20007810"],"is_preprint":false},{"year":2010,"finding":"GPVI signals through an ITAM in the associated FcRγ chain via sequential Src family kinase-mediated phosphorylation of the two ITAM tyrosines, followed by Syk SH2 domain binding and activation, culminating in PLCγ2 activation. This mechanistic pathway was established and compared to CLEC-2 signaling (which uses a hemITAM).","method":"Review consolidating genetic knockout and biochemical data from multiple studies; mechanistic pathway established by epistasis and phosphorylation cascade analysis","journal":"Journal of thrombosis and haemostasis","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — review synthesizing genetic and biochemical data; pathway mechanistically established across multiple prior experiments","pmids":["20345705"],"is_preprint":false},{"year":2011,"finding":"Cdc42 GTPase is required for GPVI-dependent (CRP-induced) filopodia formation, secretion (ATP and P-selectin), and aggregation in platelets. Cdc42-/- platelets show diminished PAK1/2 phosphorylation (Cdc42 effector) and minimal Akt phosphorylation upon CRP stimulation, establishing Cdc42 as a downstream effector in GPVI signaling.","method":"Mx-cre;Cdc42lox/lox inducible knockout mice, CRP and thrombin stimulation, platelet aggregation, P-selectin and ATP release, spreading assay, PAK1/2 phosphorylation, Akt phosphorylation, bleeding time","journal":"PloS one","confidence":"High","confidence_rationale":"Tier 2 / Strong — inducible genetic knockout, multiple functional readouts (aggregation, secretion, spreading), pathway marker phosphorylation","pmids":["21789221"],"is_preprint":false},{"year":2013,"finding":"An adenine insertion (c.711_712insA) in exon 6 of GP6 generates a premature stop codon, producing a truncated GPVI protein (~49 kDa) that lacks the transmembrane domain and is absent from the platelet surface. Platelets from homozygous patients have absent surface GPVI, no aggregation in response to collagen, convulxin, or CRP, and no 14C-5-HT secretion with these agonists, directly establishing that the transmembrane domain is required for functional surface expression.","method":"DNA sequencing, mRNA sequencing, western blotting, flow cytometry, immunofluorescence confocal microscopy, platelet aggregation, 14C-5-HT secretion","journal":"Journal of thrombosis and haemostasis","confidence":"High","confidence_rationale":"Tier 2 / Strong — natural loss-of-function mutation with molecular characterization, multiple orthogonal functional assays confirming complete loss of GPVI signaling","pmids":["23815599"],"is_preprint":false},{"year":2015,"finding":"GPVI is a signaling receptor for fibrin. Thrombin-induced fibrin generation activates GPVI-dependent tyrosine phosphorylation of FcRγ and Syk in human and mouse platelets; this is abolished in GPVI-deficient mouse platelets. Mouse platelets spread fully on fibrin (but not fibrinogen) in a GPVI- and Src kinase-dependent manner, associated with phosphatidylserine exposure. The GPVI ectodomain binds directly to immobilized monomeric and polymerized fibrin.","method":"GPVI-/- mouse platelets, eptifibatide (αIIbβ3 blocker), PAR4 peptide controls, Syk and FcRγ phosphorylation western blot, platelet spreading on fibrin/fibrinogen, Src kinase inhibitor, SPR-like binding assay","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 / Strong — GPVI-/- genetic model, direct ectodomain binding to fibrin, multiple functional readouts, controls distinguishing fibrin from fibrinogen","pmids":["26282541"],"is_preprint":false},{"year":2016,"finding":"GPVI-mediated inhibition of proplatelet formation in megakaryocytes is specifically controlled by collagen type I signaling through receptor-proximal Src family kinases; Syk and LAT are dispensable for this pathway. Integrin α2β1 mediates megakaryocyte binding to collagens. Collagen type IV at the vascular niche may displace collagen I from megakaryocytes, preventing GPVI-mediated inhibition.","method":"Megakaryocytes from GPVI-/-, Syk-/-, LAT-/-, and α2β1-/- mice; blocking antibodies; Src family kinase inhibitors; adhesion assays; proplatelet formation assays","journal":"Journal of cell science","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple genetic knockout models, receptor-blocking antibodies, kinase inhibitors, well-defined functional readout (PPF)","pmids":["27505889"],"is_preprint":false},{"year":2016,"finding":"TULA-2 protein phosphatase negatively regulates Syk activation downstream of GPVI by dephosphorylating Tyr346, a regulatory (interdomain B) site of Syk that is phosphorylated early after receptor ligation and is critical for initiating full Syk activation. TULA-2 dephosphorylates Tyr346 with high efficiency but is less efficient at other Syk regulatory sites, establishing Tyr346 dephosphorylation as a key checkpoint in GPVI/Syk signaling.","method":"In vitro phosphatase assay with recombinant TULA-2 and Syk peptides/protein, phosphospecific antibodies to multiple Syk sites, GPVI-stimulated platelet signaling with TULA-2 loss-of-function","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro phosphatase assay with site-specific quantification, phosphospecific antibody panel, clear mechanistic checkpoint identification","pmids":["27609517"],"is_preprint":false},{"year":2017,"finding":"Pre-existing GPVI dimers on resting platelets coalesce into clusters upon platelet adhesion to collagenous substrates (fibrous collagen I, soluble collagen III, GPVI-specific collagen peptides). GPVI dimer cluster formation depends on a dynamic actin cytoskeleton. Cluster size is substrate-dependent (collagen III most effective). Some GPVI dimer clusters colocalize with areas of phosphotyrosine, indicating signaling activity. This clustering mechanism increases avidity for collagen and facilitates platelet activation.","method":"Total internal reflection fluorescence microscopy, direct stochastic optical reconstruction microscopy (dSTORM), confocal microscopy, fluorophore-conjugated GPVI dimer-specific Fab, actin inhibitors","journal":"Journal of thrombosis and haemostasis","confidence":"High","confidence_rationale":"Tier 2 / Strong — super-resolution microscopy (dSTORM) plus TIRF real-time imaging, multiple substrates, colocalization with phosphotyrosine signals","pmids":["28058806"],"is_preprint":false},{"year":2017,"finding":"Fibrin and D-dimer (but not the E fragment of fibrin) bind to GPVI and activate platelets. D-dimer binds GPVI by SPR with KD of 302 nM for monomeric GPVI. Immobilized D-dimer induces platelet spreading via Src and Syk kinase activation. Fibrin binds selectively to monomeric GPVI (KD 302 nM), in contrast to collagen which binds primarily to dimeric GPVI. Spreading of human platelets on fibrin is abolished in GPVI-deficient patients, confirming fibrin activates human platelets through GPVI.","method":"GPVI-deficient human patient platelets, proteolytic fibrin fragments (D-dimer, E fragment), SPR binding assays, platelet spreading on fibrin/D-dimer, Src/Syk inhibitors, soluble D-dimer inhibition assays","journal":"Blood advances","confidence":"High","confidence_rationale":"Tier 1 / Strong — SPR with KD determination, human GPVI-deficient platelets, fragment-specific binding characterization, multiple functional assays","pmids":["29296791"],"is_preprint":false},{"year":2008,"finding":"Globular adiponectin (gAd) activates platelets through the GPVI-FcRγ chain complex. gAd stimulates tyrosine phosphorylation of Syk and PLCγ2 (pattern similar to collagen), and this activation is abolished in FcRγ-null platelets (which also lack GPVI). GPVI was confirmed as the receptor for gAd by increased luciferase activity in GPVI-transfected Jurkat T-cells. Full-length adiponectin does not activate platelets through this mechanism.","method":"FcRγ-/- and PLCγ2-/- mouse platelets, NFAT luciferase reporter assay in GPVI-transfected Jurkat cells, platelet aggregation, tyrosine phosphorylation western blot","journal":"Journal of thrombosis and haemostasis","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic knockout (FcRγ-/-), GPVI transfected reporter cell assay, multiple orthogonal methods","pmids":["18419742"],"is_preprint":false},{"year":2009,"finding":"PPARγ ligands inhibit GPVI signaling through a non-genomic mechanism. PPARγ associates with Syk and LAT after platelet activation, and PPARγ agonists prevent this association, reducing tyrosine phosphorylation of multiple GPVI signaling components. The inhibitory effect is reversed by the PPARγ antagonist GW9662.","method":"Optical aggregometry, spectrofluorimetry (Ca2+ measurement), immunoblot of tyrosine phosphorylation, Co-immunoprecipitation of PPARγ with Syk/LAT, in vitro thrombus formation under flow","journal":"Journal of thrombosis and haemostasis","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Co-IP showing PPARγ-Syk/LAT association, antagonist reversal, multiple signaling readouts, single lab","pmids":["20040043"],"is_preprint":false},{"year":2014,"finding":"Nox1, but not Nox2, is the key NADPH oxidase regulating GPVI-dependent reactive oxygen species (ROS) production in platelets. Nox1-derived ROS is essential for CRP-dependent thromboxane A2 production via p38 MAPK signaling downstream of GPVI. Neither Nox1 nor Nox2 was significantly involved in CRP-induced platelet aggregation/integrin αIIbβ3 activation, spreading, or granule release. Both Nox1 and Nox2 are involved in collagen-mediated thrombus formation at arterial shear.","method":"Nox2-/- mouse platelets, ML171 (Nox1-specific pharmacological inhibitor), H2DCF-DA oxidation assay for ROS, TxA2 ELISA, p38 MAPK inhibitor, ex vivo perfusion","journal":"Redox biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic (Nox2-/-) plus pharmacological (Nox1 inhibitor) approaches, multiple downstream readouts, ex vivo flow","pmids":["24494191"],"is_preprint":false},{"year":2014,"finding":"FAK (focal adhesion kinase) is downstream of Lyn, Syk, PI3K, and Btk, and upstream of Rac1, PLCγ2, Ca2+ release, PKC, Hic-5, Nox1, and αIIbβ3 activation in the GPVI signaling pathway. FAK, but not Pyk2, is essential for GPVI-dependent ROS production, aggregation, phosphatidylserine exposure, P-selectin expression, and integrin αIIbβ3 activation.","method":"Pyk2-/- mouse platelets, FAK inhibitor PF-228, Pyk2 inhibitor Tyrphostin A9, CRP stimulation, ROS assay, aggregation, flow cytometry (PS exposure, P-selectin), immunoprecipitation and pulldown for pathway position","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic (Pyk2-/-) plus pharmacological FAK inhibition, epistasis by inhibitor cascade, multiple functional readouts, single lab","pmids":["25415317"],"is_preprint":false},{"year":2014,"finding":"CEACAM2 negatively regulates GPVI-FcRγ chain signaling in platelets. CEACAM2-/- platelets exhibit enhanced GPVI agonist (CRP, collagen)-induced aggregation, increased adhesion to type I collagen, and hyperresponsive α and dense granule release. In vivo thrombi in Ceacam2-/- mice are larger and more stable than wild-type, establishing CEACAM2 as a novel negative regulator of platelet GPVI-collagen interactions acting through immunoreceptor tyrosine-based inhibitory motifs.","method":"Ceacam2-/- mouse platelets, GPVI and CLEC-2 selective agonists, platelet aggregation, granule secretion, adhesion to collagen I, intravital microscopy (FeCl3 and laser injury models)","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic knockout, selective agonists for pathway specificity, in vitro and in vivo thrombosis models","pmids":["25085348"],"is_preprint":false},{"year":2020,"finding":"Phosphoproteomics downstream of GPVI/ITAM identified >3000 significant phosphorylation events on >1300 proteins. Key signaling relations were mapped including FcRγ→Syk→PLCγ2→PKCδ→DAPP1. Ras/MAPK axis proteins (KSR1, SOS1, STAT1, Hsp27) and >40 Rab GTPases are regulated downstream of GPVI. GPVI-mediated Rab7 S72 phosphorylation and endolysosomal maturation were blocked by TAK1 inhibition, establishing TAK1 as an upstream regulator of GPVI-induced endolysosomal signaling.","method":"Peptide TMT labeling, SPS-MS3 phosphoproteomics, causal inference analysis, TAK1 inhibitor functional validation","journal":"Blood","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — systems phosphoproteomics with TAK1 inhibitor validation for Rab7 finding; broad discovery study, individual findings partially validated","pmids":["32640021"],"is_preprint":false},{"year":2020,"finding":"GP6-homozygous platelets (carrying premature stop codon before transmembrane domain) show complete loss of surface GPVI and absence of collagen-induced spreading. Under flow, GP6hom blood fails to form stable platelet aggregates or expose phosphatidylserine (PS) on collagen or noncollagen surfaces (VWF/laminin/rhodocytin), but platelet adhesion is preserved. Thrombin generation is partially reduced. The partial adhesion retention explains the mild bleeding diathesis of GP6hom patients.","method":"Western blotting, flow cytometry, platelet spreading on collagen/VWF, whole blood perfusion over collagen or noncollagen surfaces under flow, thrombin generation assay, population frequency analysis","journal":"Blood advances","confidence":"High","confidence_rationale":"Tier 2 / Strong — natural human loss-of-function (homozygous), multiple orthogonal functional assays under flow and static conditions, distinguishes aggregation from adhesion","pmids":["32603422"],"is_preprint":false},{"year":2021,"finding":"Dimeric GPVI binds fibrinogen with much higher affinity and slower dissociation rate than monomeric GPVI due to avidity effects. The αC-region of fibrinogen is the primary binding site for GPVI. GPVI interaction with fibrinogen and fibrin (including non-polymerizing fibrin variant) occurs at similar levels, demonstrating GPVI binding is independent of fibrin polymerization. Fibrin polymerization into fibers clusters GPVI through the αC-region, potentially driving downstream signaling.","method":"Multiple protein-protein interaction methods (surface-based and solution-based), SPR, competition assays with fibrinogen fragments, non-polymerizing fibrin variant, dimeric vs monomeric GPVI constructs","journal":"Arteriosclerosis, thrombosis, and vascular biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — multiple orthogonal binding methods, fragment mapping, non-polymerizing fibrin variant control, avidity mechanism established","pmids":["33472402"],"is_preprint":false},{"year":2021,"finding":"Crystal structure of monomeric GPVI in complex with nanobody Nb2 revealed: (1) Nb2 binding epitope adjacent to the CRP binding groove in the D1 domain; (2) a novel domain-swapped GPVI dimer conformation via the C-C' loop hinge in D2. Truncation of the C-C' loop hinge prevents domain-swapping and abolishes GPVI signaling in a cell-based NFAT reporter assay without blocking ligand binding, establishing that the C-C' loop/domain-swapped structure is required for GPVI signaling but not ligand binding.","method":"X-ray crystallography of GPVI-Nb2 complex, C-C' loop truncation mutagenesis, NFAT luciferase reporter assay, Nb2 collagen/CRP displacement binding assay","journal":"Blood","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure plus mutagenesis plus functional reporter assay in single study, separation of ligand binding from signaling function","pmids":["33512486"],"is_preprint":false},{"year":2021,"finding":"Both CLEC-2 and GPVI (via FcRγ chain) are platelet hemin receptors. Hemin directly binds to both CLEC-2 and GPVI as shown by western blotting and SPR, activating platelets by stimulating SYK and PLCγ2 phosphorylation. Hemin-induced murine platelet aggregation is partially reduced in CLEC-2-depleted or FcRγ-deficient platelets and almost completely inhibited in CLEC-2-depleted FcRγ-deficient double-knockout platelets.","method":"SPR binding of hemin to CLEC-2 and GPVI, western blotting, CLEC-2-depleted and FcRγ-/- mouse platelets, double-knockout model, GPVI antibody JAQ-1 inhibition, platelet aggregation","journal":"Blood advances","confidence":"High","confidence_rationale":"Tier 2 / Strong — SPR binding assay plus multiple genetic models (single and double knockout), functional inhibition with antibody","pmids":["33843987"],"is_preprint":false},{"year":2022,"finding":"Anti-GPVI nanobody Nb2 blocks GPVI-mediated signaling (Syk, LAT, PLCγ2 phosphorylation) and thrombus formation on collagen and atherosclerotic plaque. Non-blocking Nb28 reveals GPVI forms distinct distribution patterns on collagen (clusters along fibers) vs. plaque; clustering on collagen fibers is abolished by Nb2. GPVI clustering is required for downstream signaling and thrombus formation. Adhesion to plaque is maintained despite GPVI signaling blockade and is not inhibited by α2β1 alone.","method":"Anti-GPVI nanobodies, western blot of Syk/LAT/PLCγ2 phosphorylation, whole blood thrombus formation under flow, super-resolution/fluorescence microscopy with labeled Nb28, integrin blocking antibodies","journal":"Journal of thrombosis and haemostasis","confidence":"High","confidence_rationale":"Tier 2 / Strong — live cell super-resolution microscopy, phosphorylation western blots, flow-based thrombus assay, multiple inhibitor combinations","pmids":["35894121"],"is_preprint":false},{"year":2023,"finding":"GPVI plays a key role in neutrophil recruitment, platelet-neutrophil complex (PNC) formation, and NETosis during acute lung injury (ALI). Anti-GPVI treatment results in less stable platelet-neutrophil interactions, reduced neutrophil crawling and adhesion on endothelial cells, reduced neutrophil transmigration and alveolar infiltrates, and strongly reduced NET formation. GPVI-deficient mice are markedly protected from pulmonary inflammation in LPS-induced ALI, without increased pulmonary bleeding.","method":"GPVI-deficient mice, anti-GPVI antibody treatment, LPS-induced ALI model, intravital confocal microscopy of ventilated lung, high-resolution multicolor microscopy of lung sections, PNC quantification, NET staining","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic knockout plus antibody depletion, intravital microscopy, multiple functional readouts (neutrophil crawling, adhesion, transmigration, NETs), in vivo model","pmids":["37441848"],"is_preprint":false},{"year":2003,"finding":"PLCγ1 partially compensates for PLCγ2 downstream of GPVI in mouse (but not human) platelets. CRP and collagen stimulate tyrosine phosphorylation of PLCγ1 at Tyr783 in mouse platelets through a Src kinase-dependent pathway. In PLCγ2-/- platelets, weak collagen-induced integrin activation persists via PLCγ1 and PI3-kinase signaling downstream of GPVI. Adhesion of PLCγ2-/- platelets to collagen under shear is severely reduced but not abolished (unlike FcRγ-/- platelets).","method":"PLCγ2-/- and FcRγ-/- mouse platelets, PLCγ1 phosphospecific antibody (pTyr783), PP1 (Src inhibitor), wortmannin (PI3K inhibitor), αIIbβ3 and α2β1 inhibitors, Born aggregometry, collagen adhesion under flow","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple genetic knockout models, phosphospecific antibodies, kinase inhibitors, flow adhesion assay","pmids":["12730118"],"is_preprint":false}],"current_model":"GPVI is a platelet and megakaryocyte immunoglobulin-superfamily receptor that exists as dimers (and higher-order clusters) on the platelet surface, associates non-covalently with the FcRγ chain, and is activated by collagen, fibrin/fibrinogen (via the αC-region), laminin, EMMPRIN, globular adiponectin, hemin, and several snake venom proteins; upon ligand-induced clustering, constitutively-active Lyn (recruited via a GPVI proline-rich domain) phosphorylates the FcRγ ITAM tyrosines, recruiting and activating Syk (regulated by c-Cbl ubiquitination and TULA-2 dephosphorylation of pTyr346), which—with co-activation of LAT, SLP-76, Gads, and Tec/Btk kinases—drives PLCγ2 activation and downstream Ca2+, PKC, Cdc42, Rac1, and Nox1/ROS-dependent platelet responses including granule secretion, integrin αIIbβ3 inside-out activation, procoagulant PS exposure, and NETosis induction; GPVI surface expression is controlled by ADAM10/ADAM17-mediated ectodomain shedding (generating soluble GPVI) or receptor internalization, both downstream of the FcRγ ITAM, and GP6 transcription during megakaryopoiesis is initiated by TPO-driven CpG demethylation of the GP6 promoter."},"narrative":{"mechanistic_narrative":"GP6 encodes GPVI, an immunoglobulin-superfamily collagen receptor expressed early during megakaryocyte differentiation and on platelets, where it functions as the principal activating receptor for collagen and related vascular ligands and as a driver of platelet adhesion, activation, and procoagulant responses [PMID:11278467, PMID:11723134]. GPVI couples to the FcRγ chain via a transmembrane arginine and intracellular C-terminal tail, and upon ligand-induced clustering, Src-family kinases phosphorylate the FcRγ ITAM tyrosines, recruiting and activating Syk and culminating in PLCγ2 activation [PMID:11152687, PMID:20345705]. Receptor avidity is achieved through dimerization and higher-order clustering: dimeric GPVI binds fibrous collagen with high affinity, while pre-existing dimers coalesce into actin cytoskeleton-dependent clusters on collagenous substrates that colocalize with phosphotyrosine signaling [PMID:12356768, PMID:28058806], and a domain-swapped dimer conformation mediated by a D2 C-C' loop hinge is structurally required for signaling independent of ligand binding [PMID:33512486]. Beyond collagen, GPVI is a signaling receptor for laminin, EMMPRIN/CD147, globular adiponectin, hemin, fibrin and the fibrinogen αC-region, and snake venom proteins such as convulxin and alborhagin, each triggering the FcRγ–Syk–PLCγ2 cascade [PMID:11344165, PMID:16219796, PMID:19350111, PMID:26282541, PMID:18419742, PMID:33472402, PMID:33843987]. The proximal signal is shaped by a constitutive GPVI proline-rich-domain interaction with Lyn [PMID:19940238], by c-Cbl-mediated ubiquitination that enhances Syk activity, and by TULA-2 dephosphorylation of Syk Tyr346 [PMID:15701717, PMID:27609517], with downstream propagation through LAT, SLP-76, Tec/Btk kinases, FAK, Cdc42, and Nox1-derived ROS to control granule secretion, integrin αIIbβ3 activation, and phosphatidylserine exposure [PMID:10567557, PMID:12842985, PMID:21789221, PMID:24494191, PMID:25415317]. GPVI surface levels are governed by ADAM10/ADAM17-mediated ectodomain shedding and by receptor internalization, both operating downstream of the FcRγ ITAM, while GP6 transcription is initiated by TPO-driven CpG demethylation of its promoter during megakaryopoiesis [PMID:15701720, PMID:17374738, PMID:20644114]. A homozygous GP6 frameshift producing a truncated protein lacking the transmembrane domain ablates surface GPVI and collagen responsiveness, causing a mild bleeding diathesis [PMID:23815599, PMID:32603422]. GPVI also extends beyond hemostasis to inflammation, promoting platelet-neutrophil complex formation and NETosis in acute lung injury [PMID:37441848].","teleology":[{"year":1997,"claim":"Established that the platelet collagen receptor p62/GPVI transduces collagen-like activation, defining a Src-kinase-to-Syk phosphorylation cascade through the FcRγ chain.","evidence":"Convulxin binding and tyrosine-phosphorylation cascade mapping in platelet lysates with kinase inhibitor","pmids":["9153205"],"confidence":"High","gaps":["Did not establish the receptor's quaternary state","Direct collagen-binding site not yet mapped"]},{"year":2001,"claim":"Defined the structural requirements for GPVI–FcRγ coupling and demonstrated that GPVI density determines collagen adhesion and signaling, while showing GPVI is functional from early megakaryopoiesis.","evidence":"Reconstitution in RBL-2H3/K562/Jurkat cells with transmembrane and C-tail mutagenesis, density-titrated cell lines, and megakaryocyte differentiation assays","pmids":["11152687","11278467","11344165","11723134"],"confidence":"High","gaps":["Whether GPVI signals as monomer or dimer unresolved","Relationship to α2β1 in collagen binding not yet defined"]},{"year":2002,"claim":"Resolved that GPVI dimerization, not the monomer, confers high-affinity collagen binding and that the convulxin and collagen sites are distinct, establishing avidity as central to receptor function.","evidence":"SPR kinetics and aggregation-inhibition assays comparing monomeric and dimeric recombinant GPVI ectodomains","pmids":["12356768"],"confidence":"High","gaps":["Dimer existence on native platelets inferred, not yet imaged","Structural basis of dimerization unknown"]},{"year":2005,"claim":"Built the downstream signaling map by ordering LAT, SLP-76, Tec/Btk, and PLCγ1/γ2 contributions and defining differential adaptor requirements relative to TCR signaling.","evidence":"Genetic knockout mouse platelets (LAT, SLP-76, Syk, Btk/Tec, PLCγ2, FcRγ) with epistasis and functional readouts; XLA human platelets","pmids":["10567557","11901197","12842985","12730118"],"confidence":"High","gaps":["Species differences in PLCγ1 compensation not mechanistically explained","Quantitative contribution of minor LAT-independent branch unclear"]},{"year":2005,"claim":"Identified post-translational control points of proximal signaling — c-Cbl-dependent Syk ubiquitination enhancing kinase activity — and the cooperative role of α2β1 in increasing collagen avidity.","evidence":"c-Cbl-/- platelets with ubiquitin-IP and in vitro kinase assays; pharmacological α2β1/ADP/COX dissection under flow","pmids":["15701717","12871331","15886326"],"confidence":"High","gaps":["Mechanism by which ubiquitination raises Syk activity not fully resolved","α2β1 avidity role established largely pharmacologically"]},{"year":2005,"claim":"Connected GPVI transcriptional control and ligand repertoire expansion, showing TPO-driven promoter demethylation drives GP6 expression and GPVI serves as a laminin receptor.","evidence":"Bisulfite sequencing with TPO treatment; GPVI-/- and Syk-/- platelets, α6 blockade, and SPR for laminin binding","pmids":["15701720","16219796"],"confidence":"High","gaps":["Transcription factors acting on the demethylated promoter not identified","Laminin-binding site on GPVI not mapped"]},{"year":2007,"claim":"Defined the regulation of GPVI surface expression, distinguishing ADAM-mediated ectodomain shedding from internalization, both downstream of the FcRγ ITAM.","evidence":"MALDI-TOF-MS peptide cleavage, ADAM10/ADAM17 enzyme assays, and FcRγ ITAM point-mutant, LAT-/-, and PLCγ2-/- mice with flow cytometry","pmids":["17445093","17374738"],"confidence":"High","gaps":["In vivo dominant sheddase not yet pinned down in this work","Signals rendering GPVI susceptible to cleavage incompletely defined"]},{"year":2009,"claim":"Characterized the proximal signaling architecture — constitutive Lyn priming via the GPVI proline-rich domain, tetraspanin microdomain localization, and Lyn/PKCδ/SHIP-1 negative regulation of secretion.","evidence":"Reciprocal Co-IP, PRD-deletion mutants, Lyn-/-/SHIP-1-/-/PKCδ-/- platelets, and Tspan9 Co-IP/colocalization","pmids":["19940238","18795891","19587372"],"confidence":"High","gaps":["Tetraspanin microdomain functional consequence rests on single-lab Co-IP","How Lyn-priming integrates with negative SHIP-1 regulation temporally unclear"]},{"year":2009,"claim":"Expanded the GPVI ligand repertoire to EMMPRIN, globular adiponectin, and an alborhagin venom protein, and informed collagen recognition through the homologous LAIR-1 structure.","evidence":"SPR/ELISA binding with transfected CHO/Jurkat reporters, FcRγ-/- platelets, and LAIR-1 crystallography with NMR and mutagenesis","pmids":["19350111","18419742","20007810","20040043"],"confidence":"High","gaps":["Physiological relevance of EMMPRIN and adiponectin engagement in vivo not established","Conserved collagen-contact residues inferred from a homolog, not GPVI structure"]},{"year":2014,"claim":"Extended the downstream effector network to Cdc42, FAK, Nox1-derived ROS, and identified CEACAM2 as an ITIM-bearing negative regulator of GPVI signaling.","evidence":"Inducible Cdc42 knockout, Pyk2-/- platelets with FAK inhibitor, Nox2-/- with Nox1 inhibitor, and Ceacam2-/- platelets with in vivo thrombosis models","pmids":["21789221","25415317","24494191","25085348"],"confidence":"High","gaps":["FAK pathway position rests partly on inhibitor epistasis from a single lab","Mechanism of CEACAM2 recruitment to GPVI not detailed"]},{"year":2017,"claim":"Established fibrin and fibrinogen as GPVI ligands and resolved the spatial mechanism of activation as actin-dependent clustering of pre-existing dimers.","evidence":"GPVI-/- mouse and GPVI-deficient human platelets, SPR fragment mapping (D-dimer, αC-region), and dSTORM/TIRF imaging of dimer clusters","pmids":["26282541","29296791","28058806"],"confidence":"High","gaps":["Differential monomer vs dimer preference for fibrin vs collagen mechanistically incomplete","Link between cluster geometry and ITAM phosphorylation kinetics not quantified"]},{"year":2020,"claim":"Confirmed the physiological consequence of GPVI loss in humans and mapped the genome-wide phosphosignaling output including a TAK1-Rab7 endolysosomal branch.","evidence":"Homozygous GP6 loss-of-function human platelets under flow; TMT/SPS-MS3 phosphoproteomics with TAK1 inhibitor validation","pmids":["32603422","23815599","32640021"],"confidence":"High","gaps":["Many phosphoproteomic relations remain individually unvalidated","Mechanism preserving partial adhesion despite GPVI loss not fully defined"]},{"year":2021,"claim":"Provided the structural and biophysical basis for GPVI signaling competence, defining the domain-swapped dimer via the D2 C-C' loop and the fibrinogen αC-region/avidity interaction.","evidence":"GPVI-Nb2 crystallography with C-C' loop truncation and NFAT reporter; multi-method binding with fibrinogen fragments, non-polymerizing fibrin variant, and hemin SPR","pmids":["33512486","33472402","33843987"],"confidence":"High","gaps":["How domain-swapping is triggered by ligand clustering in the membrane unclear","Physiological role of hemin engagement in vivo not established"]},{"year":2023,"claim":"Demonstrated GPVI function beyond hemostasis, establishing its role in platelet-neutrophil complex formation and NETosis driving inflammatory injury.","evidence":"GPVI-deficient mice and anti-GPVI antibody in LPS-induced acute lung injury with intravital microscopy and NET quantification","pmids":["37441848"],"confidence":"High","gaps":["GPVI ligand mediating neutrophil interactions in inflamed lung not identified","Whether the same FcRγ/Syk cascade drives the NETosis phenotype not dissected"]},{"year":null,"claim":"The identity of the third in vivo GPVI-cleaving sheddase and the membrane-level trigger converting ligand clustering into domain-swapped, signaling-competent dimers remain open.","evidence":"","pmids":[],"confidence":"Medium","gaps":["Third platelet GPVI sheddase beyond ADAM10/ADAM17 unidentified","Coupling between cluster geometry and the D2 domain-swap conformational change unresolved"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0098631","term_label":"cell adhesion mediator activity","supporting_discovery_ids":[4,5,11,27]},{"term_id":"GO:0060089","term_label":"molecular transducer activity","supporting_discovery_ids":[1,24,32]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[19,30]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[1,4,20,30]}],"pathway":[{"term_id":"R-HSA-109582","term_label":"Hemostasis","supporting_discovery_ids":[4,12,27,38]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[1,6,24]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[43]}],"complexes":["GPVI-FcRγ receptor complex","tetraspanin (Tspan9) microdomain"],"partners":["FCER1G","LYN","SYK","PLCG2","LAT","TSPAN9","CBL","ITGA6"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9HCN6","full_name":"Platelet glycoprotein VI","aliases":["Glycoprotein 6"],"length_aa":339,"mass_kda":36.9,"function":"Collagen receptor involved in collagen-induced platelet adhesion and activation. Plays a key role in platelet procoagulant activity and subsequent thrombin and fibrin formation. This procoagulant function may contribute to arterial and venous thrombus formation. The signaling pathway involves the FcR gamma-chain, the Src kinases (likely FYN or LYN) and SYK, the adapter protein LAT and leads to the activation of PLCG2","subcellular_location":"Cell membrane","url":"https://www.uniprot.org/uniprotkb/Q9HCN6/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/GP6","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/GP6","total_profiled":1310},"omim":[{"mim_id":"620484","title":"THROMBOCYTOPENIA 10; THC10","url":"https://www.omim.org/entry/620484"},{"mim_id":"617443","title":"BLEEDING DISORDER, PLATELET-TYPE, 21; BDPLT21","url":"https://www.omim.org/entry/617443"},{"mim_id":"614201","title":"BLEEDING DISORDER, PLATELET-TYPE, 11; BDPLT11","url":"https://www.omim.org/entry/614201"},{"mim_id":"605546","title":"GLYCOPROTEIN VI, PLATELET; GP6","url":"https://www.omim.org/entry/605546"},{"mim_id":"600925","title":"PROTEIN-TYROSINE PHOSPHATASE, RECEPTOR-TYPE, J; PTPRJ","url":"https://www.omim.org/entry/600925"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Plasma membrane","reliability":"Supported"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in some","driving_tissues":[{"tissue":"brain","ntpm":2.3},{"tissue":"testis","ntpm":2.5}],"url":"https://www.proteinatlas.org/search/GP6"},"hgnc":{"alias_symbol":["GPVI"],"prev_symbol":[]},"alphafold":{"accession":"Q9HCN6","domains":[{"cath_id":"2.60.40.10","chopping":"30-108","consensus_level":"high","plddt":95.988,"start":30,"end":108},{"cath_id":"2.60.40.10","chopping":"113-204","consensus_level":"high","plddt":93.3032,"start":113,"end":204}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9HCN6","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9HCN6-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9HCN6-F1-predicted_aligned_error_v6.png","plddt_mean":76.06},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=GP6","jax_strain_url":"https://www.jax.org/strain/search?query=GP6"},"sequence":{"accession":"Q9HCN6","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9HCN6.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9HCN6/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9HCN6"}},"corpus_meta":[{"pmid":"9049358","id":"PMC_9049358","title":"The use of general primers GP5 and GP6 elongated at their 3' ends with adjacent highly conserved sequences improves human papillomavirus detection by PCR.","date":"1995","source":"The Journal of general virology","url":"https://pubmed.ncbi.nlm.nih.gov/9049358","citation_count":1073,"is_preprint":false},{"pmid":"12649139","id":"PMC_12649139","title":"Platelet-collagen interaction: is GPVI the central receptor?","date":"2003","source":"Blood","url":"https://pubmed.ncbi.nlm.nih.gov/12649139","citation_count":880,"is_preprint":false},{"pmid":"9041439","id":"PMC_9041439","title":"A general primer GP5+/GP6(+)-mediated PCR-enzyme immunoassay method for rapid detection of 14 high-risk and 6 low-risk human papillomavirus genotypes in cervical scrapings.","date":"1997","source":"Journal of clinical microbiology","url":"https://pubmed.ncbi.nlm.nih.gov/9041439","citation_count":484,"is_preprint":false},{"pmid":"9163434","id":"PMC_9163434","title":"PCR detection of human papillomavirus: comparison between MY09/MY11 and GP5+/GP6+ primer systems.","date":"1997","source":"Journal of clinical microbiology","url":"https://pubmed.ncbi.nlm.nih.gov/9163434","citation_count":378,"is_preprint":false},{"pmid":"16102042","id":"PMC_16102042","title":"GPVI and integrin alphaIIb beta3 signaling in platelets.","date":"2005","source":"Journal of thrombosis and haemostasis : JTH","url":"https://pubmed.ncbi.nlm.nih.gov/16102042","citation_count":323,"is_preprint":false},{"pmid":"9153205","id":"PMC_9153205","title":"Platelet activation and signal transduction by convulxin, a C-type lectin from Crotalus durissus terrificus (tropical rattlesnake) venom via the p62/GPVI collagen receptor.","date":"1997","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/9153205","citation_count":299,"is_preprint":false},{"pmid":"30601137","id":"PMC_30601137","title":"Functional significance of the platelet immune receptors GPVI and CLEC-2.","date":"2019","source":"The Journal of clinical investigation","url":"https://pubmed.ncbi.nlm.nih.gov/30601137","citation_count":251,"is_preprint":false},{"pmid":"17445093","id":"PMC_17445093","title":"Controlled shedding of platelet glycoprotein (GP)VI and GPIb-IX-V by ADAM family metalloproteinases.","date":"2007","source":"Journal of thrombosis and haemostasis : JTH","url":"https://pubmed.ncbi.nlm.nih.gov/17445093","citation_count":218,"is_preprint":false},{"pmid":"26282541","id":"PMC_26282541","title":"Fibrin activates GPVI in human and mouse platelets.","date":"2015","source":"Blood","url":"https://pubmed.ncbi.nlm.nih.gov/26282541","citation_count":178,"is_preprint":false},{"pmid":"18199790","id":"PMC_18199790","title":"Homogeneous amplification of genital human alpha papillomaviruses by PCR using novel broad-spectrum GP5+ and GP6+ primers.","date":"2008","source":"Journal of clinical microbiology","url":"https://pubmed.ncbi.nlm.nih.gov/18199790","citation_count":161,"is_preprint":false},{"pmid":"10567557","id":"PMC_10567557","title":"LAT is required for tyrosine phosphorylation of phospholipase cgamma2 and platelet activation by the collagen receptor GPVI.","date":"1999","source":"Molecular and cellular biology","url":"https://pubmed.ncbi.nlm.nih.gov/10567557","citation_count":153,"is_preprint":false},{"pmid":"20345705","id":"PMC_20345705","title":"GPVI and CLEC-2 in hemostasis and vascular integrity.","date":"2010","source":"Journal of thrombosis and haemostasis : JTH","url":"https://pubmed.ncbi.nlm.nih.gov/20345705","citation_count":150,"is_preprint":false},{"pmid":"12356768","id":"PMC_12356768","title":"Analysis of the interaction of platelet collagen receptor glycoprotein VI (GPVI) with collagen. 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dimer.","date":"2021","source":"Blood","url":"https://pubmed.ncbi.nlm.nih.gov/33512486","citation_count":26,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":55028,"output_tokens":13667,"usd":0.185045,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":25924,"output_tokens":5137,"usd":0.129023,"stage2_stop_reason":"end_turn"},"total_usd":0.314068,"stage1_batch_id":"msgbatch_01DUPACZi7AAWwuZiUjseQF4","stage2_batch_id":"msgbatch_018yt7chE3f48PEFtDwtj6AE","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1997,\n      \"finding\": \"Convulxin, a C-type lectin from snake venom, activates platelets through the p62/GPVI collagen receptor (not GPIb), inducing tyrosine phosphorylation of FcRγ chain, PLCγ2, Syk (p72), c-Cbl, and p36-38 in a pattern similar to collagen. Convulxin binds directly to GPVI immunoprecipitated from platelet lysates, and convulxin subunits inhibit both aggregation and tyrosine phosphorylation in response to collagen. Src family kinases phosphorylate FcRγ chain upon GPVI clustering, activating Syk downstream.\",\n      \"method\": \"125I-convulxin binding, immunoprecipitation, tyrosine phosphorylation western blot, platelet aggregation assay, tyrosine kinase inhibitor (piceatannol)\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal binding assay, Co-IP, phosphorylation cascade mapping, inhibitor studies, replicated across multiple methods in single study\",\n      \"pmids\": [\"9153205\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"GPVI expression in RBL-2H3 cells (which express FcRγ chain) confers adhesive and signaling responses to convulxin via the Fc receptor γ-chain. Both the transmembrane arginine of GPVI and its intracellular C-terminal tail are required for coupling to FcRγ and signal transduction. GPVI expression was sufficient for convulxin responses but not for full collagen signaling, suggesting an additional collagen-binding receptor is required.\",\n      \"method\": \"Transfection of wild-type and mutant GPVI into RBL-2H3 cells, adhesion assays, signaling readouts, mutagenesis of transmembrane domain and C-tail\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — direct mutagenesis of functional domains with multiple orthogonal readouts (adhesion, signaling), reconstitution in heterologous cells\",\n      \"pmids\": [\"11152687\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"GPVI is expressed early during megakaryocyte differentiation (CD41+ stage) and associates with the FcRγ chain in megakaryocytes as in platelets. Functional GPVI-mediated signaling (tyrosine phosphorylation of Syk, FcRγ, PLCγ2) is activated by convulxin in mature megakaryocytes; adhesion to immobilized convulxin occurs only at late stages of maturation when GPVI expression increases.\",\n      \"method\": \"Flow cytometry, RT-PCR, immunoblotting, ligand blotting, adhesion assays, tyrosine phosphorylation analysis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (flow cytometry, biochemistry, functional adhesion), clear functional-expression correlation\",\n      \"pmids\": [\"11278467\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"Alborhagin, a viper venom metalloproteinase, activates platelets through GPVI (not through a distinct receptor), inducing a tyrosine phosphorylation pattern similar to convulxin including PLCγ2 phosphorylation. Alborhagin-induced aggregation in mouse platelets is inhibited by anti-GPVI monoclonal antibody JAQ1, and alborhagin induces GPVI-dependent responses in GPVI-transfected K562 and Jurkat cells.\",\n      \"method\": \"Platelet aggregation, tyrosine phosphorylation western blot, transfected cell signaling, antibody inhibition (JAQ1), Src kinase inhibitor PP1\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — GPVI-transfected cell confirmation, antibody blockade, multiple orthogonal readouts\",\n      \"pmids\": [\"11344165\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"GPVI mediates both adhesive and signaling responses to collagen in a receptor density-dependent manner. GPVI-FcRγ expression alone is sufficient to confer collagen adhesion and signaling in RBL-2H3 cells when GPVI density matches that on human platelets; collagen responses are proportional to receptor density. GPVI receptor density on human platelets is tightly regulated and independent of α2β1 expression.\",\n      \"method\": \"Novel anti-GPVI monoclonal antibody quantification, GPVI-expressing RBL-2H3 cell lines at varying receptor densities, adhesion and signaling assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — reconstitution in heterologous cells with defined receptor densities, multiple orthogonal readouts, resolves conflicting prior data\",\n      \"pmids\": [\"11723134\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Dimeric GPVI (GPVI-Fc fusion) but not monomeric GPVI (GPVIex) exhibits high affinity binding to fibrous collagen (KD ~576 nM by SPR), inhibits collagen-induced platelet aggregation, and inhibits collagen-related peptide-induced aggregation. Monomeric GPVI inhibited convulxin- but not collagen-induced aggregation. The convulxin binding site on GPVI is distinct from the collagen-binding site. GPVI is proposed to exist as a dimeric form on platelets for high-affinity collagen interaction.\",\n      \"method\": \"ELISA, surface plasmon resonance (SPR), platelet aggregation inhibition assays with soluble monomeric and dimeric recombinant GPVI ectodomains\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro binding reconstitution with SPR kinetics, multiple functional assays, rigorous controls distinguishing monomer vs. dimer\",\n      \"pmids\": [\"12356768\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"LAT (linker for activation of T cells) is required for tyrosine phosphorylation of PLCγ2 downstream of the GPVI collagen receptor. LAT is tyrosine-phosphorylated downstream of Syk (not SLP-76 or Btk) following GPVI activation. In LAT-deficient platelets, PLCγ2 phosphorylation is substantially reduced, leading to marked inhibition of phosphatidic acid formation, pleckstrin phosphorylation, P-selectin expression, and reduced αIIbβ3 activation in response to CRP. A minor LAT-independent pathway of PLCγ2 phosphorylation exists.\",\n      \"method\": \"LAT-deficient mouse platelets, Syk-deficient mouse platelets, XLA human platelets (Btk-deficient), SLP-76-deficient mouse platelets, tyrosine phosphorylation western blot, metabolite measurement, flow cytometry\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic knockout models with multiple orthogonal biochemical and functional readouts, epistasis established\",\n      \"pmids\": [\"10567557\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"SLP-76 plays a greater role than LAT in platelet activation downstream of GPVI. LAT-deficient platelets aggregate normally to high concentrations of collagen and convulxin, while SLP-76-deficient platelets do not. LAT is not tyrosine-phosphorylated after fibrinogen binding to integrin αIIbβ3; collagen-stimulated LAT-deficient platelets spread normally on fibrinogen. These findings establish differential requirements for LAT and SLP-76 in GPVI versus TCR signaling.\",\n      \"method\": \"LAT-/- and SLP-76-/- mouse platelets, platelet aggregation, P-selectin expression, spreading assays, phosphorylation western blot\",\n      \"journal\": \"The Journal of experimental medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — comparative genetic knockout models, multiple orthogonal functional readouts, epistasis between adaptor proteins\",\n      \"pmids\": [\"11901197\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"Tec family kinase Tec compensates for Btk downstream of GPVI in regulating PLCγ2 phosphorylation. Btk/Tec double-deficient platelets fail to undergo Ca2+ increase, aggregation, secretion, and spreading in response to collagen or CRP, while single knockouts show partial defects. A residual GPVI signal persists in Btk/Tec double-deficient platelets as CRP synergizes with ADP to mediate aggregation.\",\n      \"method\": \"Btk-/-, Tec-/-, and Btk/Tec double-knockout mouse platelets, aggregation, Ca2+ measurement, secretion, spreading, PLCγ2 phosphorylation western blot\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — double-knockout epistasis, multiple functional readouts, establishes Tec as redundant kinase for PLCγ2 activation downstream of GPVI\",\n      \"pmids\": [\"12842985\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"Activation of GPVI by collagen is regulated by α2β1 integrin and secondary mediators (ADP, thromboxanes). α2β1 blockade has a greater inhibitory effect on collagen-induced PLC activation, Ca2+ elevation, and dense granule secretion than ADP receptor antagonists plus COX inhibitor. The major role of α2β1 is proposed to increase avidity of collagen for the platelet surface, thereby enhancing GPVI activation. Secondary mediators indirectly regulate GPVI signaling via activation of α2β1.\",\n      \"method\": \"Platelet aggregation, tyrosine phosphorylation, Ca2+ measurement, dense granule secretion with pharmacological inhibitors of α2β1, ADP receptors, and COX; integrin αIIbβ3 antagonist to isolate signaling from aggregation\",\n      \"journal\": \"Journal of thrombosis and haemostasis\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — pharmacological approach with multiple signaling readouts, single lab\",\n      \"pmids\": [\"12871331\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"GPVI undergoes metalloproteinase-dependent ectodomain shedding generating a ~55 kDa soluble fragment released from platelets. This shedding is induced by mitochondrial injury (CCCP) and is inhibited by the broad-spectrum metalloproteinase inhibitor GM6001. CRP or thrombin stimulation induces shedding of GPIbα but not GPVI, suggesting different metalloproteinases regulate the two receptors or an additional signal is needed to render GPVI susceptible to cleavage.\",\n      \"method\": \"Mouse platelet stimulation with CCCP, GM6001 inhibition, immunoblotting of supernatant and lysate, platelet aggregation and CRP/thrombin response assays\",\n      \"journal\": \"Thrombosis and haemostasis\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct biochemical detection of ectodomain in supernatant, metalloproteinase inhibitor, functional readouts, single lab\",\n      \"pmids\": [\"15116256\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"GPVI is a receptor for laminin. Lamellipodia (but not filopodia) formation on laminin requires GPVI, as demonstrated in GPVI-knockout mouse platelets. Integrin α6β1 is essential for adhesion to laminin and brings laminin to GPVI. GPVI-laminin interaction was confirmed by surface plasmon resonance spectroscopy and by lamellipodia formation on laminin in the presence of collagenase. Syk kinase is required for signaling downstream of both GPVI and α6β1.\",\n      \"method\": \"GPVI-/- mouse platelets, α6-blocking antibody, Syk-/- mouse platelets, surface plasmon resonance, collagenase treatment, platelet spreading microscopy\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — SPR binding plus genetic knockout functional validation, collagenase control, multiple orthogonal approaches in single study\",\n      \"pmids\": [\"16219796\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Both GPVI and integrin α2β1 play independent critical roles in platelet adhesion to collagen under flow; loss of both receptors completely ablates this response. Intracellular signaling mediated by SLP-76 (but not LAT) is critical for platelet adhesion to collagen under flow. Reduced GPVI receptor density results in severe defects in platelet adhesion to collagen under flow.\",\n      \"method\": \"Pharmacological and genetic approaches with human and mouse platelets under flow conditions; SLP-76-/- and LAT-/- mice; GPVI density variation; tail bleeding times\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic knockout plus pharmacological inhibition, flow assays, epistasis between adaptor proteins and receptor density\",\n      \"pmids\": [\"15886326\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Syk undergoes rapid ubiquitination upon GPVI activation by collagen, CRP, and convulxin; this process is downstream of Src family kinases. Ubiquitinated Syk is ~5-fold more kinase-active than non-ubiquitinated Syk and represents 60-75% of active (pTyr525/526) Syk. In c-Cbl-deficient mice, Syk is not ubiquitinated and fails to be dephosphorylated, implicating c-Cbl as the E3 ubiquitin ligase for Syk in GPVI signaling.\",\n      \"method\": \"c-Cbl-/- mouse platelets, Src kinase inhibitor, ubiquitin immunoprecipitation, in vitro kinase assay on separated ubiquitinated/non-ubiquitinated Syk, phosphospecific antibody to Syk\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic knockout (c-Cbl-/-), in vitro kinase assay, multiple orthogonal methods identifying c-Cbl as E3 ligase\",\n      \"pmids\": [\"15701717\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Thrombopoietin (TPO) initiates CpG demethylation of the GP6 promoter, which drives GPVI transcription during megakaryocyte differentiation. The GP6 promoter CpG island is fully methylated in GPVI-non-expressive cell lines and completely unmethylated in GPVI-expressive lines. TPO treatment of UT-7/EPO-Mpl cells induces GP6 promoter demethylation correlated with increased mRNA.\",\n      \"method\": \"Sodium bisulfite genomic sequencing of CpG island, RT-PCR for mRNA quantification, TPO treatment of primary cord blood cells and cell lines\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct bisulfite sequencing of promoter methylation, multiple cell lines and primary cells, TPO treatment experiments\",\n      \"pmids\": [\"15701720\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"Collagen type I-induced stress fiber formation in megakaryocytes requires α2β1 integrin (essential) and GPVI (triggers rapid and sustained ERK1/2 MAPK activation). Both pathways converge via synergistic MAPK/ERK1/2 and Rho/ROCK activation. After longer adhesion, proplatelet formation is inhibited by α2β1 but not by GPVI, through the Rho/ROCK pathway. This demonstrates differential regulation of megakaryocyte actin dynamics by the two collagen receptors.\",\n      \"method\": \"Function-blocking antibodies against α2β1 and GPVI-specific ligands in primary human megakaryocytes, kinase inhibitors (ROCK, ERK1/2), F-actin staining, proplatelet formation assay\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — receptor-specific ligands and blocking antibodies, kinase inhibitors, multiple functional readouts, primary human cells\",\n      \"pmids\": [\"15265786\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"ADAM10, but not ADAM17, cleaves a GPVI-based synthetic peptide within the extracellular membrane-proximal sequence (PAR;Q243YY) as identified by MALDI-TOF-MS. GPVI and GPIbα are shed by distinct ADAM family members (ADAM10 for GPVI, ADAM17 for GPIbα) in response to distinct stimuli (CCCP, PMA, W7, NEM). A significant fraction of GPIbα but not GPVI exists in cleaved state on resting platelets.\",\n      \"method\": \"Immunoblotting with affinity-purified cytoplasmic domain antibodies, MALDI-TOF-MS of synthetic peptide cleavage products, ADAM10/ADAM17 recombinant enzyme assays, metalloproteinase inhibitors, mutagenesis of cleavage sequences in transfected cells\",\n      \"journal\": \"Journal of thrombosis and haemostasis\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro peptide cleavage with mass spectrometry, mutagenesis validation, biochemical fractionation\",\n      \"pmids\": [\"17445093\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"GPVI down-regulation occurs through two distinct pathways downstream of the FcRγ-ITAM: ectodomain shedding (via metalloproteinases, requiring LAT and PLCγ2 signaling) or internalization/intracellular clearing (independent of shedding). Both pathways are abrogated in mice with a point mutation in the FcRγ ITAM. In LAT-/- or PLCγ2-/- mice, GPVI shedding is abolished but receptor is down-regulated through internalization without thrombocytopenia or altered thrombin responses.\",\n      \"method\": \"FcRγ ITAM point mutant mice, LAT-/- mice, PLCγ2-/- mice, anti-GPVI antibody (JAQ1) treatment, flow cytometry of GPVI surface levels, platelet count monitoring, thrombin response assays\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple genetic models (point mutant + knockouts), epistasis establishing two distinct signaling branches, multiple functional readouts\",\n      \"pmids\": [\"17374738\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"GPVI ectodomain shedding in vitro can be mediated by either ADAM10 or ADAM17 in response to distinct stimuli. In vivo antibody (JAQ1)-induced GPVI shedding occurs even in mice lacking both ADAM10 and ADAM17 in platelets, establishing the existence of a third GPVI-cleaving platelet enzyme.\",\n      \"method\": \"Megakaryocyte-specific ADAM10-deficient mice, Adam17ex/ex mice, ADAM10/ADAM17 double-deficient platelets in vitro and in vivo, anti-GPVI antibody JAQ1 treatment, flow cytometry\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — two genetic models plus double deficiency, in vitro and in vivo shedding experiments\",\n      \"pmids\": [\"20644114\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"GPVI constitutively associates in resting platelets with the Src-family kinase Lyn via a unique intracellular proline-rich domain (PRD), which directly activates Lyn presumably through SH3 displacement. Lyn is bound to GPVI in an activated state. Platelets lacking Lyn exhibit defective collagen adhesion under flow similar to platelets with GPVI receptors lacking the PRD. This PRD-Lyn interaction constitutes a molecular priming mechanism enabling rapid hemostatic response.\",\n      \"method\": \"Co-immunoprecipitation of Lyn with GPVI, PRD deletion mutant GPVI, Lyn-/- mouse platelets, platelet adhesion to collagen under flow, in vitro binding assays\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, PRD mutant analysis, Lyn-/- genetic model, flow adhesion assay, multiple orthogonal methods\",\n      \"pmids\": [\"19940238\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"GPVI is a component of tetraspanin microdomains on platelets, specifically associating with Tspan9 and integrin α6β1, but not with GPIbα or integrins αIIbβ3 or α2β1. This selective tetraspanin microdomain association was established by Co-IP and co-localization.\",\n      \"method\": \"Co-immunoprecipitation, western blotting, flow cytometry with novel Tspan9 antibodies, megakaryocyte SAGE and DNA microarrays\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Co-IP evidence of GPVI in specific tetraspanin microdomain, single lab\",\n      \"pmids\": [\"18795891\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"GPVI functions as a receptor for EMMPRIN (CD147/basigin) and mediates platelet rolling on immobilized EMMPRIN under arterial shear. Anti-GPVI blocking antibodies reduce platelet rolling on EMMPRIN-Fc. CHO cells transfected with GPVI show enhanced rolling on EMMPRIN-Fc; CHO cells transfected with EMMPRIN show enhanced rolling on GPVI-Fc. Direct GPVI-EMMPRIN binding was confirmed by ELISA and SPR with KD = 88 nM.\",\n      \"method\": \"Flow-based platelet rolling assay, blocking monoclonal antibodies (anti-EMMPRIN, anti-GPVI), CHO cell transfection, modified ELISA, surface plasmon resonance\",\n      \"journal\": \"Thrombosis and haemostasis\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — SPR with KD measurement, transfected cell assays, blocking antibodies, multiple orthogonal methods\",\n      \"pmids\": [\"19350111\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Lyn, PKC-δ, and SHIP-1 form a complex that specifically regulates GPVI-mediated (but not PAR-mediated) dense granule secretion. GPVI stimulation induces rapid SHIP-1 phosphorylation at Y1020 and association of SHIP-1 and Lyn with PKC-δ. In Lyn-/- platelets, GPVI-mediated phosphorylations of SHIP-1 (Y1020) and PKC-δ (Y311) are inhibited. In Lyn-/- or SHIP-1-/- platelets, GPVI-mediated dense granule secretion is potentiated (negative regulation), while PAR-mediated secretion is inhibited. Lyn-mediated phosphorylation of PKC-δ and SHIP-1 negatively regulates GPVI-specific dense granule secretion.\",\n      \"method\": \"PKC-δ-/-, Lyn-/-, SHIP-1-/- mouse platelets, Co-IP, phosphospecific antibodies, dense granule secretion assays (convulxin vs PAR agonists)\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple genetic knockout models, reciprocal Co-IP, phosphospecific antibodies, comparing GPVI vs. PAR pathways\",\n      \"pmids\": [\"19587372\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"The crystal structure of LAIR-1 (a collagen receptor homologous to GPVI's collagen-binding domain) was determined. NMR titrations and mutagenesis mapped the collagen-binding site: residues R59, E61, and W109 are key for collagen interaction. These residues are strictly conserved in both LAIR-1 and GPVI but are located outside the previously proposed GPVI collagen-binding site, revealing an unanticipated common mechanism of collagen recognition.\",\n      \"method\": \"X-ray crystallography (LAIR-1 structure), NMR titrations with collagen peptides, mutagenesis of collagen-contact residues\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure plus NMR plus mutagenesis, directly informing GPVI collagen-binding mechanism through conserved residues\",\n      \"pmids\": [\"20007810\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"GPVI signals through an ITAM in the associated FcRγ chain via sequential Src family kinase-mediated phosphorylation of the two ITAM tyrosines, followed by Syk SH2 domain binding and activation, culminating in PLCγ2 activation. This mechanistic pathway was established and compared to CLEC-2 signaling (which uses a hemITAM).\",\n      \"method\": \"Review consolidating genetic knockout and biochemical data from multiple studies; mechanistic pathway established by epistasis and phosphorylation cascade analysis\",\n      \"journal\": \"Journal of thrombosis and haemostasis\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — review synthesizing genetic and biochemical data; pathway mechanistically established across multiple prior experiments\",\n      \"pmids\": [\"20345705\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Cdc42 GTPase is required for GPVI-dependent (CRP-induced) filopodia formation, secretion (ATP and P-selectin), and aggregation in platelets. Cdc42-/- platelets show diminished PAK1/2 phosphorylation (Cdc42 effector) and minimal Akt phosphorylation upon CRP stimulation, establishing Cdc42 as a downstream effector in GPVI signaling.\",\n      \"method\": \"Mx-cre;Cdc42lox/lox inducible knockout mice, CRP and thrombin stimulation, platelet aggregation, P-selectin and ATP release, spreading assay, PAK1/2 phosphorylation, Akt phosphorylation, bleeding time\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — inducible genetic knockout, multiple functional readouts (aggregation, secretion, spreading), pathway marker phosphorylation\",\n      \"pmids\": [\"21789221\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"An adenine insertion (c.711_712insA) in exon 6 of GP6 generates a premature stop codon, producing a truncated GPVI protein (~49 kDa) that lacks the transmembrane domain and is absent from the platelet surface. Platelets from homozygous patients have absent surface GPVI, no aggregation in response to collagen, convulxin, or CRP, and no 14C-5-HT secretion with these agonists, directly establishing that the transmembrane domain is required for functional surface expression.\",\n      \"method\": \"DNA sequencing, mRNA sequencing, western blotting, flow cytometry, immunofluorescence confocal microscopy, platelet aggregation, 14C-5-HT secretion\",\n      \"journal\": \"Journal of thrombosis and haemostasis\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — natural loss-of-function mutation with molecular characterization, multiple orthogonal functional assays confirming complete loss of GPVI signaling\",\n      \"pmids\": [\"23815599\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"GPVI is a signaling receptor for fibrin. Thrombin-induced fibrin generation activates GPVI-dependent tyrosine phosphorylation of FcRγ and Syk in human and mouse platelets; this is abolished in GPVI-deficient mouse platelets. Mouse platelets spread fully on fibrin (but not fibrinogen) in a GPVI- and Src kinase-dependent manner, associated with phosphatidylserine exposure. The GPVI ectodomain binds directly to immobilized monomeric and polymerized fibrin.\",\n      \"method\": \"GPVI-/- mouse platelets, eptifibatide (αIIbβ3 blocker), PAR4 peptide controls, Syk and FcRγ phosphorylation western blot, platelet spreading on fibrin/fibrinogen, Src kinase inhibitor, SPR-like binding assay\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — GPVI-/- genetic model, direct ectodomain binding to fibrin, multiple functional readouts, controls distinguishing fibrin from fibrinogen\",\n      \"pmids\": [\"26282541\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"GPVI-mediated inhibition of proplatelet formation in megakaryocytes is specifically controlled by collagen type I signaling through receptor-proximal Src family kinases; Syk and LAT are dispensable for this pathway. Integrin α2β1 mediates megakaryocyte binding to collagens. Collagen type IV at the vascular niche may displace collagen I from megakaryocytes, preventing GPVI-mediated inhibition.\",\n      \"method\": \"Megakaryocytes from GPVI-/-, Syk-/-, LAT-/-, and α2β1-/- mice; blocking antibodies; Src family kinase inhibitors; adhesion assays; proplatelet formation assays\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple genetic knockout models, receptor-blocking antibodies, kinase inhibitors, well-defined functional readout (PPF)\",\n      \"pmids\": [\"27505889\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"TULA-2 protein phosphatase negatively regulates Syk activation downstream of GPVI by dephosphorylating Tyr346, a regulatory (interdomain B) site of Syk that is phosphorylated early after receptor ligation and is critical for initiating full Syk activation. TULA-2 dephosphorylates Tyr346 with high efficiency but is less efficient at other Syk regulatory sites, establishing Tyr346 dephosphorylation as a key checkpoint in GPVI/Syk signaling.\",\n      \"method\": \"In vitro phosphatase assay with recombinant TULA-2 and Syk peptides/protein, phosphospecific antibodies to multiple Syk sites, GPVI-stimulated platelet signaling with TULA-2 loss-of-function\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro phosphatase assay with site-specific quantification, phosphospecific antibody panel, clear mechanistic checkpoint identification\",\n      \"pmids\": [\"27609517\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Pre-existing GPVI dimers on resting platelets coalesce into clusters upon platelet adhesion to collagenous substrates (fibrous collagen I, soluble collagen III, GPVI-specific collagen peptides). GPVI dimer cluster formation depends on a dynamic actin cytoskeleton. Cluster size is substrate-dependent (collagen III most effective). Some GPVI dimer clusters colocalize with areas of phosphotyrosine, indicating signaling activity. This clustering mechanism increases avidity for collagen and facilitates platelet activation.\",\n      \"method\": \"Total internal reflection fluorescence microscopy, direct stochastic optical reconstruction microscopy (dSTORM), confocal microscopy, fluorophore-conjugated GPVI dimer-specific Fab, actin inhibitors\",\n      \"journal\": \"Journal of thrombosis and haemostasis\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — super-resolution microscopy (dSTORM) plus TIRF real-time imaging, multiple substrates, colocalization with phosphotyrosine signals\",\n      \"pmids\": [\"28058806\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Fibrin and D-dimer (but not the E fragment of fibrin) bind to GPVI and activate platelets. D-dimer binds GPVI by SPR with KD of 302 nM for monomeric GPVI. Immobilized D-dimer induces platelet spreading via Src and Syk kinase activation. Fibrin binds selectively to monomeric GPVI (KD 302 nM), in contrast to collagen which binds primarily to dimeric GPVI. Spreading of human platelets on fibrin is abolished in GPVI-deficient patients, confirming fibrin activates human platelets through GPVI.\",\n      \"method\": \"GPVI-deficient human patient platelets, proteolytic fibrin fragments (D-dimer, E fragment), SPR binding assays, platelet spreading on fibrin/D-dimer, Src/Syk inhibitors, soluble D-dimer inhibition assays\",\n      \"journal\": \"Blood advances\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — SPR with KD determination, human GPVI-deficient platelets, fragment-specific binding characterization, multiple functional assays\",\n      \"pmids\": [\"29296791\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Globular adiponectin (gAd) activates platelets through the GPVI-FcRγ chain complex. gAd stimulates tyrosine phosphorylation of Syk and PLCγ2 (pattern similar to collagen), and this activation is abolished in FcRγ-null platelets (which also lack GPVI). GPVI was confirmed as the receptor for gAd by increased luciferase activity in GPVI-transfected Jurkat T-cells. Full-length adiponectin does not activate platelets through this mechanism.\",\n      \"method\": \"FcRγ-/- and PLCγ2-/- mouse platelets, NFAT luciferase reporter assay in GPVI-transfected Jurkat cells, platelet aggregation, tyrosine phosphorylation western blot\",\n      \"journal\": \"Journal of thrombosis and haemostasis\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic knockout (FcRγ-/-), GPVI transfected reporter cell assay, multiple orthogonal methods\",\n      \"pmids\": [\"18419742\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"PPARγ ligands inhibit GPVI signaling through a non-genomic mechanism. PPARγ associates with Syk and LAT after platelet activation, and PPARγ agonists prevent this association, reducing tyrosine phosphorylation of multiple GPVI signaling components. The inhibitory effect is reversed by the PPARγ antagonist GW9662.\",\n      \"method\": \"Optical aggregometry, spectrofluorimetry (Ca2+ measurement), immunoblot of tyrosine phosphorylation, Co-immunoprecipitation of PPARγ with Syk/LAT, in vitro thrombus formation under flow\",\n      \"journal\": \"Journal of thrombosis and haemostasis\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Co-IP showing PPARγ-Syk/LAT association, antagonist reversal, multiple signaling readouts, single lab\",\n      \"pmids\": [\"20040043\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Nox1, but not Nox2, is the key NADPH oxidase regulating GPVI-dependent reactive oxygen species (ROS) production in platelets. Nox1-derived ROS is essential for CRP-dependent thromboxane A2 production via p38 MAPK signaling downstream of GPVI. Neither Nox1 nor Nox2 was significantly involved in CRP-induced platelet aggregation/integrin αIIbβ3 activation, spreading, or granule release. Both Nox1 and Nox2 are involved in collagen-mediated thrombus formation at arterial shear.\",\n      \"method\": \"Nox2-/- mouse platelets, ML171 (Nox1-specific pharmacological inhibitor), H2DCF-DA oxidation assay for ROS, TxA2 ELISA, p38 MAPK inhibitor, ex vivo perfusion\",\n      \"journal\": \"Redox biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic (Nox2-/-) plus pharmacological (Nox1 inhibitor) approaches, multiple downstream readouts, ex vivo flow\",\n      \"pmids\": [\"24494191\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"FAK (focal adhesion kinase) is downstream of Lyn, Syk, PI3K, and Btk, and upstream of Rac1, PLCγ2, Ca2+ release, PKC, Hic-5, Nox1, and αIIbβ3 activation in the GPVI signaling pathway. FAK, but not Pyk2, is essential for GPVI-dependent ROS production, aggregation, phosphatidylserine exposure, P-selectin expression, and integrin αIIbβ3 activation.\",\n      \"method\": \"Pyk2-/- mouse platelets, FAK inhibitor PF-228, Pyk2 inhibitor Tyrphostin A9, CRP stimulation, ROS assay, aggregation, flow cytometry (PS exposure, P-selectin), immunoprecipitation and pulldown for pathway position\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic (Pyk2-/-) plus pharmacological FAK inhibition, epistasis by inhibitor cascade, multiple functional readouts, single lab\",\n      \"pmids\": [\"25415317\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"CEACAM2 negatively regulates GPVI-FcRγ chain signaling in platelets. CEACAM2-/- platelets exhibit enhanced GPVI agonist (CRP, collagen)-induced aggregation, increased adhesion to type I collagen, and hyperresponsive α and dense granule release. In vivo thrombi in Ceacam2-/- mice are larger and more stable than wild-type, establishing CEACAM2 as a novel negative regulator of platelet GPVI-collagen interactions acting through immunoreceptor tyrosine-based inhibitory motifs.\",\n      \"method\": \"Ceacam2-/- mouse platelets, GPVI and CLEC-2 selective agonists, platelet aggregation, granule secretion, adhesion to collagen I, intravital microscopy (FeCl3 and laser injury models)\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic knockout, selective agonists for pathway specificity, in vitro and in vivo thrombosis models\",\n      \"pmids\": [\"25085348\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Phosphoproteomics downstream of GPVI/ITAM identified >3000 significant phosphorylation events on >1300 proteins. Key signaling relations were mapped including FcRγ→Syk→PLCγ2→PKCδ→DAPP1. Ras/MAPK axis proteins (KSR1, SOS1, STAT1, Hsp27) and >40 Rab GTPases are regulated downstream of GPVI. GPVI-mediated Rab7 S72 phosphorylation and endolysosomal maturation were blocked by TAK1 inhibition, establishing TAK1 as an upstream regulator of GPVI-induced endolysosomal signaling.\",\n      \"method\": \"Peptide TMT labeling, SPS-MS3 phosphoproteomics, causal inference analysis, TAK1 inhibitor functional validation\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — systems phosphoproteomics with TAK1 inhibitor validation for Rab7 finding; broad discovery study, individual findings partially validated\",\n      \"pmids\": [\"32640021\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"GP6-homozygous platelets (carrying premature stop codon before transmembrane domain) show complete loss of surface GPVI and absence of collagen-induced spreading. Under flow, GP6hom blood fails to form stable platelet aggregates or expose phosphatidylserine (PS) on collagen or noncollagen surfaces (VWF/laminin/rhodocytin), but platelet adhesion is preserved. Thrombin generation is partially reduced. The partial adhesion retention explains the mild bleeding diathesis of GP6hom patients.\",\n      \"method\": \"Western blotting, flow cytometry, platelet spreading on collagen/VWF, whole blood perfusion over collagen or noncollagen surfaces under flow, thrombin generation assay, population frequency analysis\",\n      \"journal\": \"Blood advances\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — natural human loss-of-function (homozygous), multiple orthogonal functional assays under flow and static conditions, distinguishes aggregation from adhesion\",\n      \"pmids\": [\"32603422\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Dimeric GPVI binds fibrinogen with much higher affinity and slower dissociation rate than monomeric GPVI due to avidity effects. The αC-region of fibrinogen is the primary binding site for GPVI. GPVI interaction with fibrinogen and fibrin (including non-polymerizing fibrin variant) occurs at similar levels, demonstrating GPVI binding is independent of fibrin polymerization. Fibrin polymerization into fibers clusters GPVI through the αC-region, potentially driving downstream signaling.\",\n      \"method\": \"Multiple protein-protein interaction methods (surface-based and solution-based), SPR, competition assays with fibrinogen fragments, non-polymerizing fibrin variant, dimeric vs monomeric GPVI constructs\",\n      \"journal\": \"Arteriosclerosis, thrombosis, and vascular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — multiple orthogonal binding methods, fragment mapping, non-polymerizing fibrin variant control, avidity mechanism established\",\n      \"pmids\": [\"33472402\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Crystal structure of monomeric GPVI in complex with nanobody Nb2 revealed: (1) Nb2 binding epitope adjacent to the CRP binding groove in the D1 domain; (2) a novel domain-swapped GPVI dimer conformation via the C-C' loop hinge in D2. Truncation of the C-C' loop hinge prevents domain-swapping and abolishes GPVI signaling in a cell-based NFAT reporter assay without blocking ligand binding, establishing that the C-C' loop/domain-swapped structure is required for GPVI signaling but not ligand binding.\",\n      \"method\": \"X-ray crystallography of GPVI-Nb2 complex, C-C' loop truncation mutagenesis, NFAT luciferase reporter assay, Nb2 collagen/CRP displacement binding assay\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure plus mutagenesis plus functional reporter assay in single study, separation of ligand binding from signaling function\",\n      \"pmids\": [\"33512486\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Both CLEC-2 and GPVI (via FcRγ chain) are platelet hemin receptors. Hemin directly binds to both CLEC-2 and GPVI as shown by western blotting and SPR, activating platelets by stimulating SYK and PLCγ2 phosphorylation. Hemin-induced murine platelet aggregation is partially reduced in CLEC-2-depleted or FcRγ-deficient platelets and almost completely inhibited in CLEC-2-depleted FcRγ-deficient double-knockout platelets.\",\n      \"method\": \"SPR binding of hemin to CLEC-2 and GPVI, western blotting, CLEC-2-depleted and FcRγ-/- mouse platelets, double-knockout model, GPVI antibody JAQ-1 inhibition, platelet aggregation\",\n      \"journal\": \"Blood advances\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — SPR binding assay plus multiple genetic models (single and double knockout), functional inhibition with antibody\",\n      \"pmids\": [\"33843987\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Anti-GPVI nanobody Nb2 blocks GPVI-mediated signaling (Syk, LAT, PLCγ2 phosphorylation) and thrombus formation on collagen and atherosclerotic plaque. Non-blocking Nb28 reveals GPVI forms distinct distribution patterns on collagen (clusters along fibers) vs. plaque; clustering on collagen fibers is abolished by Nb2. GPVI clustering is required for downstream signaling and thrombus formation. Adhesion to plaque is maintained despite GPVI signaling blockade and is not inhibited by α2β1 alone.\",\n      \"method\": \"Anti-GPVI nanobodies, western blot of Syk/LAT/PLCγ2 phosphorylation, whole blood thrombus formation under flow, super-resolution/fluorescence microscopy with labeled Nb28, integrin blocking antibodies\",\n      \"journal\": \"Journal of thrombosis and haemostasis\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — live cell super-resolution microscopy, phosphorylation western blots, flow-based thrombus assay, multiple inhibitor combinations\",\n      \"pmids\": [\"35894121\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"GPVI plays a key role in neutrophil recruitment, platelet-neutrophil complex (PNC) formation, and NETosis during acute lung injury (ALI). Anti-GPVI treatment results in less stable platelet-neutrophil interactions, reduced neutrophil crawling and adhesion on endothelial cells, reduced neutrophil transmigration and alveolar infiltrates, and strongly reduced NET formation. GPVI-deficient mice are markedly protected from pulmonary inflammation in LPS-induced ALI, without increased pulmonary bleeding.\",\n      \"method\": \"GPVI-deficient mice, anti-GPVI antibody treatment, LPS-induced ALI model, intravital confocal microscopy of ventilated lung, high-resolution multicolor microscopy of lung sections, PNC quantification, NET staining\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic knockout plus antibody depletion, intravital microscopy, multiple functional readouts (neutrophil crawling, adhesion, transmigration, NETs), in vivo model\",\n      \"pmids\": [\"37441848\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"PLCγ1 partially compensates for PLCγ2 downstream of GPVI in mouse (but not human) platelets. CRP and collagen stimulate tyrosine phosphorylation of PLCγ1 at Tyr783 in mouse platelets through a Src kinase-dependent pathway. In PLCγ2-/- platelets, weak collagen-induced integrin activation persists via PLCγ1 and PI3-kinase signaling downstream of GPVI. Adhesion of PLCγ2-/- platelets to collagen under shear is severely reduced but not abolished (unlike FcRγ-/- platelets).\",\n      \"method\": \"PLCγ2-/- and FcRγ-/- mouse platelets, PLCγ1 phosphospecific antibody (pTyr783), PP1 (Src inhibitor), wortmannin (PI3K inhibitor), αIIbβ3 and α2β1 inhibitors, Born aggregometry, collagen adhesion under flow\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple genetic knockout models, phosphospecific antibodies, kinase inhibitors, flow adhesion assay\",\n      \"pmids\": [\"12730118\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"GPVI is a platelet and megakaryocyte immunoglobulin-superfamily receptor that exists as dimers (and higher-order clusters) on the platelet surface, associates non-covalently with the FcRγ chain, and is activated by collagen, fibrin/fibrinogen (via the αC-region), laminin, EMMPRIN, globular adiponectin, hemin, and several snake venom proteins; upon ligand-induced clustering, constitutively-active Lyn (recruited via a GPVI proline-rich domain) phosphorylates the FcRγ ITAM tyrosines, recruiting and activating Syk (regulated by c-Cbl ubiquitination and TULA-2 dephosphorylation of pTyr346), which—with co-activation of LAT, SLP-76, Gads, and Tec/Btk kinases—drives PLCγ2 activation and downstream Ca2+, PKC, Cdc42, Rac1, and Nox1/ROS-dependent platelet responses including granule secretion, integrin αIIbβ3 inside-out activation, procoagulant PS exposure, and NETosis induction; GPVI surface expression is controlled by ADAM10/ADAM17-mediated ectodomain shedding (generating soluble GPVI) or receptor internalization, both downstream of the FcRγ ITAM, and GP6 transcription during megakaryopoiesis is initiated by TPO-driven CpG demethylation of the GP6 promoter.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"GP6 encodes GPVI, an immunoglobulin-superfamily collagen receptor expressed early during megakaryocyte differentiation and on platelets, where it functions as the principal activating receptor for collagen and related vascular ligands and as a driver of platelet adhesion, activation, and procoagulant responses [#2, #4]. GPVI couples to the FcRγ chain via a transmembrane arginine and intracellular C-terminal tail, and upon ligand-induced clustering, Src-family kinases phosphorylate the FcRγ ITAM tyrosines, recruiting and activating Syk and culminating in PLCγ2 activation [#1, #24]. Receptor avidity is achieved through dimerization and higher-order clustering: dimeric GPVI binds fibrous collagen with high affinity, while pre-existing dimers coalesce into actin cytoskeleton-dependent clusters on collagenous substrates that colocalize with phosphotyrosine signaling [#5, #30], and a domain-swapped dimer conformation mediated by a D2 C-C' loop hinge is structurally required for signaling independent of ligand binding [#40]. Beyond collagen, GPVI is a signaling receptor for laminin, EMMPRIN/CD147, globular adiponectin, hemin, fibrin and the fibrinogen αC-region, and snake venom proteins such as convulxin and alborhagin, each triggering the FcRγ–Syk–PLCγ2 cascade [#3, #11, #21, #27, #32, #39, #41]. The proximal signal is shaped by a constitutive GPVI proline-rich-domain interaction with Lyn [#19], by c-Cbl-mediated ubiquitination that enhances Syk activity, and by TULA-2 dephosphorylation of Syk Tyr346 [#13, #29], with downstream propagation through LAT, SLP-76, Tec/Btk kinases, FAK, Cdc42, and Nox1-derived ROS to control granule secretion, integrin αIIbβ3 activation, and phosphatidylserine exposure [#6, #8, #25, #34, #35]. GPVI surface levels are governed by ADAM10/ADAM17-mediated ectodomain shedding and by receptor internalization, both operating downstream of the FcRγ ITAM, while GP6 transcription is initiated by TPO-driven CpG demethylation of its promoter during megakaryopoiesis [#14, #17, #18]. A homozygous GP6 frameshift producing a truncated protein lacking the transmembrane domain ablates surface GPVI and collagen responsiveness, causing a mild bleeding diathesis [#26, #38]. GPVI also extends beyond hemostasis to inflammation, promoting platelet-neutrophil complex formation and NETosis in acute lung injury [#43].\",\n  \"teleology\": [\n    {\n      \"year\": 1997,\n      \"claim\": \"Established that the platelet collagen receptor p62/GPVI transduces collagen-like activation, defining a Src-kinase-to-Syk phosphorylation cascade through the FcRγ chain.\",\n      \"evidence\": \"Convulxin binding and tyrosine-phosphorylation cascade mapping in platelet lysates with kinase inhibitor\",\n      \"pmids\": [\"9153205\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not establish the receptor's quaternary state\", \"Direct collagen-binding site not yet mapped\"]\n    },\n    {\n      \"year\": 2001,\n      \"claim\": \"Defined the structural requirements for GPVI–FcRγ coupling and demonstrated that GPVI density determines collagen adhesion and signaling, while showing GPVI is functional from early megakaryopoiesis.\",\n      \"evidence\": \"Reconstitution in RBL-2H3/K562/Jurkat cells with transmembrane and C-tail mutagenesis, density-titrated cell lines, and megakaryocyte differentiation assays\",\n      \"pmids\": [\"11152687\", \"11278467\", \"11344165\", \"11723134\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether GPVI signals as monomer or dimer unresolved\", \"Relationship to α2β1 in collagen binding not yet defined\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Resolved that GPVI dimerization, not the monomer, confers high-affinity collagen binding and that the convulxin and collagen sites are distinct, establishing avidity as central to receptor function.\",\n      \"evidence\": \"SPR kinetics and aggregation-inhibition assays comparing monomeric and dimeric recombinant GPVI ectodomains\",\n      \"pmids\": [\"12356768\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Dimer existence on native platelets inferred, not yet imaged\", \"Structural basis of dimerization unknown\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Built the downstream signaling map by ordering LAT, SLP-76, Tec/Btk, and PLCγ1/γ2 contributions and defining differential adaptor requirements relative to TCR signaling.\",\n      \"evidence\": \"Genetic knockout mouse platelets (LAT, SLP-76, Syk, Btk/Tec, PLCγ2, FcRγ) with epistasis and functional readouts; XLA human platelets\",\n      \"pmids\": [\"10567557\", \"11901197\", \"12842985\", \"12730118\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Species differences in PLCγ1 compensation not mechanistically explained\", \"Quantitative contribution of minor LAT-independent branch unclear\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Identified post-translational control points of proximal signaling — c-Cbl-dependent Syk ubiquitination enhancing kinase activity — and the cooperative role of α2β1 in increasing collagen avidity.\",\n      \"evidence\": \"c-Cbl-/- platelets with ubiquitin-IP and in vitro kinase assays; pharmacological α2β1/ADP/COX dissection under flow\",\n      \"pmids\": [\"15701717\", \"12871331\", \"15886326\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism by which ubiquitination raises Syk activity not fully resolved\", \"α2β1 avidity role established largely pharmacologically\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Connected GPVI transcriptional control and ligand repertoire expansion, showing TPO-driven promoter demethylation drives GP6 expression and GPVI serves as a laminin receptor.\",\n      \"evidence\": \"Bisulfite sequencing with TPO treatment; GPVI-/- and Syk-/- platelets, α6 blockade, and SPR for laminin binding\",\n      \"pmids\": [\"15701720\", \"16219796\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Transcription factors acting on the demethylated promoter not identified\", \"Laminin-binding site on GPVI not mapped\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Defined the regulation of GPVI surface expression, distinguishing ADAM-mediated ectodomain shedding from internalization, both downstream of the FcRγ ITAM.\",\n      \"evidence\": \"MALDI-TOF-MS peptide cleavage, ADAM10/ADAM17 enzyme assays, and FcRγ ITAM point-mutant, LAT-/-, and PLCγ2-/- mice with flow cytometry\",\n      \"pmids\": [\"17445093\", \"17374738\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo dominant sheddase not yet pinned down in this work\", \"Signals rendering GPVI susceptible to cleavage incompletely defined\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Characterized the proximal signaling architecture — constitutive Lyn priming via the GPVI proline-rich domain, tetraspanin microdomain localization, and Lyn/PKCδ/SHIP-1 negative regulation of secretion.\",\n      \"evidence\": \"Reciprocal Co-IP, PRD-deletion mutants, Lyn-/-/SHIP-1-/-/PKCδ-/- platelets, and Tspan9 Co-IP/colocalization\",\n      \"pmids\": [\"19940238\", \"18795891\", \"19587372\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Tetraspanin microdomain functional consequence rests on single-lab Co-IP\", \"How Lyn-priming integrates with negative SHIP-1 regulation temporally unclear\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Expanded the GPVI ligand repertoire to EMMPRIN, globular adiponectin, and an alborhagin venom protein, and informed collagen recognition through the homologous LAIR-1 structure.\",\n      \"evidence\": \"SPR/ELISA binding with transfected CHO/Jurkat reporters, FcRγ-/- platelets, and LAIR-1 crystallography with NMR and mutagenesis\",\n      \"pmids\": [\"19350111\", \"18419742\", \"20007810\", \"20040043\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Physiological relevance of EMMPRIN and adiponectin engagement in vivo not established\", \"Conserved collagen-contact residues inferred from a homolog, not GPVI structure\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Extended the downstream effector network to Cdc42, FAK, Nox1-derived ROS, and identified CEACAM2 as an ITIM-bearing negative regulator of GPVI signaling.\",\n      \"evidence\": \"Inducible Cdc42 knockout, Pyk2-/- platelets with FAK inhibitor, Nox2-/- with Nox1 inhibitor, and Ceacam2-/- platelets with in vivo thrombosis models\",\n      \"pmids\": [\"21789221\", \"25415317\", \"24494191\", \"25085348\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"FAK pathway position rests partly on inhibitor epistasis from a single lab\", \"Mechanism of CEACAM2 recruitment to GPVI not detailed\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Established fibrin and fibrinogen as GPVI ligands and resolved the spatial mechanism of activation as actin-dependent clustering of pre-existing dimers.\",\n      \"evidence\": \"GPVI-/- mouse and GPVI-deficient human platelets, SPR fragment mapping (D-dimer, αC-region), and dSTORM/TIRF imaging of dimer clusters\",\n      \"pmids\": [\"26282541\", \"29296791\", \"28058806\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Differential monomer vs dimer preference for fibrin vs collagen mechanistically incomplete\", \"Link between cluster geometry and ITAM phosphorylation kinetics not quantified\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Confirmed the physiological consequence of GPVI loss in humans and mapped the genome-wide phosphosignaling output including a TAK1-Rab7 endolysosomal branch.\",\n      \"evidence\": \"Homozygous GP6 loss-of-function human platelets under flow; TMT/SPS-MS3 phosphoproteomics with TAK1 inhibitor validation\",\n      \"pmids\": [\"32603422\", \"23815599\", \"32640021\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Many phosphoproteomic relations remain individually unvalidated\", \"Mechanism preserving partial adhesion despite GPVI loss not fully defined\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Provided the structural and biophysical basis for GPVI signaling competence, defining the domain-swapped dimer via the D2 C-C' loop and the fibrinogen αC-region/avidity interaction.\",\n      \"evidence\": \"GPVI-Nb2 crystallography with C-C' loop truncation and NFAT reporter; multi-method binding with fibrinogen fragments, non-polymerizing fibrin variant, and hemin SPR\",\n      \"pmids\": [\"33512486\", \"33472402\", \"33843987\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How domain-swapping is triggered by ligand clustering in the membrane unclear\", \"Physiological role of hemin engagement in vivo not established\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Demonstrated GPVI function beyond hemostasis, establishing its role in platelet-neutrophil complex formation and NETosis driving inflammatory injury.\",\n      \"evidence\": \"GPVI-deficient mice and anti-GPVI antibody in LPS-induced acute lung injury with intravital microscopy and NET quantification\",\n      \"pmids\": [\"37441848\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"GPVI ligand mediating neutrophil interactions in inflamed lung not identified\", \"Whether the same FcRγ/Syk cascade drives the NETosis phenotype not dissected\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The identity of the third in vivo GPVI-cleaving sheddase and the membrane-level trigger converting ligand clustering into domain-swapped, signaling-competent dimers remain open.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Third platelet GPVI sheddase beyond ADAM10/ADAM17 unidentified\", \"Coupling between cluster geometry and the D2 domain-swap conformational change unresolved\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0098631\", \"supporting_discovery_ids\": [4, 5, 11, 27]},\n      {\"term_id\": \"GO:0060089\", \"supporting_discovery_ids\": [1, 24, 32]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [19, 30]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [1, 4, 20, 30]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-109582\", \"supporting_discovery_ids\": [4, 12, 27, 38]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [1, 6, 24]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [43]}\n    ],\n    \"complexes\": [\"GPVI-FcRγ receptor complex\", \"tetraspanin (Tspan9) microdomain\"],\n    \"partners\": [\"FCER1G\", \"LYN\", \"SYK\", \"PLCG2\", \"LAT\", \"TSPAN9\", \"CBL\", \"ITGA6\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":8,"faith_total":8,"faith_pct":100.0}}