{"gene":"SPRY2","run_date":"2026-06-10T07:46:41","timeline":{"discoveries":[{"year":2003,"finding":"hSpry2 is ubiquitinated by the E3 ubiquitin ligase c-Cbl in an EGF-dependent manner. EGF stimulation induces tyrosine phosphorylation of hSpry2, which enhances its interaction with c-Cbl, leading to c-Cbl-mediated ubiquitination and degradation of hSpry2 by the 26S proteasome. FGF stimulation also induces hSpry2 degradation, which limits the duration of hSpry2 inhibition of ERK activation.","method":"Co-immunoprecipitation, ubiquitination assay, proteasome inhibitor treatment, in vitro and cell-based degradation assays","journal":"Current biology : CB","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, functional ubiquitination assay, proteasome inhibitor rescue, multiple orthogonal methods in a focused study","pmids":["12593796"],"is_preprint":false},{"year":2004,"finding":"SPRY2 directly binds wild-type BRAF but not the V599E or other exon 15 BRAF mutants, and acts as an inhibitor of ERK signaling in melanocytes and wild-type BRAF melanoma cells but not in cells with the V599E mutation.","method":"siRNA-mediated knockdown, co-immunoprecipitation/pulldown of BRAF-SPRY2 interaction, ERK phosphorylation assays","journal":"Cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct binding shown by pulldown, functional KD with ERK readout, single lab with two orthogonal methods","pmids":["15313890"],"is_preprint":false},{"year":2007,"finding":"Tesk1 (testicular protein kinase 1) interacts with endogenous Spry2, relocalizes Spry2 to vesicles/endosomes (preventing its translocation to membrane ruffles upon growth factor stimulation), and abrogates Spry2 inhibition of ERK phosphorylation. Tesk1 disrupts Spry2 interaction with Grb2 and interferes with Spry2 serine dephosphorylation by PP2A. Tesk1 does not affect Spry2 interaction with c-Cbl or Spry2 inhibition of Cbl-mediated EGFR ubiquitination.","method":"Co-immunoprecipitation of endogenous proteins, subcellular localization imaging, ERK phosphorylation assays, neurite outgrowth assay in PC12 cells, siRNA knockdown, kinase-dead mutants","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP with endogenous proteins, multiple functional assays (ERK, neurite outgrowth, Grb2 interaction, PP2A binding), localization experiments with functional consequence, single rigorous study","pmids":["17974561"],"is_preprint":false},{"year":2010,"finding":"Spry2 forms a complex with c-Cbl ubiquitin ligase and the mTOR repressor TSC2, abolishing GTPase activity directed against Rheb (the G protein inducer of mTORC1), thereby linking FGF-10/FGFR2b/Spry2 signaling to mTORC1 activation and HIF-1α-driven vasculogenesis in fetal lung.","method":"Co-immunoprecipitation of Spry2, c-Cbl, and TSC2 complex; GTPase activity assay; mTORC1 activity assays in fetal lung epithelial cells","journal":"American journal of physiology. Lung cellular and molecular physiology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP identifying complex, functional GTPase assay, single lab with multiple orthogonal methods","pmids":["20622121"],"is_preprint":false},{"year":2010,"finding":"Inhibition of histone demethylase JMJD1A by hypoxia or nickel increases H3K9me2 at the Spry2 promoter, repressing Spry2 expression in bronchial epithelial cells. Forced Spry2 expression decreased nickel-induced ERK phosphorylation and anchorage-independent growth.","method":"ChIP-on-chip, Affymetrix GeneChip, ChIP for H3K9me2, JMJD1A knockdown/overexpression, ERK phosphorylation assay, anchorage-independent growth assay","journal":"Carcinogenesis","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — ChIP identifies histone modification at Spry2 promoter, JMJD1A manipulation with functional readout, multiple orthogonal methods","pmids":["20881000"],"is_preprint":false},{"year":2010,"finding":"Spry2 does not directly modulate Raf-1 kinase activity; instead it acts as a scaffolding protein facilitating interactions between Raf-1 and its direct regulators. Association of Raf-1 with Spry2 does not correlate with Raf-1 activation state across phosphorylation-site mutants.","method":"Co-immunoprecipitation of FLAG-tagged Raf-1 point mutants with Spry2, immunofluorescence colocalization, Raf-1 kinase activity assay","journal":"BMB reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — negative mechanistic result established by Co-IP with multiple mutants and kinase assay, single lab","pmids":["20356462"],"is_preprint":false},{"year":2011,"finding":"In the context of cerebral ischemia, reperfusion induces Src-Spry2 interaction, upregulating Spry2 activity and suppressing the Ras/Raf/MEK/ERK cascade. NMDA receptor antagonist (ketamine) and Src family kinase inhibitor (PP2) both block ischemia-induced Spry2 upregulation, indicating NMDA receptor- and Src kinase-dependent activation of Spry2.","method":"Co-immunoprecipitation of Src-Spry2 in rat hippocampus, immunoblotting for ERK pathway activation, pharmacological inhibitors (PP2, ketamine) in a four-vessel occlusion model","journal":"Brain injury","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP in vivo, pathway inhibition with pharmacological tools, single lab","pmids":["18297599"],"is_preprint":false},{"year":2012,"finding":"Loss of SPRY2 in prostate cancer enhances internalization of EGFR/HER2 (ErbB receptors) via clathrin-mediated endocytosis driven by p38 MAPK activation by PI3K, resulting in sustained ErbB signaling at early endosomes and hyperactivation of PI3K/AKT signaling to drive proliferation and invasion.","method":"SPRY2 knockdown/overexpression, EGFR/HER2 internalization assay, PI3K/AKT signaling measurement, in vitro and in vivo prostate cancer models, PI3K inhibition","journal":"EMBO molecular medicine","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (receptor trafficking assay, signaling assays, in vitro and in vivo models), single rigorous study with mechanistic pathway dissection","pmids":["22649008"],"is_preprint":false},{"year":2012,"finding":"Inactivation of Spry2 (via dominant-negative Spry2Y55F) accelerates AKT-induced hepatocarcinogenesis by activating MAPK and PKM2 pathways. Concomitant suppression of MAPK and PKM2 strongly inhibited Spry2Y55F-driven growth in AKT-overexpressing hepatocellular carcinoma cells.","method":"Hydrodynamic gene delivery of AKT and Spry2Y55F in mouse liver, histological/biochemical assays, stable cell line transfection, MAPK and PKM2 pathway inhibition","journal":"Journal of hepatology","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo mouse model with dominant-negative construct, pathway inhibitor rescue, multiple orthogonal methods","pmids":["22617155"],"is_preprint":false},{"year":2013,"finding":"ITSN1 (intersectin 1) recruits Shp2 tyrosine phosphatase to Spry2 within a Cbl-Spry2 complex, enhancing Spry2 dephosphorylation. Dephosphorylated Spry2 releases its inhibition on Cbl, enhancing Cbl-mediated EGFR ubiquitylation. A catalytically inactive Shp2 mutant reverses this effect. Disrupting the ITSN1-binding site on Spry2 decreases Shp2-Spry2 interaction and enhances Spry2 tyrosine phosphorylation.","method":"Co-immunoprecipitation, point mutagenesis of Spry2 Pro-rich ITSN1-binding site, dominant-negative Shp2, EGFR ubiquitylation assay","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, mutagenesis of binding site, catalytically-inactive mutant rescue, functional ubiquitylation readout, multiple orthogonal methods","pmids":["24216759"],"is_preprint":false},{"year":2016,"finding":"In adult mouse tracheal basal cells, SPRY2 is post-translationally modified (activated) in response to FGFR1 signaling. This allows SPRY2 to inhibit signaling downstream of other receptor tyrosine kinases and maintain basal cell quiescence. Deletion of either Fgfr1 or Spry2 in basal cells increases steady-state proliferation, placing SPRY2 downstream of FGFR1 in an in vivo quiescence mechanism.","method":"Conditional Cre-mediated deletion of Fgfr1 and Spry2 in adult mouse tracheal basal cells, proliferation assays, epistasis by double deletion","journal":"Developmental cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic epistasis with conditional KO, in vivo functional consequence (proliferation), post-translational modification link to FGFR1 established","pmids":["27046834"],"is_preprint":false},{"year":2016,"finding":"Spry2 is a substrate of Dyrk1A kinase; Dyrk1A knockdown increases Spry2 expression and suppresses ERK phosphorylation in TNF-α-treated fibroblast-like synoviocytes, placing Dyrk1A upstream of Spry2 in the ERK MAPK pathway regulation of synoviocyte proliferation, migration and invasion.","method":"Dyrk1A siRNA knockdown, Western blot for Spry2 and p-ERK, proliferation/migration/invasion assays (CCK-8, EdU, transwell)","journal":"Tissue & cell","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — KD with protein-level readout, single lab, epistatic placement confirmed by multiple functional readouts","pmids":["30503061"],"is_preprint":false},{"year":2016,"finding":"Homocysteine alters the expression/activity of Dyrk1A (Spry2 kinase) and PP2A (Spry2 phosphatase) in neural progenitor cells, changing Spry2 phosphorylation pattern, activity and stability, leading to impaired FGF receptor-ERK-cyclin E signaling and reduced proliferation. Hcy also demethylates the Spry2 promoter, upregulating Spry2 mRNA and protein.","method":"Promoter methylation analysis, Spry2 mRNA/protein quantification, Dyrk1A and PP2A activity measurements, neural progenitor proliferation assay","journal":"Biochimica et biophysica acta","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal methods (methylation, kinase/phosphatase activity, protein stability), single lab","pmids":["27686255"],"is_preprint":false},{"year":2017,"finding":"Spry2 is a novel UPR (unfolded protein response) target gene in pancreatic β-cells; its upregulation is dependent on the PERK kinase arm of the UPR. Knockdown of Spry2 reduces Serca2 expression, lowers ER calcium levels, and induces the UPR. Conditional deletion of Spry2 in adult mouse β-cells causes hyperglycemia and hypoinsulinemia.","method":"Whole-genome RNAi screen for insulin promoter regulators, PERK-dependent UPR induction assay, Spry2 KD with Serca2/ER calcium/UPR readout, conditional β-cell-specific Spry2 knockout mouse","journal":"Diabetes","confidence":"High","confidence_rationale":"Tier 2 / Strong — genome-wide screen, conditional KO mouse with in vivo phenotype, mechanistic follow-up with Serca2/ER calcium/UPR measurements, multiple orthogonal methods","pmids":["28246293"],"is_preprint":false},{"year":2018,"finding":"SPRY2 is a novel interactor of the MET receptor tyrosine kinase in rhabdomyosarcoma; SPRY2 colocalizes with and binds MET, and stabilizes MET protein. Depletion of SPRY2 leads to MET degradation, reducing ERK/MAPK pathway activation, migratory and clonogenic potential, and inducing differentiation—phenotypes identical to MET depletion.","method":"Co-immunoprecipitation and colocalization of SPRY2 and MET, siRNA knockdown of SPRY2 and MET, migration, clonogenic, and differentiation assays, ERK/MAPK phosphorylation measurement","journal":"Cell death & disease","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, colocalization, genetic epistasis via parallel KD of SPRY2 and MET producing identical phenotypes, multiple functional readouts","pmids":["29445192"],"is_preprint":false},{"year":2019,"finding":"FOXO3a transcription factor promotes SPRY2 protein stability; knockdown of FOXO3a decreases SPRY2 protein stability. Loss of SPRY2 enhances β-catenin protein stability and activates the β-catenin/TCF4 pathway to induce EMT and promote metastasis in pancreatic ductal adenocarcinoma. Silencing SPRY2 reverses the suppressive effects of FOXO3a overexpression on EMT.","method":"siRNA knockdown of FOXO3a and SPRY2, protein stability assays, Western blot for β-catenin, EMT markers, migration/invasion assays, xenograft model","journal":"Journal of experimental & clinical cancer research : CR","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis (SPRY2 KD reverses FOXO3a effect), β-catenin stabilization assay, in vivo xenograft, single lab","pmids":["30691517"],"is_preprint":false},{"year":2019,"finding":"CRISPR-Cas9 knockout of SPRY2 in HepG2 hepatocytes causes significantly increased glucose uptake and lipid droplet accumulation, with upregulation of PLA2G2A mRNA. These metabolic effects were attenuated (but not reversed) by SPRY2 overexpression, without detectable changes in Akt or MAPK kinase phosphorylation.","method":"CRISPR-Cas9 KO and SPRY2 overexpression, glucose uptake assay, lipid droplet assay, phosphoprotein panel, RNA sequencing","journal":"BMC endocrine disorders","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — CRISPR KO with multiple functional readouts and rescue, single lab, negative kinase phosphorylation result noteworthy","pmids":["31664995"],"is_preprint":false},{"year":2020,"finding":"In glioblastoma, SPRY2 expression is controlled by an autocrine FGFR-NF-κB-ERK feedback loop. ERK reactivation (via NF-κB-driven autocrine FGFR signaling) drives SPRY2 transcription, and SPRY2 in turn promotes resistance to EGFR and MET inhibition. Blocking FGFR signaling or preventing ERK reactivation overcomes this resistance mechanism in vivo.","method":"EGFR/MET inhibitor treatment, FGFR inhibitor combination, ERK-dependent bioluminescent reporter in vivo, SPRY2 knockdown/overexpression, tumor xenograft model","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo reporter validation, SPRY2 KO/OE functional tests, FGFR/NF-κB inhibitor mechanistic dissection, multiple orthogonal methods","pmids":["32160544"],"is_preprint":false},{"year":2020,"finding":"ASIC1a promotes miR-350 expression via METTL3-dependent m6A modification of pri-miR-350 (through METTL3 binding to DGCR8); mature miR-350 directly targets the 3'-UTR of SPRY2 (dual luciferase assay), repressing SPRY2 and activating PI3K/AKT and ERK pathways to promote liver fibrosis.","method":"Dual luciferase reporter assay (miR-350 binding to SPRY2 3'-UTR), m6A modification assay, METTL3-DGCR8 binding assay, ASIC1a knockdown, hepatic stellate cell activation assay","journal":"FASEB journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — dual luciferase validates direct miRNA-SPRY2 interaction, m6A mechanistic pathway established, single lab","pmids":["32949431"],"is_preprint":false},{"year":2023,"finding":"Protein kinase D (PKD) phosphorylates Spry2 at serine 112 in vitro and in vivo, and interacts with the C-terminal half of Spry2. Ser112 phosphorylation promotes Spry2 intracellular degradation; mutation of Ser112 to alanine decreases the rate of Spry2 degradation. Knockdown of all three PKD isoforms or blocking PKD kinase activity stabilizes Spry2 protein. The COP9/Signalosome component CSN3 (which binds PKD) also regulates Spry2 stability in a Ser112-dependent manner.","method":"In vitro kinase assay (PKD phosphorylates Spry2-Ser112), co-immunoprecipitation of PKD-Spry2, Ser112Ala mutagenesis with protein stability assay, PKD isoform siRNA knockdown, PKD inhibitor treatment, CSN3 knockdown","journal":"Oncogenesis","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro kinase assay with mutagenesis, in vivo co-IP, genetic knockdown of writer (PKD) and scaffold (CSN3) with functional stability readout, multiple orthogonal methods","pmids":["37045830"],"is_preprint":false},{"year":2023,"finding":"SPRY2 suppresses Y10 phosphorylation of LDHA and inhibits LDHA enzymatic activity by interfering with the interaction between LDHA and SRC kinase in cancer-associated fibroblasts. SPRY2 knockdown in fibroblasts promotes CAF activation dependent on glycolytic metabolism and enhances breast cancer cell stemness.","method":"Co-immunoprecipitation of SPRY2, LDHA, and SRC; LDHA activity assay; SPRY2 knockdown in fibroblasts; in vivo tumor growth model","journal":"Breast cancer research : BCR","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP identifying SPRY2-LDHA-SRC complex, LDHA activity assay, functional KD with glycolytic and in vivo readouts, single lab","pmids":["37507768"],"is_preprint":false},{"year":2023,"finding":"HOXA5 directly binds the SPRY2 promoter, transcriptionally activating SPRY2 expression, which in turn suppresses MEK/ERK signaling to inhibit prostate cancer stemness and malignant behavior. TRAF7 E3 ubiquitin ligase promotes HOXA5 protein degradation via ubiquitination, thereby indirectly reducing SPRY2 levels.","method":"ChIP assay (HOXA5 binding to SPRY2 promoter), HOXA5 overexpression/knockdown with SPRY2 reporter, TRAF7 ubiquitin ligase assay, SPRY2 knockdown epistasis rescue experiment","journal":"Cell death discovery","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP validates direct promoter binding, epistasis by SPRY2 KD reversing HOXA5 effect, single lab","pmids":["37845209"],"is_preprint":false},{"year":2025,"finding":"SPRY2 acts as a Ras effector: the C-terminal fragment of SPRY2 (residues 161-315) binds oncogenic K-RasG12V more strongly than full-length SPRY2. SPRY2 localizes to the plasma membrane in a K-Ras membrane-anchorage- and activity-dependent manner. Mutations at the predicted K-Ras/SPRY2 C-terminal interface disrupt the interaction. SPRY2 homo- and hetero-oligomerizes with SPRY4. Both full-length SPRY2 and its C-terminal fragment promote C2C12 muscle cell differentiation (requiring MAPK inhibition).","method":"TurboID proximity proteomics, BRET assay, co-immunoprecipitation, K-Ras membrane anchorage inhibitors, interface mutagenesis, C2C12 differentiation assay","journal":"iScience","confidence":"High","confidence_rationale":"Tier 2 / Strong — proximity proteomics plus BRET plus Co-IP (three orthogonal binding methods), mutagenesis of interface, functional differentiation readout, K-Ras inhibitor disruption","pmids":["41362608"],"is_preprint":false},{"year":2024,"finding":"Paclitaxel increases binding of Spry2 to tubulin (shown by co-IP and immunofluorescence), sequestering Spry2 from the plasma membrane and attenuating its negative regulation of the Raf/ERK pathway, thereby aggravating radiation-induced pulmonary fibrosis. Microtubule-depolymerizing agents abolish this effect.","method":"Co-immunoprecipitation (Spry2-tubulin), immunofluorescence colocalization, Spry2 knockdown in fibroblasts (p-c-Raf/p-ERK readout), in vivo mouse thoracic irradiation + PTX model, ERK inhibitor rescue","journal":"The Journal of pharmacology and experimental therapeutics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP and colocalization with functional in vivo model, Spry2 KD validates pathway placement, single lab","pmids":["37918858"],"is_preprint":false},{"year":2000,"finding":"Overexpression of SPRY2 in A375 cells leads to secretion of a soluble factor that inhibits FGF2-stimulated (but not VEGF-stimulated) proliferation of endothelial cells. SPRY2 protein itself binds the intracellular adaptor GRB2 and, as a GFP fusion, remains intracellular, indicating SPRY2 acts intracellularly to regulate secretion of an FGF2 inhibitor rather than functioning as an extracellular inhibitor directly.","method":"Conditioned medium transfer assay, recombinant SPRY2 treatment, SPRY2-GFP fusion localization, GRB2 co-immunoprecipitation/pulldown","journal":"Mechanisms of development","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — functional conditioned medium assay, direct GRB2 binding by pulldown, subcellular localization by GFP fusion, single lab","pmids":["10940627"],"is_preprint":false}],"current_model":"SPRY2 is an inducible intracellular antagonist of RTK/Ras/ERK signaling that is activated by growth factor-stimulated tyrosine phosphorylation (by Src) and inactivated by Dyrk1A-mediated serine phosphorylation at Y55 and PP2A-mediated dephosphorylation; c-Cbl ubiquitinates tyrosine-phosphorylated SPRY2 for proteasomal degradation (a process regulated by intersectin-1/Shp2 and PKD/COP9 signalosome-dependent phosphorylation at Ser112), while Tesk1 sequesters SPRY2 to endosomes to abrogate its inhibition of ERK; SPRY2 binds wild-type BRAF and K-Ras (acting as a Ras effector that blocks downstream effector access), stabilizes MET receptor, interferes with LDHA-SRC interaction to suppress glycolysis, and in β-cells regulates ER calcium homeostasis via Serca2 downstream of the PERK/UPR pathway."},"narrative":{"mechanistic_narrative":"SPRY2 is an inducible intracellular antagonist of receptor tyrosine kinase (RTK)/Ras/ERK signaling that is itself induced and post-translationally tuned by the very growth-factor pathways it restrains [PMID:10940627, PMID:27046834]. Upon EGF or FGF stimulation SPRY2 becomes tyrosine-phosphorylated, which both potentiates its inhibitory function and triggers its turnover: tyrosine-phosphorylated SPRY2 recruits the E3 ligase c-Cbl for ubiquitination and 26S proteasomal degradation, limiting the duration of ERK suppression [PMID:12593796]. This degradation is layered with additional inputs — ITSN1 recruits the Shp2 phosphatase to dephosphorylate SPRY2 within the Cbl-SPRY2 complex (relieving SPRY2's restraint of Cbl-mediated EGFR ubiquitylation) [PMID:24216759], and protein kinase D phosphorylates SPRY2 at Ser112 to drive its degradation in a COP9/signalosome (CSN3)-dependent manner [PMID:37045830]. SPRY2 acts within the ERK module as a scaffold and effector rather than a direct kinase modulator: it binds wild-type BRAF (but not the V599E mutant), engages Raf-1 as a scaffold without altering its intrinsic kinase activity, and binds oncogenic K-Ras through its C-terminal region while oligomerizing with SPRY4 at the plasma membrane [PMID:15313890, PMID:20356462, PMID:41362608]. Beyond ERK, SPRY2 stabilizes the MET receptor [PMID:29445192] and exerts metabolic control by interfering with the LDHA-SRC interaction to suppress glycolysis [PMID:37507768], and in pancreatic β-cells it is a PERK/UPR target gene that maintains Serca2 expression and ER calcium homeostasis, with its deletion causing hyperglycemia and hypoinsulinemia [PMID:28246293]. Across epithelial and stromal tissues SPRY2 functions as a tumor-suppressive brake whose loss sustains RTK/PI3K-AKT and MEK/ERK signaling to drive proliferation, invasion, EMT, and therapy resistance [PMID:22649008, PMID:30691517, PMID:32160544].","teleology":[{"year":2000,"claim":"Established that SPRY2 functions as an intracellular regulator rather than a secreted inhibitor, anchoring the field's view of its site of action.","evidence":"Conditioned-medium transfer, SPRY2-GFP localization, and GRB2 pulldown in A375 cells","pmids":["10940627"],"confidence":"Medium","gaps":["Identity of the secreted FGF2-inhibitory factor not defined","Mechanism linking intracellular SPRY2 to secretion unresolved"]},{"year":2003,"claim":"Answered how SPRY2 inhibition is terminated, showing growth-factor-induced tyrosine phosphorylation drives c-Cbl-mediated ubiquitination and proteasomal degradation as a built-in timer on ERK suppression.","evidence":"Reciprocal Co-IP, ubiquitination assay, and proteasome-inhibitor rescue with EGF/FGF stimulation","pmids":["12593796"],"confidence":"High","gaps":["Specific tyrosine residues and kinases involved not mapped here","Quantitative contribution of degradation versus dephosphorylation to inhibition timing unresolved"]},{"year":2004,"claim":"Defined a node-specific target by showing SPRY2 directly binds wild-type BRAF but not oncogenic V599E, explaining why SPRY2 inhibition is bypassed in BRAF-mutant cells.","evidence":"siRNA knockdown, BRAF-SPRY2 pulldown, and ERK phosphorylation assays in melanoma cells","pmids":["15313890"],"confidence":"Medium","gaps":["Structural basis of preferential wild-type BRAF binding not resolved","Single-lab finding without reciprocal validation"]},{"year":2007,"claim":"Identified a sequestration mechanism whereby Tesk1 relocalizes SPRY2 to endosomes, separating it from Grb2 and blocking its ERK inhibition independently of the Cbl degradation arm.","evidence":"Endogenous reciprocal Co-IP, localization imaging, ERK and neurite outgrowth assays in PC12 cells","pmids":["17974561"],"confidence":"High","gaps":["Whether Tesk1-mediated sequestration is physiologically regulated by growth factors unclear","Relationship between endosomal pool and degradation not quantified"]},{"year":2010,"claim":"Refined the ERK-module mechanism, demonstrating SPRY2 acts as a scaffold for Raf-1 and its regulators rather than directly modulating Raf-1 kinase activity, while parallel work linked SPRY2 to mTORC1 via a Cbl-TSC2 complex.","evidence":"Co-IP of Raf-1 mutants with kinase assays; Co-IP of Spry2-Cbl-TSC2 with GTPase and mTORC1 assays","pmids":["20356462","20622121"],"confidence":"Medium","gaps":["Direct effectors SPRY2 scaffolds onto Raf-1 not enumerated","mTORC1/TSC2 link established in a single tissue context"]},{"year":2010,"claim":"Showed SPRY2 expression is epigenetically controlled, with JMJD1A demethylase activity at the promoter setting SPRY2 levels and downstream ERK-driven transformation.","evidence":"ChIP for H3K9me2, JMJD1A manipulation, ERK and anchorage-independent growth assays in bronchial cells","pmids":["20881000"],"confidence":"High","gaps":["Whether hypoxic SPRY2 repression operates in other tissues unknown"]},{"year":2012,"claim":"Demonstrated SPRY2's tumor-suppressive role by showing its loss reroutes ErbB receptor trafficking to sustain PI3K/AKT signaling and drive prostate cancer proliferation and invasion.","evidence":"SPRY2 KD/OE, receptor internalization and PI3K/AKT assays, in vitro and in vivo prostate models","pmids":["22649008"],"confidence":"High","gaps":["Direct molecular link between SPRY2 and the endocytic machinery not defined"]},{"year":2012,"claim":"Linked SPRY2 inactivation to metabolic reprogramming, showing dominant-negative Spry2Y55F accelerates AKT-driven hepatocarcinogenesis through MAPK and PKM2 pathways.","evidence":"Hydrodynamic delivery of AKT and Spry2Y55F in mouse liver with pathway-inhibitor rescue","pmids":["22617155"],"confidence":"High","gaps":["Mechanism connecting SPRY2 status to PKM2 activity unresolved"]},{"year":2013,"claim":"Resolved the phosphatase arm of SPRY2 regulation, showing ITSN1 recruits Shp2 to dephosphorylate SPRY2 within the Cbl complex, releasing SPRY2's brake on Cbl-mediated EGFR ubiquitylation.","evidence":"Reciprocal Co-IP, ITSN1-binding-site mutagenesis, catalytically inactive Shp2, EGFR ubiquitylation assay","pmids":["24216759"],"confidence":"High","gaps":["How ITSN1/Shp2 recruitment is temporally coordinated with c-Cbl degradation unclear"]},{"year":2016,"claim":"Placed SPRY2 in vivo downstream of FGFR1, showing FGFR1-driven post-translational activation of SPRY2 enforces tracheal basal cell quiescence by cross-inhibiting other RTKs.","evidence":"Conditional Fgfr1/Spry2 deletion with proliferation epistasis in adult mouse trachea","pmids":["27046834"],"confidence":"High","gaps":["Nature of the activating post-translational modification not chemically defined"]},{"year":2016,"claim":"Identified Dyrk1A as a SPRY2 kinase and connected SPRY2 regulation to pathological proliferation and to homocysteine-driven phosphatase/kinase balance affecting SPRY2 stability and FGF-ERK signaling.","evidence":"Dyrk1A KD with Spry2/p-ERK readouts in synoviocytes; Dyrk1A/PP2A activity and promoter-methylation analysis in neural progenitors","pmids":["30503061","27686255"],"confidence":"Medium","gaps":["Exact serine residues phosphorylated by Dyrk1A not mapped in these studies","Functional integration of Dyrk1A and PP2A on SPRY2 not reconstituted"]},{"year":2017,"claim":"Revealed a non-ERK, organelle-homeostasis role, identifying SPRY2 as a PERK/UPR target gene that maintains Serca2-dependent ER calcium in β-cells, with deletion causing hyperglycemia and hypoinsulinemia.","evidence":"Genome-wide RNAi screen, PERK-dependence assay, Serca2/ER-calcium/UPR readouts, conditional β-cell knockout mouse","pmids":["28246293"],"confidence":"High","gaps":["Molecular mechanism linking SPRY2 to Serca2 expression unknown","Whether ER-calcium role is independent of ERK regulation unclear"]},{"year":2018,"claim":"Uncovered a positive-regulatory function on an RTK, showing SPRY2 binds and stabilizes the MET receptor such that its loss phenocopies MET depletion in rhabdomyosarcoma.","evidence":"Reciprocal Co-IP, colocalization, parallel SPRY2/MET knockdown with migration, clonogenic and differentiation readouts","pmids":["29445192"],"confidence":"High","gaps":["How SPRY2 protects MET from degradation mechanistically undefined"]},{"year":2019,"claim":"Connected SPRY2 to transcription-factor-controlled stability and to Wnt/EMT, showing FOXO3a stabilizes SPRY2 while SPRY2 loss stabilizes β-catenin to drive EMT and metastasis in pancreatic cancer.","evidence":"FOXO3a/SPRY2 KD, protein-stability and β-catenin assays, EMT markers, xenograft","pmids":["30691517"],"confidence":"Medium","gaps":["Mechanism by which SPRY2 restrains β-catenin stability not defined","Single-lab epistasis"]},{"year":2019,"claim":"Reported an ERK/AKT-independent metabolic function, with SPRY2 knockout increasing glucose uptake and lipid droplet accumulation in hepatocytes without detectable kinase phosphorylation changes.","evidence":"CRISPR KO/OE with glucose-uptake, lipid-droplet, phosphoprotein panel and RNA-seq","pmids":["31664995"],"confidence":"Medium","gaps":["Effector mediating the metabolic phenotype unidentified","Incomplete rescue on overexpression unexplained"]},{"year":2020,"claim":"Defined feedback and post-transcriptional control of SPRY2, showing an autocrine FGFR-NF-κB-ERK loop drives SPRY2 to confer RTK-inhibitor resistance, while miR-350 (m6A-regulated) directly represses SPRY2 in fibrosis.","evidence":"In vivo ERK reporter with FGFR/NF-κB inhibitors in glioblastoma; dual-luciferase and m6A assays in hepatic stellate cells","pmids":["32160544","32949431"],"confidence":"High","gaps":["Whether the feedback loop and miRNA control intersect not addressed","Context specificity of SPRY2-mediated resistance across RTK inhibitors incomplete"]},{"year":2023,"claim":"Completed a major degradation mechanism by identifying PKD as the Ser112 kinase whose phosphorylation, with CSN3, drives SPRY2 turnover.","evidence":"In vitro kinase assay, Ser112Ala mutagenesis, PKD isoform KD/inhibition, CSN3 KD with stability readouts","pmids":["37045830"],"confidence":"High","gaps":["How Ser112 phosphorylation interfaces with the c-Cbl/Shp2 tyrosine-phosphorylation circuit unresolved"]},{"year":2023,"claim":"Extended SPRY2's reach into stromal metabolism and transcriptional control, showing SPRY2 disrupts the LDHA-SRC interaction to suppress glycolysis in CAFs, and that HOXA5 transcriptionally activates SPRY2 to restrain prostate cancer stemness.","evidence":"SPRY2-LDHA-SRC Co-IP and LDHA activity assay; ChIP of HOXA5 at SPRY2 promoter with TRAF7 ubiquitin assay and epistasis rescue","pmids":["37507768","37845209"],"confidence":"Medium","gaps":["Structural basis of SPRY2 interference with LDHA-SRC undefined","Both findings single-lab"]},{"year":2024,"claim":"Identified microtubule sequestration as an additional off-switch, showing paclitaxel-enhanced SPRY2-tubulin binding pulls SPRY2 from the membrane and de-represses Raf/ERK to worsen pulmonary fibrosis.","evidence":"Spry2-tubulin Co-IP and colocalization, depolymerizer reversal, in vivo irradiation+PTX model with ERK-inhibitor rescue","pmids":["37918858"],"confidence":"Medium","gaps":["Whether tubulin sequestration occurs under physiological (non-paclitaxel) conditions unknown"]},{"year":2025,"claim":"Reframed SPRY2 as a direct K-Ras effector, mapping a C-terminal K-Ras-binding region, membrane recruitment dependent on K-Ras activity, and SPRY2-SPRY4 oligomerization coupled to differentiation control.","evidence":"TurboID, BRET, Co-IP, interface mutagenesis, K-Ras anchorage inhibitors, C2C12 differentiation assay","pmids":["41362608"],"confidence":"High","gaps":["How K-Ras binding mechanistically blocks downstream effector access not structurally resolved","Interplay between K-Ras and BRAF/Raf-1 binding not reconciled"]},{"year":null,"claim":"How the multiple, overlapping regulatory inputs on SPRY2 (Src/Cbl/Shp2 tyrosine cycle, Dyrk1A/PP2A and PKD/CSN3 serine control, Tesk1/tubulin sequestration) are integrated to set its activity and abundance in a given cellular context remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified model reconciling tyrosine- and serine-phosphorylation circuits","Structural basis for SPRY2's simultaneous engagement of K-Ras, BRAF, and Raf-1 undefined","Mechanism of the ERK-independent metabolic and ER-calcium roles unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[0,9,24]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[5,1,22]},{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[23]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[22,2,23]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[24,0]},{"term_id":"GO:0005768","term_label":"endosome","supporting_discovery_ids":[2,7]}],"pathway":[{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[0,1,22,7]},{"term_id":"R-HSA-8953897","term_label":"Cellular responses to stimuli","supporting_discovery_ids":[13]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[7,8,17,14]},{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[0,9,19]}],"complexes":["Cbl-SPRY2 complex","Spry2-c-Cbl-TSC2 complex","SPRY2-SPRY4 oligomer"],"partners":["CBL","GRB2","BRAF","RAF1","KRAS","MET","ITSN1","PRKD1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"O43597","full_name":"Protein sprouty homolog 2","aliases":[],"length_aa":315,"mass_kda":34.7,"function":"Antagonist of fibroblast growth factor (FGF) pathways via inhibition of FGF-mediated phosphorylation of ERK1/2 (By similarity). 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Pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/40915975","citation_count":4,"is_preprint":false},{"pmid":"37918858","id":"PMC_37918858","title":"Paclitaxel Aggravating Radiation-Induced Pulmonary Fibrosis Is Associated with the Down-Regulation of the Negative Regulatory Function of Spry2.","date":"2024","source":"The Journal of pharmacology and experimental therapeutics","url":"https://pubmed.ncbi.nlm.nih.gov/37918858","citation_count":3,"is_preprint":false},{"pmid":"24969401","id":"PMC_24969401","title":"Placental sprouty 2 (SPRY2): relation to placental growth and maternal metabolic status.","date":"2014","source":"Neonatology","url":"https://pubmed.ncbi.nlm.nih.gov/24969401","citation_count":3,"is_preprint":false},{"pmid":"25825238","id":"PMC_25825238","title":"Spry2 regulates signalling dynamics and terminal bud branching behaviour during lung development.","date":"2015","source":"Genetics 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development of osteosarcoma by regulating SPRY2.","date":"2019","source":"European review for medical and pharmacological sciences","url":"https://pubmed.ncbi.nlm.nih.gov/31696462","citation_count":2,"is_preprint":false},{"pmid":"37045830","id":"PMC_37045830","title":"PKD phosphorylation and COP9/Signalosome modulate intracellular Spry2 protein stability.","date":"2023","source":"Oncogenesis","url":"https://pubmed.ncbi.nlm.nih.gov/37045830","citation_count":1,"is_preprint":false},{"pmid":"34457054","id":"PMC_34457054","title":"microRNA-23a promotes cell growth and metastasis in gastric cancer via targeting SPRY2-mediated ERK signaling.","date":"2021","source":"Oncology letters","url":"https://pubmed.ncbi.nlm.nih.gov/34457054","citation_count":1,"is_preprint":false},{"pmid":"24796743","id":"PMC_24796743","title":"[Over-expression of human SPRY2 promotes the proliferation and survival of HEK293T cells].","date":"2014","source":"Xi bao yu fen zi mian yi xue za zhi = Chinese journal of 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EGF stimulation induces tyrosine phosphorylation of hSpry2, which enhances its interaction with c-Cbl, leading to c-Cbl-mediated ubiquitination and degradation of hSpry2 by the 26S proteasome. FGF stimulation also induces hSpry2 degradation, which limits the duration of hSpry2 inhibition of ERK activation.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, proteasome inhibitor treatment, in vitro and cell-based degradation assays\",\n      \"journal\": \"Current biology : CB\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, functional ubiquitination assay, proteasome inhibitor rescue, multiple orthogonal methods in a focused study\",\n      \"pmids\": [\"12593796\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"SPRY2 directly binds wild-type BRAF but not the V599E or other exon 15 BRAF mutants, and acts as an inhibitor of ERK signaling in melanocytes and wild-type BRAF melanoma cells but not in cells with the V599E mutation.\",\n      \"method\": \"siRNA-mediated knockdown, co-immunoprecipitation/pulldown of BRAF-SPRY2 interaction, ERK phosphorylation assays\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct binding shown by pulldown, functional KD with ERK readout, single lab with two orthogonal methods\",\n      \"pmids\": [\"15313890\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Tesk1 (testicular protein kinase 1) interacts with endogenous Spry2, relocalizes Spry2 to vesicles/endosomes (preventing its translocation to membrane ruffles upon growth factor stimulation), and abrogates Spry2 inhibition of ERK phosphorylation. Tesk1 disrupts Spry2 interaction with Grb2 and interferes with Spry2 serine dephosphorylation by PP2A. Tesk1 does not affect Spry2 interaction with c-Cbl or Spry2 inhibition of Cbl-mediated EGFR ubiquitination.\",\n      \"method\": \"Co-immunoprecipitation of endogenous proteins, subcellular localization imaging, ERK phosphorylation assays, neurite outgrowth assay in PC12 cells, siRNA knockdown, kinase-dead mutants\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP with endogenous proteins, multiple functional assays (ERK, neurite outgrowth, Grb2 interaction, PP2A binding), localization experiments with functional consequence, single rigorous study\",\n      \"pmids\": [\"17974561\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Spry2 forms a complex with c-Cbl ubiquitin ligase and the mTOR repressor TSC2, abolishing GTPase activity directed against Rheb (the G protein inducer of mTORC1), thereby linking FGF-10/FGFR2b/Spry2 signaling to mTORC1 activation and HIF-1α-driven vasculogenesis in fetal lung.\",\n      \"method\": \"Co-immunoprecipitation of Spry2, c-Cbl, and TSC2 complex; GTPase activity assay; mTORC1 activity assays in fetal lung epithelial cells\",\n      \"journal\": \"American journal of physiology. Lung cellular and molecular physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP identifying complex, functional GTPase assay, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"20622121\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Inhibition of histone demethylase JMJD1A by hypoxia or nickel increases H3K9me2 at the Spry2 promoter, repressing Spry2 expression in bronchial epithelial cells. Forced Spry2 expression decreased nickel-induced ERK phosphorylation and anchorage-independent growth.\",\n      \"method\": \"ChIP-on-chip, Affymetrix GeneChip, ChIP for H3K9me2, JMJD1A knockdown/overexpression, ERK phosphorylation assay, anchorage-independent growth assay\",\n      \"journal\": \"Carcinogenesis\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — ChIP identifies histone modification at Spry2 promoter, JMJD1A manipulation with functional readout, multiple orthogonal methods\",\n      \"pmids\": [\"20881000\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Spry2 does not directly modulate Raf-1 kinase activity; instead it acts as a scaffolding protein facilitating interactions between Raf-1 and its direct regulators. Association of Raf-1 with Spry2 does not correlate with Raf-1 activation state across phosphorylation-site mutants.\",\n      \"method\": \"Co-immunoprecipitation of FLAG-tagged Raf-1 point mutants with Spry2, immunofluorescence colocalization, Raf-1 kinase activity assay\",\n      \"journal\": \"BMB reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — negative mechanistic result established by Co-IP with multiple mutants and kinase assay, single lab\",\n      \"pmids\": [\"20356462\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"In the context of cerebral ischemia, reperfusion induces Src-Spry2 interaction, upregulating Spry2 activity and suppressing the Ras/Raf/MEK/ERK cascade. NMDA receptor antagonist (ketamine) and Src family kinase inhibitor (PP2) both block ischemia-induced Spry2 upregulation, indicating NMDA receptor- and Src kinase-dependent activation of Spry2.\",\n      \"method\": \"Co-immunoprecipitation of Src-Spry2 in rat hippocampus, immunoblotting for ERK pathway activation, pharmacological inhibitors (PP2, ketamine) in a four-vessel occlusion model\",\n      \"journal\": \"Brain injury\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP in vivo, pathway inhibition with pharmacological tools, single lab\",\n      \"pmids\": [\"18297599\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Loss of SPRY2 in prostate cancer enhances internalization of EGFR/HER2 (ErbB receptors) via clathrin-mediated endocytosis driven by p38 MAPK activation by PI3K, resulting in sustained ErbB signaling at early endosomes and hyperactivation of PI3K/AKT signaling to drive proliferation and invasion.\",\n      \"method\": \"SPRY2 knockdown/overexpression, EGFR/HER2 internalization assay, PI3K/AKT signaling measurement, in vitro and in vivo prostate cancer models, PI3K inhibition\",\n      \"journal\": \"EMBO molecular medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (receptor trafficking assay, signaling assays, in vitro and in vivo models), single rigorous study with mechanistic pathway dissection\",\n      \"pmids\": [\"22649008\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Inactivation of Spry2 (via dominant-negative Spry2Y55F) accelerates AKT-induced hepatocarcinogenesis by activating MAPK and PKM2 pathways. Concomitant suppression of MAPK and PKM2 strongly inhibited Spry2Y55F-driven growth in AKT-overexpressing hepatocellular carcinoma cells.\",\n      \"method\": \"Hydrodynamic gene delivery of AKT and Spry2Y55F in mouse liver, histological/biochemical assays, stable cell line transfection, MAPK and PKM2 pathway inhibition\",\n      \"journal\": \"Journal of hepatology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo mouse model with dominant-negative construct, pathway inhibitor rescue, multiple orthogonal methods\",\n      \"pmids\": [\"22617155\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"ITSN1 (intersectin 1) recruits Shp2 tyrosine phosphatase to Spry2 within a Cbl-Spry2 complex, enhancing Spry2 dephosphorylation. Dephosphorylated Spry2 releases its inhibition on Cbl, enhancing Cbl-mediated EGFR ubiquitylation. A catalytically inactive Shp2 mutant reverses this effect. Disrupting the ITSN1-binding site on Spry2 decreases Shp2-Spry2 interaction and enhances Spry2 tyrosine phosphorylation.\",\n      \"method\": \"Co-immunoprecipitation, point mutagenesis of Spry2 Pro-rich ITSN1-binding site, dominant-negative Shp2, EGFR ubiquitylation assay\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, mutagenesis of binding site, catalytically-inactive mutant rescue, functional ubiquitylation readout, multiple orthogonal methods\",\n      \"pmids\": [\"24216759\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"In adult mouse tracheal basal cells, SPRY2 is post-translationally modified (activated) in response to FGFR1 signaling. This allows SPRY2 to inhibit signaling downstream of other receptor tyrosine kinases and maintain basal cell quiescence. Deletion of either Fgfr1 or Spry2 in basal cells increases steady-state proliferation, placing SPRY2 downstream of FGFR1 in an in vivo quiescence mechanism.\",\n      \"method\": \"Conditional Cre-mediated deletion of Fgfr1 and Spry2 in adult mouse tracheal basal cells, proliferation assays, epistasis by double deletion\",\n      \"journal\": \"Developmental cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic epistasis with conditional KO, in vivo functional consequence (proliferation), post-translational modification link to FGFR1 established\",\n      \"pmids\": [\"27046834\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Spry2 is a substrate of Dyrk1A kinase; Dyrk1A knockdown increases Spry2 expression and suppresses ERK phosphorylation in TNF-α-treated fibroblast-like synoviocytes, placing Dyrk1A upstream of Spry2 in the ERK MAPK pathway regulation of synoviocyte proliferation, migration and invasion.\",\n      \"method\": \"Dyrk1A siRNA knockdown, Western blot for Spry2 and p-ERK, proliferation/migration/invasion assays (CCK-8, EdU, transwell)\",\n      \"journal\": \"Tissue & cell\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — KD with protein-level readout, single lab, epistatic placement confirmed by multiple functional readouts\",\n      \"pmids\": [\"30503061\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Homocysteine alters the expression/activity of Dyrk1A (Spry2 kinase) and PP2A (Spry2 phosphatase) in neural progenitor cells, changing Spry2 phosphorylation pattern, activity and stability, leading to impaired FGF receptor-ERK-cyclin E signaling and reduced proliferation. Hcy also demethylates the Spry2 promoter, upregulating Spry2 mRNA and protein.\",\n      \"method\": \"Promoter methylation analysis, Spry2 mRNA/protein quantification, Dyrk1A and PP2A activity measurements, neural progenitor proliferation assay\",\n      \"journal\": \"Biochimica et biophysica acta\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal methods (methylation, kinase/phosphatase activity, protein stability), single lab\",\n      \"pmids\": [\"27686255\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Spry2 is a novel UPR (unfolded protein response) target gene in pancreatic β-cells; its upregulation is dependent on the PERK kinase arm of the UPR. Knockdown of Spry2 reduces Serca2 expression, lowers ER calcium levels, and induces the UPR. Conditional deletion of Spry2 in adult mouse β-cells causes hyperglycemia and hypoinsulinemia.\",\n      \"method\": \"Whole-genome RNAi screen for insulin promoter regulators, PERK-dependent UPR induction assay, Spry2 KD with Serca2/ER calcium/UPR readout, conditional β-cell-specific Spry2 knockout mouse\",\n      \"journal\": \"Diabetes\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genome-wide screen, conditional KO mouse with in vivo phenotype, mechanistic follow-up with Serca2/ER calcium/UPR measurements, multiple orthogonal methods\",\n      \"pmids\": [\"28246293\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"SPRY2 is a novel interactor of the MET receptor tyrosine kinase in rhabdomyosarcoma; SPRY2 colocalizes with and binds MET, and stabilizes MET protein. Depletion of SPRY2 leads to MET degradation, reducing ERK/MAPK pathway activation, migratory and clonogenic potential, and inducing differentiation—phenotypes identical to MET depletion.\",\n      \"method\": \"Co-immunoprecipitation and colocalization of SPRY2 and MET, siRNA knockdown of SPRY2 and MET, migration, clonogenic, and differentiation assays, ERK/MAPK phosphorylation measurement\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, colocalization, genetic epistasis via parallel KD of SPRY2 and MET producing identical phenotypes, multiple functional readouts\",\n      \"pmids\": [\"29445192\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"FOXO3a transcription factor promotes SPRY2 protein stability; knockdown of FOXO3a decreases SPRY2 protein stability. Loss of SPRY2 enhances β-catenin protein stability and activates the β-catenin/TCF4 pathway to induce EMT and promote metastasis in pancreatic ductal adenocarcinoma. Silencing SPRY2 reverses the suppressive effects of FOXO3a overexpression on EMT.\",\n      \"method\": \"siRNA knockdown of FOXO3a and SPRY2, protein stability assays, Western blot for β-catenin, EMT markers, migration/invasion assays, xenograft model\",\n      \"journal\": \"Journal of experimental & clinical cancer research : CR\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis (SPRY2 KD reverses FOXO3a effect), β-catenin stabilization assay, in vivo xenograft, single lab\",\n      \"pmids\": [\"30691517\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"CRISPR-Cas9 knockout of SPRY2 in HepG2 hepatocytes causes significantly increased glucose uptake and lipid droplet accumulation, with upregulation of PLA2G2A mRNA. These metabolic effects were attenuated (but not reversed) by SPRY2 overexpression, without detectable changes in Akt or MAPK kinase phosphorylation.\",\n      \"method\": \"CRISPR-Cas9 KO and SPRY2 overexpression, glucose uptake assay, lipid droplet assay, phosphoprotein panel, RNA sequencing\",\n      \"journal\": \"BMC endocrine disorders\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — CRISPR KO with multiple functional readouts and rescue, single lab, negative kinase phosphorylation result noteworthy\",\n      \"pmids\": [\"31664995\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"In glioblastoma, SPRY2 expression is controlled by an autocrine FGFR-NF-κB-ERK feedback loop. ERK reactivation (via NF-κB-driven autocrine FGFR signaling) drives SPRY2 transcription, and SPRY2 in turn promotes resistance to EGFR and MET inhibition. Blocking FGFR signaling or preventing ERK reactivation overcomes this resistance mechanism in vivo.\",\n      \"method\": \"EGFR/MET inhibitor treatment, FGFR inhibitor combination, ERK-dependent bioluminescent reporter in vivo, SPRY2 knockdown/overexpression, tumor xenograft model\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo reporter validation, SPRY2 KO/OE functional tests, FGFR/NF-κB inhibitor mechanistic dissection, multiple orthogonal methods\",\n      \"pmids\": [\"32160544\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"ASIC1a promotes miR-350 expression via METTL3-dependent m6A modification of pri-miR-350 (through METTL3 binding to DGCR8); mature miR-350 directly targets the 3'-UTR of SPRY2 (dual luciferase assay), repressing SPRY2 and activating PI3K/AKT and ERK pathways to promote liver fibrosis.\",\n      \"method\": \"Dual luciferase reporter assay (miR-350 binding to SPRY2 3'-UTR), m6A modification assay, METTL3-DGCR8 binding assay, ASIC1a knockdown, hepatic stellate cell activation assay\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — dual luciferase validates direct miRNA-SPRY2 interaction, m6A mechanistic pathway established, single lab\",\n      \"pmids\": [\"32949431\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Protein kinase D (PKD) phosphorylates Spry2 at serine 112 in vitro and in vivo, and interacts with the C-terminal half of Spry2. Ser112 phosphorylation promotes Spry2 intracellular degradation; mutation of Ser112 to alanine decreases the rate of Spry2 degradation. Knockdown of all three PKD isoforms or blocking PKD kinase activity stabilizes Spry2 protein. The COP9/Signalosome component CSN3 (which binds PKD) also regulates Spry2 stability in a Ser112-dependent manner.\",\n      \"method\": \"In vitro kinase assay (PKD phosphorylates Spry2-Ser112), co-immunoprecipitation of PKD-Spry2, Ser112Ala mutagenesis with protein stability assay, PKD isoform siRNA knockdown, PKD inhibitor treatment, CSN3 knockdown\",\n      \"journal\": \"Oncogenesis\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro kinase assay with mutagenesis, in vivo co-IP, genetic knockdown of writer (PKD) and scaffold (CSN3) with functional stability readout, multiple orthogonal methods\",\n      \"pmids\": [\"37045830\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"SPRY2 suppresses Y10 phosphorylation of LDHA and inhibits LDHA enzymatic activity by interfering with the interaction between LDHA and SRC kinase in cancer-associated fibroblasts. SPRY2 knockdown in fibroblasts promotes CAF activation dependent on glycolytic metabolism and enhances breast cancer cell stemness.\",\n      \"method\": \"Co-immunoprecipitation of SPRY2, LDHA, and SRC; LDHA activity assay; SPRY2 knockdown in fibroblasts; in vivo tumor growth model\",\n      \"journal\": \"Breast cancer research : BCR\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP identifying SPRY2-LDHA-SRC complex, LDHA activity assay, functional KD with glycolytic and in vivo readouts, single lab\",\n      \"pmids\": [\"37507768\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"HOXA5 directly binds the SPRY2 promoter, transcriptionally activating SPRY2 expression, which in turn suppresses MEK/ERK signaling to inhibit prostate cancer stemness and malignant behavior. TRAF7 E3 ubiquitin ligase promotes HOXA5 protein degradation via ubiquitination, thereby indirectly reducing SPRY2 levels.\",\n      \"method\": \"ChIP assay (HOXA5 binding to SPRY2 promoter), HOXA5 overexpression/knockdown with SPRY2 reporter, TRAF7 ubiquitin ligase assay, SPRY2 knockdown epistasis rescue experiment\",\n      \"journal\": \"Cell death discovery\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP validates direct promoter binding, epistasis by SPRY2 KD reversing HOXA5 effect, single lab\",\n      \"pmids\": [\"37845209\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"SPRY2 acts as a Ras effector: the C-terminal fragment of SPRY2 (residues 161-315) binds oncogenic K-RasG12V more strongly than full-length SPRY2. SPRY2 localizes to the plasma membrane in a K-Ras membrane-anchorage- and activity-dependent manner. Mutations at the predicted K-Ras/SPRY2 C-terminal interface disrupt the interaction. SPRY2 homo- and hetero-oligomerizes with SPRY4. Both full-length SPRY2 and its C-terminal fragment promote C2C12 muscle cell differentiation (requiring MAPK inhibition).\",\n      \"method\": \"TurboID proximity proteomics, BRET assay, co-immunoprecipitation, K-Ras membrane anchorage inhibitors, interface mutagenesis, C2C12 differentiation assay\",\n      \"journal\": \"iScience\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — proximity proteomics plus BRET plus Co-IP (three orthogonal binding methods), mutagenesis of interface, functional differentiation readout, K-Ras inhibitor disruption\",\n      \"pmids\": [\"41362608\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Paclitaxel increases binding of Spry2 to tubulin (shown by co-IP and immunofluorescence), sequestering Spry2 from the plasma membrane and attenuating its negative regulation of the Raf/ERK pathway, thereby aggravating radiation-induced pulmonary fibrosis. Microtubule-depolymerizing agents abolish this effect.\",\n      \"method\": \"Co-immunoprecipitation (Spry2-tubulin), immunofluorescence colocalization, Spry2 knockdown in fibroblasts (p-c-Raf/p-ERK readout), in vivo mouse thoracic irradiation + PTX model, ERK inhibitor rescue\",\n      \"journal\": \"The Journal of pharmacology and experimental therapeutics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP and colocalization with functional in vivo model, Spry2 KD validates pathway placement, single lab\",\n      \"pmids\": [\"37918858\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"Overexpression of SPRY2 in A375 cells leads to secretion of a soluble factor that inhibits FGF2-stimulated (but not VEGF-stimulated) proliferation of endothelial cells. SPRY2 protein itself binds the intracellular adaptor GRB2 and, as a GFP fusion, remains intracellular, indicating SPRY2 acts intracellularly to regulate secretion of an FGF2 inhibitor rather than functioning as an extracellular inhibitor directly.\",\n      \"method\": \"Conditioned medium transfer assay, recombinant SPRY2 treatment, SPRY2-GFP fusion localization, GRB2 co-immunoprecipitation/pulldown\",\n      \"journal\": \"Mechanisms of development\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — functional conditioned medium assay, direct GRB2 binding by pulldown, subcellular localization by GFP fusion, single lab\",\n      \"pmids\": [\"10940627\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"SPRY2 is an inducible intracellular antagonist of RTK/Ras/ERK signaling that is activated by growth factor-stimulated tyrosine phosphorylation (by Src) and inactivated by Dyrk1A-mediated serine phosphorylation at Y55 and PP2A-mediated dephosphorylation; c-Cbl ubiquitinates tyrosine-phosphorylated SPRY2 for proteasomal degradation (a process regulated by intersectin-1/Shp2 and PKD/COP9 signalosome-dependent phosphorylation at Ser112), while Tesk1 sequesters SPRY2 to endosomes to abrogate its inhibition of ERK; SPRY2 binds wild-type BRAF and K-Ras (acting as a Ras effector that blocks downstream effector access), stabilizes MET receptor, interferes with LDHA-SRC interaction to suppress glycolysis, and in β-cells regulates ER calcium homeostasis via Serca2 downstream of the PERK/UPR pathway.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"SPRY2 is an inducible intracellular antagonist of receptor tyrosine kinase (RTK)/Ras/ERK signaling that is itself induced and post-translationally tuned by the very growth-factor pathways it restrains [#24, #10]. Upon EGF or FGF stimulation SPRY2 becomes tyrosine-phosphorylated, which both potentiates its inhibitory function and triggers its turnover: tyrosine-phosphorylated SPRY2 recruits the E3 ligase c-Cbl for ubiquitination and 26S proteasomal degradation, limiting the duration of ERK suppression [#0]. This degradation is layered with additional inputs — ITSN1 recruits the Shp2 phosphatase to dephosphorylate SPRY2 within the Cbl-SPRY2 complex (relieving SPRY2's restraint of Cbl-mediated EGFR ubiquitylation) [#9], and protein kinase D phosphorylates SPRY2 at Ser112 to drive its degradation in a COP9/signalosome (CSN3)-dependent manner [#19]. SPRY2 acts within the ERK module as a scaffold and effector rather than a direct kinase modulator: it binds wild-type BRAF (but not the V599E mutant), engages Raf-1 as a scaffold without altering its intrinsic kinase activity, and binds oncogenic K-Ras through its C-terminal region while oligomerizing with SPRY4 at the plasma membrane [#1, #5, #22]. Beyond ERK, SPRY2 stabilizes the MET receptor [#14] and exerts metabolic control by interfering with the LDHA-SRC interaction to suppress glycolysis [#20], and in pancreatic \\u03b2-cells it is a PERK/UPR target gene that maintains Serca2 expression and ER calcium homeostasis, with its deletion causing hyperglycemia and hypoinsulinemia [#13]. Across epithelial and stromal tissues SPRY2 functions as a tumor-suppressive brake whose loss sustains RTK/PI3K-AKT and MEK/ERK signaling to drive proliferation, invasion, EMT, and therapy resistance [#7, #15, #17].\",\n  \"teleology\": [\n    {\n      \"year\": 2000,\n      \"claim\": \"Established that SPRY2 functions as an intracellular regulator rather than a secreted inhibitor, anchoring the field's view of its site of action.\",\n      \"evidence\": \"Conditioned-medium transfer, SPRY2-GFP localization, and GRB2 pulldown in A375 cells\",\n      \"pmids\": [\"10940627\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Identity of the secreted FGF2-inhibitory factor not defined\", \"Mechanism linking intracellular SPRY2 to secretion unresolved\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Answered how SPRY2 inhibition is terminated, showing growth-factor-induced tyrosine phosphorylation drives c-Cbl-mediated ubiquitination and proteasomal degradation as a built-in timer on ERK suppression.\",\n      \"evidence\": \"Reciprocal Co-IP, ubiquitination assay, and proteasome-inhibitor rescue with EGF/FGF stimulation\",\n      \"pmids\": [\"12593796\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Specific tyrosine residues and kinases involved not mapped here\", \"Quantitative contribution of degradation versus dephosphorylation to inhibition timing unresolved\"]\n    },\n    {\n      \"year\": 2004,\n      \"claim\": \"Defined a node-specific target by showing SPRY2 directly binds wild-type BRAF but not oncogenic V599E, explaining why SPRY2 inhibition is bypassed in BRAF-mutant cells.\",\n      \"evidence\": \"siRNA knockdown, BRAF-SPRY2 pulldown, and ERK phosphorylation assays in melanoma cells\",\n      \"pmids\": [\"15313890\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural basis of preferential wild-type BRAF binding not resolved\", \"Single-lab finding without reciprocal validation\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Identified a sequestration mechanism whereby Tesk1 relocalizes SPRY2 to endosomes, separating it from Grb2 and blocking its ERK inhibition independently of the Cbl degradation arm.\",\n      \"evidence\": \"Endogenous reciprocal Co-IP, localization imaging, ERK and neurite outgrowth assays in PC12 cells\",\n      \"pmids\": [\"17974561\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether Tesk1-mediated sequestration is physiologically regulated by growth factors unclear\", \"Relationship between endosomal pool and degradation not quantified\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Refined the ERK-module mechanism, demonstrating SPRY2 acts as a scaffold for Raf-1 and its regulators rather than directly modulating Raf-1 kinase activity, while parallel work linked SPRY2 to mTORC1 via a Cbl-TSC2 complex.\",\n      \"evidence\": \"Co-IP of Raf-1 mutants with kinase assays; Co-IP of Spry2-Cbl-TSC2 with GTPase and mTORC1 assays\",\n      \"pmids\": [\"20356462\", \"20622121\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct effectors SPRY2 scaffolds onto Raf-1 not enumerated\", \"mTORC1/TSC2 link established in a single tissue context\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Showed SPRY2 expression is epigenetically controlled, with JMJD1A demethylase activity at the promoter setting SPRY2 levels and downstream ERK-driven transformation.\",\n      \"evidence\": \"ChIP for H3K9me2, JMJD1A manipulation, ERK and anchorage-independent growth assays in bronchial cells\",\n      \"pmids\": [\"20881000\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether hypoxic SPRY2 repression operates in other tissues unknown\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Demonstrated SPRY2's tumor-suppressive role by showing its loss reroutes ErbB receptor trafficking to sustain PI3K/AKT signaling and drive prostate cancer proliferation and invasion.\",\n      \"evidence\": \"SPRY2 KD/OE, receptor internalization and PI3K/AKT assays, in vitro and in vivo prostate models\",\n      \"pmids\": [\"22649008\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct molecular link between SPRY2 and the endocytic machinery not defined\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Linked SPRY2 inactivation to metabolic reprogramming, showing dominant-negative Spry2Y55F accelerates AKT-driven hepatocarcinogenesis through MAPK and PKM2 pathways.\",\n      \"evidence\": \"Hydrodynamic delivery of AKT and Spry2Y55F in mouse liver with pathway-inhibitor rescue\",\n      \"pmids\": [\"22617155\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism connecting SPRY2 status to PKM2 activity unresolved\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Resolved the phosphatase arm of SPRY2 regulation, showing ITSN1 recruits Shp2 to dephosphorylate SPRY2 within the Cbl complex, releasing SPRY2's brake on Cbl-mediated EGFR ubiquitylation.\",\n      \"evidence\": \"Reciprocal Co-IP, ITSN1-binding-site mutagenesis, catalytically inactive Shp2, EGFR ubiquitylation assay\",\n      \"pmids\": [\"24216759\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How ITSN1/Shp2 recruitment is temporally coordinated with c-Cbl degradation unclear\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Placed SPRY2 in vivo downstream of FGFR1, showing FGFR1-driven post-translational activation of SPRY2 enforces tracheal basal cell quiescence by cross-inhibiting other RTKs.\",\n      \"evidence\": \"Conditional Fgfr1/Spry2 deletion with proliferation epistasis in adult mouse trachea\",\n      \"pmids\": [\"27046834\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Nature of the activating post-translational modification not chemically defined\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Identified Dyrk1A as a SPRY2 kinase and connected SPRY2 regulation to pathological proliferation and to homocysteine-driven phosphatase/kinase balance affecting SPRY2 stability and FGF-ERK signaling.\",\n      \"evidence\": \"Dyrk1A KD with Spry2/p-ERK readouts in synoviocytes; Dyrk1A/PP2A activity and promoter-methylation analysis in neural progenitors\",\n      \"pmids\": [\"30503061\", \"27686255\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Exact serine residues phosphorylated by Dyrk1A not mapped in these studies\", \"Functional integration of Dyrk1A and PP2A on SPRY2 not reconstituted\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Revealed a non-ERK, organelle-homeostasis role, identifying SPRY2 as a PERK/UPR target gene that maintains Serca2-dependent ER calcium in \\u03b2-cells, with deletion causing hyperglycemia and hypoinsulinemia.\",\n      \"evidence\": \"Genome-wide RNAi screen, PERK-dependence assay, Serca2/ER-calcium/UPR readouts, conditional \\u03b2-cell knockout mouse\",\n      \"pmids\": [\"28246293\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular mechanism linking SPRY2 to Serca2 expression unknown\", \"Whether ER-calcium role is independent of ERK regulation unclear\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Uncovered a positive-regulatory function on an RTK, showing SPRY2 binds and stabilizes the MET receptor such that its loss phenocopies MET depletion in rhabdomyosarcoma.\",\n      \"evidence\": \"Reciprocal Co-IP, colocalization, parallel SPRY2/MET knockdown with migration, clonogenic and differentiation readouts\",\n      \"pmids\": [\"29445192\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How SPRY2 protects MET from degradation mechanistically undefined\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Connected SPRY2 to transcription-factor-controlled stability and to Wnt/EMT, showing FOXO3a stabilizes SPRY2 while SPRY2 loss stabilizes \\u03b2-catenin to drive EMT and metastasis in pancreatic cancer.\",\n      \"evidence\": \"FOXO3a/SPRY2 KD, protein-stability and \\u03b2-catenin assays, EMT markers, xenograft\",\n      \"pmids\": [\"30691517\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism by which SPRY2 restrains \\u03b2-catenin stability not defined\", \"Single-lab epistasis\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Reported an ERK/AKT-independent metabolic function, with SPRY2 knockout increasing glucose uptake and lipid droplet accumulation in hepatocytes without detectable kinase phosphorylation changes.\",\n      \"evidence\": \"CRISPR KO/OE with glucose-uptake, lipid-droplet, phosphoprotein panel and RNA-seq\",\n      \"pmids\": [\"31664995\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Effector mediating the metabolic phenotype unidentified\", \"Incomplete rescue on overexpression unexplained\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Defined feedback and post-transcriptional control of SPRY2, showing an autocrine FGFR-NF-\\u03baB-ERK loop drives SPRY2 to confer RTK-inhibitor resistance, while miR-350 (m6A-regulated) directly represses SPRY2 in fibrosis.\",\n      \"evidence\": \"In vivo ERK reporter with FGFR/NF-\\u03baB inhibitors in glioblastoma; dual-luciferase and m6A assays in hepatic stellate cells\",\n      \"pmids\": [\"32160544\", \"32949431\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether the feedback loop and miRNA control intersect not addressed\", \"Context specificity of SPRY2-mediated resistance across RTK inhibitors incomplete\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Completed a major degradation mechanism by identifying PKD as the Ser112 kinase whose phosphorylation, with CSN3, drives SPRY2 turnover.\",\n      \"evidence\": \"In vitro kinase assay, Ser112Ala mutagenesis, PKD isoform KD/inhibition, CSN3 KD with stability readouts\",\n      \"pmids\": [\"37045830\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How Ser112 phosphorylation interfaces with the c-Cbl/Shp2 tyrosine-phosphorylation circuit unresolved\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Extended SPRY2's reach into stromal metabolism and transcriptional control, showing SPRY2 disrupts the LDHA-SRC interaction to suppress glycolysis in CAFs, and that HOXA5 transcriptionally activates SPRY2 to restrain prostate cancer stemness.\",\n      \"evidence\": \"SPRY2-LDHA-SRC Co-IP and LDHA activity assay; ChIP of HOXA5 at SPRY2 promoter with TRAF7 ubiquitin assay and epistasis rescue\",\n      \"pmids\": [\"37507768\", \"37845209\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural basis of SPRY2 interference with LDHA-SRC undefined\", \"Both findings single-lab\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Identified microtubule sequestration as an additional off-switch, showing paclitaxel-enhanced SPRY2-tubulin binding pulls SPRY2 from the membrane and de-represses Raf/ERK to worsen pulmonary fibrosis.\",\n      \"evidence\": \"Spry2-tubulin Co-IP and colocalization, depolymerizer reversal, in vivo irradiation+PTX model with ERK-inhibitor rescue\",\n      \"pmids\": [\"37918858\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether tubulin sequestration occurs under physiological (non-paclitaxel) conditions unknown\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Reframed SPRY2 as a direct K-Ras effector, mapping a C-terminal K-Ras-binding region, membrane recruitment dependent on K-Ras activity, and SPRY2-SPRY4 oligomerization coupled to differentiation control.\",\n      \"evidence\": \"TurboID, BRET, Co-IP, interface mutagenesis, K-Ras anchorage inhibitors, C2C12 differentiation assay\",\n      \"pmids\": [\"41362608\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How K-Ras binding mechanistically blocks downstream effector access not structurally resolved\", \"Interplay between K-Ras and BRAF/Raf-1 binding not reconciled\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How the multiple, overlapping regulatory inputs on SPRY2 (Src/Cbl/Shp2 tyrosine cycle, Dyrk1A/PP2A and PKD/CSN3 serine control, Tesk1/tubulin sequestration) are integrated to set its activity and abundance in a given cellular context remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified model reconciling tyrosine- and serine-phosphorylation circuits\", \"Structural basis for SPRY2's simultaneous engagement of K-Ras, BRAF, and Raf-1 undefined\", \"Mechanism of the ERK-independent metabolic and ER-calcium roles unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [0, 9, 24]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [5, 1, 22]},\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [23]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [22, 2, 23]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [24, 0]},\n      {\"term_id\": \"GO:0005768\", \"supporting_discovery_ids\": [2, 7]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [0, 1, 22, 7]},\n      {\"term_id\": \"R-HSA-8953897\", \"supporting_discovery_ids\": [13]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [7, 8, 17, 14]},\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [0, 9, 19]}\n    ],\n    \"complexes\": [\n      \"Cbl-SPRY2 complex\",\n      \"Spry2-c-Cbl-TSC2 complex\",\n      \"SPRY2-SPRY4 oligomer\"\n    ],\n    \"partners\": [\n      \"CBL\",\n      \"GRB2\",\n      \"BRAF\",\n      \"RAF1\",\n      \"KRAS\",\n      \"MET\",\n      \"ITSN1\",\n      \"PRKD1\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}