{"gene":"SPRY2","run_date":"2026-04-28T20:42:08","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, targeting hSpry2 for degradation by the 26S proteasome. FGF-induced degradation of hSpry2 limits the duration of its inhibitory effect on ERK activation.","method":"Co-immunoprecipitation, ubiquitination assay, proteasome inhibitor treatment, in vitro binding","journal":"Current biology : CB","confidence":"High","confidence_rationale":"Tier 2 — reciprocal Co-IP, functional ubiquitination assay, proteasome inhibitor validation; moderate/strong evidence from single rigorous study","pmids":["12593796"],"is_preprint":false},{"year":2004,"finding":"SPRY2 acts as an inhibitor of ERK signaling in melanocytes and wild-type BRAF melanoma cells by directly binding wild-type BRAF but not the V599E or other exon 15 BRAF mutants; siRNA-mediated knockdown confirmed the ERK inhibitory function.","method":"siRNA knockdown, co-immunoprecipitation/binding assay, ERK phosphorylation assay","journal":"Cancer research","confidence":"High","confidence_rationale":"Tier 2 — direct binding assay with BRAF mutant panel plus functional siRNA knockdown with defined pathway readout","pmids":["15313890"],"is_preprint":false},{"year":2000,"finding":"Human SPRY2 protein binds the intracellular adaptor protein GRB2; a SPRY2/GFP fusion protein remains intracellular, and overexpression of SPRY2 leads to secretion of a soluble factor that inhibits FGF2- but not VEGF-stimulated endothelial cell proliferation, acting non-cell-autonomously.","method":"Co-immunoprecipitation (GRB2 binding), GFP fusion localization, conditioned medium proliferation assay","journal":"Mechanisms of development","confidence":"Medium","confidence_rationale":"Tier 2/3 — GRB2 pulldown and live-cell localization, but non-cell-autonomous mechanism not fully reconstituted","pmids":["10940627"],"is_preprint":false},{"year":2007,"finding":"Tesk1 (testicular protein kinase 1) forms an endogenous complex with Spry2 in cell lines and mouse tissues, relocalizes Spry2 to vesicles/endosomes independently of its kinase activity, abrogates Spry2's interaction with Grb2, and interferes with Spry2 serine dephosphorylation by PP2A, thereby nullifying Spry2's inhibition of ERK phosphorylation and FGF/NGF-induced neurite outgrowth.","method":"Co-immunoprecipitation (endogenous), subcellular fractionation/localization, kinase-dead mutant analysis, siRNA knockdown, neurite outgrowth assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 — reciprocal endogenous Co-IP, multiple orthogonal methods, defined cellular phenotype with mechanism","pmids":["17974561"],"is_preprint":false},{"year":2010,"finding":"In fetal lung epithelial cells, FGF-10 stimulation induces formation of a complex between Spry2, the c-Cbl ubiquitin ligase, and the mTOR repressor TSC2, which abolishes GTPase activity directed against Rheb (the G-protein inducer of mTORC1), linking the FGF-10/FGFR2b/Spry2 pathway to mTORC1 activation and HIF-1α-driven vasculogenesis.","method":"Co-immunoprecipitation (Spry2-cCbl-TSC2 complex), GTPase activity assay, fetal lung explant culture, HIF-1α reporter assay","journal":"American journal of physiology. Lung cellular and molecular physiology","confidence":"Medium","confidence_rationale":"Tier 2/3 — co-IP of endogenous complex with functional GTPase readout; single lab, moderate mechanistic follow-up","pmids":["20622121"],"is_preprint":false},{"year":2010,"finding":"Histone demethylase JMJD1A maintains active H3K9me2 demethylation at the Spry2 promoter; inhibition of JMJD1A by hypoxia or nickel increases H3K9me2 at the Spry2 promoter, represses Spry2 expression, and potentiates ERK phosphorylation and anchorage-independent growth.","method":"ChIP-on-chip (H3K9me2 at Spry2 promoter), Affymetrix GeneChip, forced Spry2 expression, ERK phosphorylation assay, anchorage-independent growth assay","journal":"Carcinogenesis","confidence":"High","confidence_rationale":"Tier 2 — ChIP identifies writer (JMJD1A) at Spry2 locus with functional rescue experiment; multiple orthogonal methods","pmids":["20881000"],"is_preprint":false},{"year":2012,"finding":"SPRY2 loss in prostate cancer cells results in enhanced internalization of EGFR/HER2 via clathrin-mediated endocytosis and their sustained signalling at the early endosome; this involves p38 MAPK activation by PI3K to facilitate ErbB receptor endocytosis, creating a positive feedback loop driving PI3K/AKT-dependent proliferation and invasion in a PTEN-dependent manner.","method":"siRNA/shRNA knockdown, receptor internalization assay, endosome fractionation, pharmacological PI3K/p38 inhibition, in vitro invasion assay, in vivo xenograft","journal":"EMBO molecular medicine","confidence":"High","confidence_rationale":"Tier 2 — multiple orthogonal methods (knockdown, fractionation, pharmacological epistasis, in vivo validation) in single rigorous study","pmids":["22649008"],"is_preprint":false},{"year":2012,"finding":"Inactivation of Spry2 (via dominant-negative Spry2Y55F) accelerates AKT-driven hepatocarcinogenesis in mice by activating MAPK and PKM2 pathways; concomitant suppression of MAPK and PKM2 strongly inhibited the growth induced by Spry2Y55F loss.","method":"Hydrodynamic gene delivery in vivo (mouse liver), dominant-negative Spry2Y55F overexpression, histological/biochemical assays, pharmacological pathway inhibition","journal":"Journal of hepatology","confidence":"High","confidence_rationale":"Tier 2 — in vivo epistasis with dominant-negative mutant plus pharmacological rescue; strong mechanistic evidence","pmids":["22617155"],"is_preprint":false},{"year":2013,"finding":"Intersectin 1 (ITSN1) recruits the Shp2 tyrosine phosphatase to the Cbl-Spry2 complex, enhancing Spry2 dephosphorylation; this disrupts the inhibitory effect of phospho-Spry2 on Cbl and enhances Cbl-mediated EGFR ubiquitylation and lysosomal trafficking. A catalytically inactive Shp2 mutant reverses this effect.","method":"Co-immunoprecipitation, phospho-Spry2 western blot, EGFR ubiquitylation assay, catalytic mutant Shp2 expression, point mutation of Spry2 ITSN1-binding site","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 1/2 — reconstituted ternary complex, mutagenesis at interaction interface, functional ubiquitylation readout","pmids":["24216759"],"is_preprint":false},{"year":2016,"finding":"In adult mouse tracheal basal cells, SPRY2 is post-translationally modified in response to FGFR1 signaling, allowing SPRY2 to inhibit intracellular signaling downstream of other receptor tyrosine kinases and restrain basal cell proliferation; deletion of Fgfr1 or Spry2 each caused increased steady-state basal cell proliferation.","method":"Conditional knockout (Fgfr1 and Spry2) in adult mouse trachea, BrdU proliferation assay, phospho-protein analysis","journal":"Developmental cell","confidence":"High","confidence_rationale":"Tier 2 — clean in vivo KO with defined proliferation phenotype and post-translational modification link; replicated across two gene deletions","pmids":["27046834"],"is_preprint":false},{"year":2017,"finding":"Spry2 is a novel UPR target whose upregulation is PERK-dependent; Spry2 knockdown reduces Serca2 expression, lowers ER calcium levels, and induces the unfolded protein response in pancreatic β-cells. Spry2 deletion in the adult mouse β-cell causes hyperglycemia and hypoinsulinemia.","method":"Whole-genome RNAi screen, PERK pathway analysis, Serca2/calcium measurements, conditional β-cell-specific Spry2 knockout mouse, glucose/insulin measurements","journal":"Diabetes","confidence":"High","confidence_rationale":"Tier 2 — genome-wide unbiased screen confirmed with conditional KO mouse and multiple mechanistic readouts","pmids":["28246293"],"is_preprint":false},{"year":2018,"finding":"SPRY2 physically interacts with MET receptor in rhabdomyosarcoma cells, colocalizing with and binding MET; depletion of SPRY2 leads to MET receptor degradation, reducing migratory and clonogenic potential and inducing differentiation by downregulating ERK/MAPK signaling.","method":"Co-immunoprecipitation, co-localization (immunofluorescence), siRNA knockdown of SPRY2/MET, ERK phosphorylation assay, migration/invasion assay, differentiation assay","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2/3 — reciprocal co-IP with functional validation; single lab, multiple methods","pmids":["29445192"],"is_preprint":false},{"year":2020,"finding":"In glioblastoma, NF-κB pathway activation drives autocrine FGFR signaling, which reactivates ERK; ERK controls SPRY2 transcription, and SPRY2 in turn promotes resistance to EGFR and MET inhibitors. Blocking FGFR prevents ERK reactivation and SPRY2 rebound, overcoming resistance.","method":"Pharmacological inhibition (EGFR, MET, FGFR, NF-κB inhibitors), ERK-bioluminescent reporter in vivo, shRNA knockdown, xenograft mouse model","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 2 — multiple pharmacological epistasis experiments plus in vivo reporter validation; mechanistic pathway placement confirmed","pmids":["32160544"],"is_preprint":false},{"year":2011,"finding":"miR-21 post-transcriptionally downregulates Spry2 protein in glioma cells, disrupting the negative feedback circuit of Ras/MAPK signaling; upregulation of miR-21 is triggered by tumor microenvironmental factors (hyaluronan and growth factors) in PTEN-deficient glioma cells, and loss of Spry2 promotes invasion.","method":"miR-21 overexpression/knockdown, western blot for Spry2 protein vs. mRNA, invasion assay, correlation analysis in human tumor tissues","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 3 — functional knockdown/overexpression with pathway readout but direct 3'UTR targeting not fully validated in this paper","pmids":["21278789"],"is_preprint":false},{"year":2013,"finding":"miR-21 modulates ERK-MAPK signaling during mesenchymal stem cell differentiation by directly repressing SPRY2 expression, increasing the duration and magnitude of ERK-MAPK activity to promote adipogenesis and osteogenesis.","method":"miR-21 overexpression/knockdown in MSCs, western blot for SPRY2 and pERK, adipogenic/osteogenic differentiation assay","journal":"Journal of cellular biochemistry","confidence":"Medium","confidence_rationale":"Tier 3 — functional knockdown with pathway and differentiation readouts; single lab, no direct 3'UTR luciferase assay reported","pmids":["23239100"],"is_preprint":false},{"year":2010,"finding":"Spry2 interacts with Raf-1 but does not directly regulate Raf-1 kinase activity; Spry2 acts as a scaffolding protein facilitating interactions between Raf-1 and its regulators, as shown by lack of correlation between Raf-1 mutant activation state and Spry2 binding strength.","method":"FLAG-tagged Raf-1 phosphorylation site mutants, co-immunoprecipitation, immunofluorescence co-localization, kinase activity assay","journal":"BMB reports","confidence":"Medium","confidence_rationale":"Tier 2 — systematic mutagenesis panel with kinase assay and co-IP; single lab study refining mechanism","pmids":["20356462"],"is_preprint":false},{"year":2023,"finding":"SPRY2 suppresses Y10 phosphorylation of LDHA and LDHA enzymatic activity in fibroblasts by interfering with the interaction between LDHA and the kinase SRC, thereby inhibiting glycolytic metabolism and cancer-associated fibroblast activation.","method":"Co-immunoprecipitation (LDHA-SRC interaction), LDHA kinase activity assay, SPRY2 knockdown in fibroblasts, in vivo tumor growth and metastasis assay","journal":"Breast cancer research : BCR","confidence":"Medium","confidence_rationale":"Tier 2/3 — co-IP of LDHA-SRC disrupted by SPRY2 with functional glycolysis and in vivo readout; single lab","pmids":["37507768"],"is_preprint":false},{"year":2023,"finding":"Protein kinase D (PKD) phosphorylates Spry2 at serine 112 and interacts with the C-terminal half of Spry2 in vivo; phosphorylation at Ser112 promotes Spry2 degradation via the proteasome. The COP9/Signalosome (via CSN3) binds PKD and contributes to Spry2 instability; knockdown of PKD isoforms or CSN3 stabilizes Spry2.","method":"In vitro kinase assay (PKD phosphorylating Spry2), site-directed mutagenesis (Ser112Ala), Co-IP (PKD-Spry2 and CSN3-PKD), protein stability assay (cycloheximide chase), siRNA knockdown","journal":"Oncogenesis","confidence":"High","confidence_rationale":"Tier 1/2 — in vitro kinase assay plus mutagenesis plus in vivo co-IP plus stability assay; single rigorous study with multiple orthogonal methods","pmids":["37045830"],"is_preprint":false},{"year":2008,"finding":"Cerebral ischemia/reperfusion induces Src-mediated upregulation of Spry2 in rat hippocampus in a NMDA receptor-dependent manner, contributing to suppression of the Ras/Raf/MEK/ERK cascade; NMDA receptor antagonist ketamine and Src inhibitor PP2 both blocked ischemia-induced Spry2 upregulation.","method":"Co-immunoprecipitation (Src-Spry2), western blot for Spry2 activity, pharmacological inhibition (ketamine, PP2), four-vessel occlusion ischemia model","journal":"Brain injury","confidence":"Medium","confidence_rationale":"Tier 2/3 — Co-IP and pharmacological epistasis in vivo; single lab, moderate mechanistic characterization","pmids":["18297599"],"is_preprint":false},{"year":2025,"finding":"SPRY2 acts as a K-Ras effector: the SPRY2 C-terminal fragment (residues 161–315) binds oncogenic RasG12V more strongly than full-length SPRY2; both forms localize to the plasma membrane in a K-Ras activity-dependent manner. Mutations at the predicted K-Ras/SPRY2 interface disrupt the interaction. SPRY2 also homo- and hetero-oligomerizes with SPRY4. Active K-Ras is proposed to recruit SPRY2 dimers to the membrane where they block Ras effector access.","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 1/2 — proximity proteomics plus BRET plus Co-IP plus mutagenesis at interface plus functional differentiation readout; single rigorous study with multiple orthogonal methods","pmids":["41362608"],"is_preprint":false},{"year":2024,"finding":"Paclitaxel increases binding of Spry2 to tubulin, sequestering Spry2 away from the plasma membrane; reduced membrane Spry2 relieves its negative regulation of Raf/ERK signaling, upregulating p-c-Raf and p-ERK and aggravating pulmonary fibrosis. Microtubule depolymerizing agents abolish PTX's inhibition of Spry2 membrane distribution.","method":"Co-immunoprecipitation (Spry2-tubulin), immunofluorescence colocalization, plasma membrane fractionation, Spry2 knockdown in fibroblasts, microtubule depolymerization rescue, ERK/Raf phosphorylation assay, in vivo fibrosis model","journal":"The Journal of pharmacology and experimental therapeutics","confidence":"Medium","confidence_rationale":"Tier 2 — co-IP and fractionation with pharmacological rescue; single lab but multiple orthogonal methods","pmids":["37918858"],"is_preprint":false},{"year":2016,"finding":"Homocysteine alters the Spry2/Dyrk1A/PP2A triad in neural progenitor cells: elevated Hcy causes Spry2 promoter demethylation and sustained upregulation of Spry2 mRNA/protein, while Dyrk1A (Spry2-activating kinase) levels decrease and PP2A (Spry2-inactivating phosphatase) levels increase, altering Spry2 phosphorylation status and inhibiting FGFR-ERK-cyclin E-dependent proliferation.","method":"Spry2 promoter methylation analysis, Dyrk1A/PP2A protein expression, Spry2 phosphorylation assay, neural progenitor proliferation assay, methyltransferase inhibitor treatment","journal":"Biochimica et biophysica acta","confidence":"Medium","confidence_rationale":"Tier 2/3 — multi-component pathway analysis with promoter methylation and phosphorylation readouts; single lab","pmids":["27686255"],"is_preprint":false},{"year":2007,"finding":"Spry2-deficient mice develop cleft palate due to excessive cell proliferation and abnormal FGF-responsive transcription factor expression (Etv5, Msx1, Barx1) and disorganized Shh in the palate; Spry2 dosage is critical as hypomorphic BAC transgenic rescue shows intermediate phenotypes, establishing Spry2 as a negative regulator of FGF signaling in palate development.","method":"Spry2 knockout mouse, Spry2-BAC transgene rescue, in situ hybridization, immunohistochemistry, cell proliferation assay","journal":"Mechanisms of development","confidence":"High","confidence_rationale":"Tier 2 — clean KO with defined cellular phenotype, dosage-sensitivity shown by BAC transgene rescue, epistasis with FGF pathway markers","pmids":["17693063"],"is_preprint":false},{"year":2011,"finding":"Spry1 and Spry2 function downstream of FGF signaling in the lens (FGF is necessary and sufficient for their induction) and, via negative modulation of ERK, allow lens vesicle separation from the overlying ectoderm. Combined loss causes persistent keratolenticular stalks with inhibited apoptosis of stalk cells and concomitant ERK activation.","method":"Conditional double knockout (Le-Cre), in situ hybridization, immunohistochemistry, ERK phosphorylation assay, apoptosis assay","journal":"Investigative ophthalmology & visual science","confidence":"High","confidence_rationale":"Tier 2 — clean conditional KO with defined developmental and ERK phenotype; FGF induction confirmed","pmids":["21743007"],"is_preprint":false}],"current_model":"SPRY2 is an inducible, membrane-associated intracellular antagonist of receptor tyrosine kinase (RTK) signaling that, upon growth factor stimulation, is tyrosine-phosphorylated (by Src and other kinases), translocates to membrane ruffles, and inhibits the Ras/Raf/MEK/ERK cascade by binding Grb2, scaffolding Raf-1, and interacting with oncogenic K-Ras at the plasma membrane; it is regulated by c-Cbl–mediated ubiquitination and proteasomal degradation (promoted by EGF/FGF stimulation), by PKD-mediated phosphorylation at Ser112 through the COP9/Signalosome, by Tesk1-dependent relocalization to endosomes (abrogating Grb2 interaction and PP2A-mediated reactivation), and by JMJD1A-controlled H3K9 demethylation at its promoter; in addition, SPRY2 modulates EGFR/ErbB endosomal trafficking via a Cbl-Shp2-intersectin axis, stabilizes MET receptor signaling in rhabdomyosarcoma, suppresses LDHA/SRC-driven glycolysis in fibroblasts, and links FGFR1 signaling to restraint of stem-cell proliferation in the airway epithelium and to ER calcium homeostasis and insulin transcription in pancreatic β-cells."},"narrative":{"teleology":[{"year":2000,"claim":"Identifying SPRY2 as a GRB2-binding intracellular protein that inhibits FGF-driven proliferation established it as a candidate negative regulator of RTK signaling in human cells.","evidence":"Co-immunoprecipitation of SPRY2-GRB2, GFP fusion localization, and conditioned-medium proliferation assay in endothelial cells","pmids":["10940627"],"confidence":"Medium","gaps":["Non-cell-autonomous inhibition mechanism not fully reconstituted","Direct 3′UTR or signaling node target not identified","No in vivo loss-of-function data"]},{"year":2003,"claim":"Demonstrating that c-Cbl ubiquitinates SPRY2 in an EGF-dependent manner and targets it for proteasomal degradation revealed the first post-translational mechanism limiting the duration of SPRY2 activity.","evidence":"Co-IP, ubiquitination assay, and proteasome inhibitor treatment in EGF/FGF-stimulated cells","pmids":["12593796"],"confidence":"High","gaps":["Structural basis of Cbl-SPRY2 interaction unknown","Whether other E3 ligases contribute not tested"]},{"year":2004,"claim":"Showing that SPRY2 binds wild-type BRAF but not oncogenic V599E BRAF to inhibit ERK signaling pinpointed the Raf level of the cascade as a direct node of SPRY2 action and explained why mutant-BRAF tumors escape SPRY2 inhibition.","evidence":"Co-IP/binding assay with BRAF mutant panel and siRNA knockdown with ERK phosphorylation readout in melanocytes","pmids":["15313890"],"confidence":"High","gaps":["SPRY2-BRAF binding interface not mapped","Whether SPRY2 affects BRAF dimerization not addressed"]},{"year":2007,"claim":"Two studies resolved distinct regulatory layers: Tesk1 relocalizes SPRY2 to endosomes to abrogate GRB2 binding and PP2A-mediated reactivation, while Spry2-knockout mice develop cleft palate from excess FGF-driven proliferation, confirming SPRY2 as an essential dosage-sensitive negative-feedback regulator of FGF signaling in vivo.","evidence":"Endogenous co-IP, kinase-dead mutant analysis, and neurite outgrowth assay (Tesk1); Spry2 KO and BAC-transgene rescue with proliferation and FGF-target gene analysis (palate)","pmids":["17974561","17693063"],"confidence":"High","gaps":["Tesk1 interaction domain on SPRY2 not mapped","Downstream FGF target selectivity in palate not fully dissected"]},{"year":2010,"claim":"Multiple discoveries expanded the SPRY2 mechanism: SPRY2 scaffolds Raf-1 without directly regulating its kinase activity, a Spry2-cCbl-TSC2 complex links FGF-10 signaling to mTORC1 activation in fetal lung, and JMJD1A-dependent H3K9 demethylation at the SPRY2 promoter was identified as a transcriptional on-switch.","evidence":"Raf-1 phosphorylation-site mutant panel with Co-IP and kinase assay; Spry2-cCbl-TSC2 Co-IP and GTPase assay in lung explants; ChIP-on-chip for H3K9me2 at SPRY2 promoter with functional rescue","pmids":["20356462","20622121","20881000"],"confidence":"Medium","gaps":["Raf-1 scaffolding model lacks structural detail","TSC2-SPRY2 interaction stoichiometry unclear","JMJD1A regulation confirmed only in nickel/hypoxia context"]},{"year":2011,"claim":"miR-21 was identified as a post-transcriptional repressor of SPRY2 in glioma and mesenchymal stem cells, revealing how the SPRY2 feedback loop can be hijacked to sustain ERK signaling in tumor and differentiation contexts.","evidence":"miR-21 overexpression/knockdown with SPRY2 protein quantification, invasion assay in glioma, and differentiation assay in MSCs","pmids":["21278789","23239100"],"confidence":"Medium","gaps":["Direct 3′UTR luciferase validation not provided in these studies","Other miRNAs targeting SPRY2 not systematically evaluated"]},{"year":2011,"claim":"Conditional double knockout of Spry1/Spry2 in the lens demonstrated that FGF-induced Sprouty proteins restrain ERK to permit lens vesicle separation, establishing a non-redundant developmental requirement.","evidence":"Le-Cre conditional double KO with ERK phosphorylation and apoptosis assays","pmids":["21743007"],"confidence":"High","gaps":["Individual contributions of Spry1 vs. Spry2 in this tissue not fully resolved"]},{"year":2012,"claim":"Two studies placed SPRY2 at the intersection of RTK trafficking and cancer: SPRY2 loss enhanced EGFR/HER2 endosomal signaling and PI3K/AKT-driven invasion in prostate cancer, while dominant-negative SPRY2Y55F accelerated AKT-driven hepatocarcinogenesis via MAPK and PKM2.","evidence":"siRNA/shRNA with receptor internalization and endosome fractionation (prostate); hydrodynamic gene delivery of Spry2Y55F in mouse liver with pharmacological rescue (liver)","pmids":["22649008","22617155"],"confidence":"High","gaps":["Whether SPRY2 directly contacts endosomal machinery not shown","PKM2 regulation mechanism downstream of SPRY2 not dissected"]},{"year":2013,"claim":"Reconstitution of an ITSN1–Shp2–Cbl–SPRY2 complex showed that Shp2-mediated dephosphorylation of SPRY2 relieves its sequestration of Cbl, thereby promoting EGFR ubiquitylation and lysosomal degradation—mechanistically connecting SPRY2 phosphorylation status to receptor downregulation.","evidence":"Co-IP of quaternary complex, catalytic-dead Shp2 mutant, Spry2 ITSN1-binding site mutant, EGFR ubiquitylation assay","pmids":["24216759"],"confidence":"High","gaps":["In vivo relevance of ITSN1-Shp2 axis not validated","Whether other phosphatases substitute in ITSN1-null cells unknown"]},{"year":2016,"claim":"Conditional knockout studies in tracheal basal cells and neural progenitors revealed that FGFR1-dependent post-translational modification of SPRY2 restrains stem-cell proliferation, and that homocysteine disrupts the Spry2/Dyrk1A/PP2A triad to alter SPRY2 phosphorylation and neural progenitor proliferation.","evidence":"Fgfr1 and Spry2 conditional KO in adult mouse trachea with BrdU assay; Spry2 promoter methylation, Dyrk1A/PP2A expression, and proliferation assays in neural progenitors","pmids":["27046834","27686255"],"confidence":"High","gaps":["Identity of FGFR1-induced post-translational modification not determined","Dyrk1A phosphorylation site on SPRY2 not mapped"]},{"year":2017,"claim":"Discovery that SPRY2 is a PERK-dependent UPR target in β-cells, required for SERCA2 expression and ER calcium homeostasis, expanded its function beyond RTK-ERK feedback to organelle stress management and insulin secretion.","evidence":"Whole-genome RNAi screen, PERK pathway analysis, SERCA2/calcium measurements, conditional β-cell-specific Spry2 KO mouse","pmids":["28246293"],"confidence":"High","gaps":["Mechanism by which SPRY2 maintains SERCA2 levels unknown","Whether SPRY2-ER calcium link exists outside β-cells not tested"]},{"year":2018,"claim":"SPRY2 was found to physically interact with and stabilize the MET receptor in rhabdomyosarcoma, demonstrating a context where SPRY2 sustains rather than inhibits RTK signaling.","evidence":"Co-IP and colocalization of SPRY2-MET, siRNA knockdown leading to MET degradation, ERK and differentiation assays","pmids":["29445192"],"confidence":"Medium","gaps":["Mechanism of SPRY2-mediated MET stabilization not defined","Whether this is unique to rhabdomyosarcoma or generalizable unclear"]},{"year":2020,"claim":"In glioblastoma, NF-κB–driven autocrine FGFR signaling was shown to reactivate ERK, which induces SPRY2 transcription; paradoxically, SPRY2 itself promotes resistance to EGFR/MET inhibitors, revealing a context-dependent pro-survival role.","evidence":"Pharmacological epistasis (EGFR, MET, FGFR, NF-κB inhibitors), ERK-bioluminescent reporter, shRNA knockdown, xenograft model","pmids":["32160544"],"confidence":"High","gaps":["Mechanism by which SPRY2 promotes drug resistance not defined at the molecular level","Generalizability across GBM subtypes not tested"]},{"year":2023,"claim":"Two studies uncovered novel regulatory inputs: PKD phosphorylates SPRY2 at Ser112 cooperating with the COP9 signalosome to promote proteasomal turnover, and SPRY2 suppresses SRC-mediated LDHA Y10 phosphorylation to inhibit glycolysis in fibroblasts.","evidence":"In vitro kinase assay, Ser112Ala mutagenesis, CSN3 co-IP and stability assay (PKD); co-IP of LDHA-SRC disrupted by SPRY2, LDHA activity assay, in vivo tumor/metastasis model (LDHA)","pmids":["37045830","37507768"],"confidence":"High","gaps":["Whether PKD-Ser112 and c-Cbl ubiquitination are sequential or parallel degradation pathways unknown","SPRY2-SRC interaction interface not mapped"]},{"year":2024,"claim":"Paclitaxel was shown to sequester SPRY2 on tubulin, depleting it from the plasma membrane and thereby de-repressing Raf/ERK signaling—a drug-induced redistribution mechanism aggravating pulmonary fibrosis.","evidence":"Co-IP of SPRY2-tubulin, membrane fractionation, microtubule depolymerization rescue, in vivo fibrosis model","pmids":["37918858"],"confidence":"Medium","gaps":["Tubulin-binding domain on SPRY2 not identified","Whether this mechanism applies to other microtubule-targeting agents not tested"]},{"year":2025,"claim":"Proximity proteomics and BRET studies established that SPRY2 is a direct K-Ras effector: the C-terminal fragment binds active K-Ras at the plasma membrane, homo/hetero-oligomerizes with SPRY4, and blocks Ras effector access, providing the first structural-level model of SPRY2 inhibition at the Ras node.","evidence":"TurboID proximity proteomics, BRET assay, interface mutagenesis, K-Ras membrane anchorage inhibitors, C2C12 differentiation assay","pmids":["41362608"],"confidence":"High","gaps":["Full atomic-resolution structure of K-Ras–SPRY2 complex not yet available","Stoichiometry and dynamics of SPRY2 oligomers at the membrane not determined"]},{"year":null,"claim":"Key unresolved questions include the structural basis of SPRY2 interactions with K-Ras/Raf/GRB2, how SPRY2 switches between tumor-suppressive and pro-survival roles in different cancer contexts, and the mechanism linking SPRY2 to ER calcium homeostasis and SERCA2 expression.","evidence":"","pmids":[],"confidence":"Low","gaps":["No atomic-resolution structure of any SPRY2 signaling complex","Molecular determinants of context-dependent pro-tumorigenic vs. tumor-suppressive function uncharacterized","SPRY2-SERCA2 regulatory mechanism undefined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[1,3,6,7,9,15,19]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[2,8,15]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[19,20]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[2]},{"term_id":"GO:0005768","term_label":"endosome","supporting_discovery_ids":[3,6]},{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[20]}],"pathway":[{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[1,2,3,6,7,8,9,12,15,19]},{"term_id":"R-HSA-1266738","term_label":"Developmental Biology","supporting_discovery_ids":[22,23]},{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[0,8,17]},{"term_id":"R-HSA-1430728","term_label":"Metabolism","supporting_discovery_ids":[16]}],"complexes":[],"partners":["GRB2","BRAF","CBL","KRAS","TESK1","ITSN1","MET","RAF1"],"other_free_text":[]},"mechanistic_narrative":"SPRY2 is an inducible intracellular antagonist of receptor tyrosine kinase (RTK) signaling that constrains the Ras/Raf/MEK/ERK cascade across diverse developmental and homeostatic contexts. Upon growth factor stimulation, SPRY2 is tyrosine-phosphorylated, translocates to the plasma membrane, and suppresses ERK activation by binding GRB2, scaffolding Raf-1, and interacting directly with oncogenic K-Ras to block effector access [PMID:10940627, PMID:15313890, PMID:20356462, PMID:41362608]. SPRY2 abundance and activity are tightly regulated: c-Cbl–mediated ubiquitination targets it for proteasomal degradation upon EGF/FGF stimulation, PKD phosphorylation at Ser112 cooperates with the COP9 signalosome to promote its turnover, Tesk1 relocalizes it to endosomes abrogating GRB2 binding, and an intersectin-1/Shp2 axis dephosphorylates SPRY2 to relieve its inhibition of Cbl-dependent EGFR ubiquitylation and lysosomal trafficking [PMID:12593796, PMID:37045830, PMID:17974561, PMID:24216759]. Beyond canonical RTK-ERK feedback, SPRY2 modulates EGFR/HER2 endosomal signaling in prostate cancer, stabilizes MET receptor in rhabdomyosarcoma, suppresses SRC-dependent LDHA phosphorylation and glycolysis in fibroblasts, maintains ER calcium homeostasis and insulin secretion in pancreatic β-cells, and is required for palatal closure during development [PMID:22649008, PMID:29445192, PMID:37507768, PMID:28246293, PMID:17693063]."},"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). Thereby acts as an antagonist of FGF-induced retinal lens fiber differentiation, may inhibit limb bud outgrowth and may negatively modulate respiratory organogenesis (By similarity). Inhibits TGFB-induced epithelial-to-mesenchymal transition in retinal lens epithelial cells (By similarity). 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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 cellular and molecular immunology","url":"https://pubmed.ncbi.nlm.nih.gov/24796743","citation_count":0,"is_preprint":false},{"pmid":"41021275","id":"PMC_41021275","title":"Functional Variant Discovery Identifies a Novel Genetic Link Between SPRY2, Wood Smoke, and Asthma.","date":"2025","source":"American journal of respiratory cell and molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/41021275","citation_count":0,"is_preprint":false},{"pmid":"41362608","id":"PMC_41362608","title":"Proteomics- and BRET- screens identify SPRY2 as a Ras effector that impacts its membrane organization.","date":"2025","source":"iScience","url":"https://pubmed.ncbi.nlm.nih.gov/41362608","citation_count":0,"is_preprint":false},{"pmid":"41480712","id":"PMC_41480712","title":"A Novel lncRNA MSTRG.16386.1 as ceRNA Promotes Kidney Injury Induced by Deep Hypothermic Circulatory Arrest via Mediating miR-466b-5p/Spry2 Axis.","date":"2026","source":"FASEB journal : official publication of the Federation of American Societies for Experimental Biology","url":"https://pubmed.ncbi.nlm.nih.gov/41480712","citation_count":0,"is_preprint":false},{"pmid":"28274312","id":"PMC_28274312","title":"[Establishment of a human bladder cancer cell line stably co-expressing hSPRY2 and luciferase genes and its subcutaneous tumor xenograft model in nude mice].","date":"2017","source":"Xi bao yu fen zi mian yi xue za zhi = Chinese journal of cellular and molecular immunology","url":"https://pubmed.ncbi.nlm.nih.gov/28274312","citation_count":0,"is_preprint":false},{"pmid":"41212736","id":"PMC_41212736","title":"Investigating SPRY2 Single-Nucleotide Polymorphisms in the Etiology of Nonsyndromic Orofacial Cleft in the Chinese Han Population.","date":"2025","source":"The Cleft palate-craniofacial journal : official publication of the American Cleft Palate-Craniofacial Association","url":"https://pubmed.ncbi.nlm.nih.gov/41212736","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.11.19.689292","title":"Transcriptional feedback of Erk signaling waves in zebrafish scale regeneration","date":"2025-11-19","source":"bioRxiv","url":"https://doi.org/10.1101/2025.11.19.689292","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2025.06.13.659437","title":"Proteomics- and BRET-screens identify SPRY2 as Ras effector that impacts its membrane organisation","date":"2025-06-13","source":"bioRxiv","url":"https://doi.org/10.1101/2025.06.13.659437","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2025.02.09.25321494","title":"Genetic-epigenetic interactions (meQTLs) in orofacial clefts etiology","date":"2025-02-12","source":"bioRxiv","url":"https://doi.org/10.1101/2025.02.09.25321494","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2025.10.13.682042","title":"Genetic dissection of tissue composition in genetically diverse mouse populations","date":"2025-10-14","source":"bioRxiv","url":"https://doi.org/10.1101/2025.10.13.682042","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2024.11.25.24317869","title":"Genomic predictors of response to antidepressants in Major depressive disorder (MDD): A GWAS-Based Study on Indian cohort","date":"2024-11-26","source":"bioRxiv","url":"https://doi.org/10.1101/2024.11.25.24317869","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":46273,"output_tokens":6464,"usd":0.117889},"stage2":{"model":"claude-opus-4-6","input_tokens":10060,"output_tokens":4498,"usd":0.244125},"total_usd":0.362014,"stage1_batch_id":"msgbatch_011rkcQdVV1RaFq9XhcG6df6","stage2_batch_id":"msgbatch_01RzgrJafdF4EWHhj5mNi6dh","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2003,\n      \"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, targeting hSpry2 for degradation by the 26S proteasome. FGF-induced degradation of hSpry2 limits the duration of its inhibitory effect on ERK activation.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, proteasome inhibitor treatment, in vitro binding\",\n      \"journal\": \"Current biology : CB\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — reciprocal Co-IP, functional ubiquitination assay, proteasome inhibitor validation; moderate/strong evidence from single rigorous study\",\n      \"pmids\": [\"12593796\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"SPRY2 acts as an inhibitor of ERK signaling in melanocytes and wild-type BRAF melanoma cells by directly binding wild-type BRAF but not the V599E or other exon 15 BRAF mutants; siRNA-mediated knockdown confirmed the ERK inhibitory function.\",\n      \"method\": \"siRNA knockdown, co-immunoprecipitation/binding assay, ERK phosphorylation assay\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — direct binding assay with BRAF mutant panel plus functional siRNA knockdown with defined pathway readout\",\n      \"pmids\": [\"15313890\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"Human SPRY2 protein binds the intracellular adaptor protein GRB2; a SPRY2/GFP fusion protein remains intracellular, and overexpression of SPRY2 leads to secretion of a soluble factor that inhibits FGF2- but not VEGF-stimulated endothelial cell proliferation, acting non-cell-autonomously.\",\n      \"method\": \"Co-immunoprecipitation (GRB2 binding), GFP fusion localization, conditioned medium proliferation assay\",\n      \"journal\": \"Mechanisms of development\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2/3 — GRB2 pulldown and live-cell localization, but non-cell-autonomous mechanism not fully reconstituted\",\n      \"pmids\": [\"10940627\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Tesk1 (testicular protein kinase 1) forms an endogenous complex with Spry2 in cell lines and mouse tissues, relocalizes Spry2 to vesicles/endosomes independently of its kinase activity, abrogates Spry2's interaction with Grb2, and interferes with Spry2 serine dephosphorylation by PP2A, thereby nullifying Spry2's inhibition of ERK phosphorylation and FGF/NGF-induced neurite outgrowth.\",\n      \"method\": \"Co-immunoprecipitation (endogenous), subcellular fractionation/localization, kinase-dead mutant analysis, siRNA knockdown, neurite outgrowth assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — reciprocal endogenous Co-IP, multiple orthogonal methods, defined cellular phenotype with mechanism\",\n      \"pmids\": [\"17974561\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"In fetal lung epithelial cells, FGF-10 stimulation induces formation of a complex between Spry2, the c-Cbl ubiquitin ligase, and the mTOR repressor TSC2, which abolishes GTPase activity directed against Rheb (the G-protein inducer of mTORC1), linking the FGF-10/FGFR2b/Spry2 pathway to mTORC1 activation and HIF-1α-driven vasculogenesis.\",\n      \"method\": \"Co-immunoprecipitation (Spry2-cCbl-TSC2 complex), GTPase activity assay, fetal lung explant culture, HIF-1α reporter assay\",\n      \"journal\": \"American journal of physiology. Lung cellular and molecular physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2/3 — co-IP of endogenous complex with functional GTPase readout; single lab, moderate mechanistic follow-up\",\n      \"pmids\": [\"20622121\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Histone demethylase JMJD1A maintains active H3K9me2 demethylation at the Spry2 promoter; inhibition of JMJD1A by hypoxia or nickel increases H3K9me2 at the Spry2 promoter, represses Spry2 expression, and potentiates ERK phosphorylation and anchorage-independent growth.\",\n      \"method\": \"ChIP-on-chip (H3K9me2 at Spry2 promoter), Affymetrix GeneChip, forced Spry2 expression, ERK phosphorylation assay, anchorage-independent growth assay\",\n      \"journal\": \"Carcinogenesis\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — ChIP identifies writer (JMJD1A) at Spry2 locus with functional rescue experiment; multiple orthogonal methods\",\n      \"pmids\": [\"20881000\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"SPRY2 loss in prostate cancer cells results in enhanced internalization of EGFR/HER2 via clathrin-mediated endocytosis and their sustained signalling at the early endosome; this involves p38 MAPK activation by PI3K to facilitate ErbB receptor endocytosis, creating a positive feedback loop driving PI3K/AKT-dependent proliferation and invasion in a PTEN-dependent manner.\",\n      \"method\": \"siRNA/shRNA knockdown, receptor internalization assay, endosome fractionation, pharmacological PI3K/p38 inhibition, in vitro invasion assay, in vivo xenograft\",\n      \"journal\": \"EMBO molecular medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — multiple orthogonal methods (knockdown, fractionation, pharmacological epistasis, in vivo validation) in single rigorous study\",\n      \"pmids\": [\"22649008\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Inactivation of Spry2 (via dominant-negative Spry2Y55F) accelerates AKT-driven hepatocarcinogenesis in mice by activating MAPK and PKM2 pathways; concomitant suppression of MAPK and PKM2 strongly inhibited the growth induced by Spry2Y55F loss.\",\n      \"method\": \"Hydrodynamic gene delivery in vivo (mouse liver), dominant-negative Spry2Y55F overexpression, histological/biochemical assays, pharmacological pathway inhibition\",\n      \"journal\": \"Journal of hepatology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — in vivo epistasis with dominant-negative mutant plus pharmacological rescue; strong mechanistic evidence\",\n      \"pmids\": [\"22617155\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Intersectin 1 (ITSN1) recruits the Shp2 tyrosine phosphatase to the Cbl-Spry2 complex, enhancing Spry2 dephosphorylation; this disrupts the inhibitory effect of phospho-Spry2 on Cbl and enhances Cbl-mediated EGFR ubiquitylation and lysosomal trafficking. A catalytically inactive Shp2 mutant reverses this effect.\",\n      \"method\": \"Co-immunoprecipitation, phospho-Spry2 western blot, EGFR ubiquitylation assay, catalytic mutant Shp2 expression, point mutation of Spry2 ITSN1-binding site\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1/2 — reconstituted ternary complex, mutagenesis at interaction interface, functional ubiquitylation readout\",\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 in response to FGFR1 signaling, allowing SPRY2 to inhibit intracellular signaling downstream of other receptor tyrosine kinases and restrain basal cell proliferation; deletion of Fgfr1 or Spry2 each caused increased steady-state basal cell proliferation.\",\n      \"method\": \"Conditional knockout (Fgfr1 and Spry2) in adult mouse trachea, BrdU proliferation assay, phospho-protein analysis\",\n      \"journal\": \"Developmental cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — clean in vivo KO with defined proliferation phenotype and post-translational modification link; replicated across two gene deletions\",\n      \"pmids\": [\"27046834\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Spry2 is a novel UPR target whose upregulation is PERK-dependent; Spry2 knockdown reduces Serca2 expression, lowers ER calcium levels, and induces the unfolded protein response in pancreatic β-cells. Spry2 deletion in the adult mouse β-cell causes hyperglycemia and hypoinsulinemia.\",\n      \"method\": \"Whole-genome RNAi screen, PERK pathway analysis, Serca2/calcium measurements, conditional β-cell-specific Spry2 knockout mouse, glucose/insulin measurements\",\n      \"journal\": \"Diabetes\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — genome-wide unbiased screen confirmed with conditional KO mouse and multiple mechanistic readouts\",\n      \"pmids\": [\"28246293\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"SPRY2 physically interacts with MET receptor in rhabdomyosarcoma cells, colocalizing with and binding MET; depletion of SPRY2 leads to MET receptor degradation, reducing migratory and clonogenic potential and inducing differentiation by downregulating ERK/MAPK signaling.\",\n      \"method\": \"Co-immunoprecipitation, co-localization (immunofluorescence), siRNA knockdown of SPRY2/MET, ERK phosphorylation assay, migration/invasion assay, differentiation assay\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2/3 — reciprocal co-IP with functional validation; single lab, multiple methods\",\n      \"pmids\": [\"29445192\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"In glioblastoma, NF-κB pathway activation drives autocrine FGFR signaling, which reactivates ERK; ERK controls SPRY2 transcription, and SPRY2 in turn promotes resistance to EGFR and MET inhibitors. Blocking FGFR prevents ERK reactivation and SPRY2 rebound, overcoming resistance.\",\n      \"method\": \"Pharmacological inhibition (EGFR, MET, FGFR, NF-κB inhibitors), ERK-bioluminescent reporter in vivo, shRNA knockdown, xenograft mouse model\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — multiple pharmacological epistasis experiments plus in vivo reporter validation; mechanistic pathway placement confirmed\",\n      \"pmids\": [\"32160544\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"miR-21 post-transcriptionally downregulates Spry2 protein in glioma cells, disrupting the negative feedback circuit of Ras/MAPK signaling; upregulation of miR-21 is triggered by tumor microenvironmental factors (hyaluronan and growth factors) in PTEN-deficient glioma cells, and loss of Spry2 promotes invasion.\",\n      \"method\": \"miR-21 overexpression/knockdown, western blot for Spry2 protein vs. mRNA, invasion assay, correlation analysis in human tumor tissues\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — functional knockdown/overexpression with pathway readout but direct 3'UTR targeting not fully validated in this paper\",\n      \"pmids\": [\"21278789\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"miR-21 modulates ERK-MAPK signaling during mesenchymal stem cell differentiation by directly repressing SPRY2 expression, increasing the duration and magnitude of ERK-MAPK activity to promote adipogenesis and osteogenesis.\",\n      \"method\": \"miR-21 overexpression/knockdown in MSCs, western blot for SPRY2 and pERK, adipogenic/osteogenic differentiation assay\",\n      \"journal\": \"Journal of cellular biochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — functional knockdown with pathway and differentiation readouts; single lab, no direct 3'UTR luciferase assay reported\",\n      \"pmids\": [\"23239100\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Spry2 interacts with Raf-1 but does not directly regulate Raf-1 kinase activity; Spry2 acts as a scaffolding protein facilitating interactions between Raf-1 and its regulators, as shown by lack of correlation between Raf-1 mutant activation state and Spry2 binding strength.\",\n      \"method\": \"FLAG-tagged Raf-1 phosphorylation site mutants, co-immunoprecipitation, immunofluorescence co-localization, kinase activity assay\",\n      \"journal\": \"BMB reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — systematic mutagenesis panel with kinase assay and co-IP; single lab study refining mechanism\",\n      \"pmids\": [\"20356462\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"SPRY2 suppresses Y10 phosphorylation of LDHA and LDHA enzymatic activity in fibroblasts by interfering with the interaction between LDHA and the kinase SRC, thereby inhibiting glycolytic metabolism and cancer-associated fibroblast activation.\",\n      \"method\": \"Co-immunoprecipitation (LDHA-SRC interaction), LDHA kinase activity assay, SPRY2 knockdown in fibroblasts, in vivo tumor growth and metastasis assay\",\n      \"journal\": \"Breast cancer research : BCR\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2/3 — co-IP of LDHA-SRC disrupted by SPRY2 with functional glycolysis and in vivo readout; single lab\",\n      \"pmids\": [\"37507768\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Protein kinase D (PKD) phosphorylates Spry2 at serine 112 and interacts with the C-terminal half of Spry2 in vivo; phosphorylation at Ser112 promotes Spry2 degradation via the proteasome. The COP9/Signalosome (via CSN3) binds PKD and contributes to Spry2 instability; knockdown of PKD isoforms or CSN3 stabilizes Spry2.\",\n      \"method\": \"In vitro kinase assay (PKD phosphorylating Spry2), site-directed mutagenesis (Ser112Ala), Co-IP (PKD-Spry2 and CSN3-PKD), protein stability assay (cycloheximide chase), siRNA knockdown\",\n      \"journal\": \"Oncogenesis\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1/2 — in vitro kinase assay plus mutagenesis plus in vivo co-IP plus stability assay; single rigorous study with multiple orthogonal methods\",\n      \"pmids\": [\"37045830\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Cerebral ischemia/reperfusion induces Src-mediated upregulation of Spry2 in rat hippocampus in a NMDA receptor-dependent manner, contributing to suppression of the Ras/Raf/MEK/ERK cascade; NMDA receptor antagonist ketamine and Src inhibitor PP2 both blocked ischemia-induced Spry2 upregulation.\",\n      \"method\": \"Co-immunoprecipitation (Src-Spry2), western blot for Spry2 activity, pharmacological inhibition (ketamine, PP2), four-vessel occlusion ischemia model\",\n      \"journal\": \"Brain injury\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2/3 — Co-IP and pharmacological epistasis in vivo; single lab, moderate mechanistic characterization\",\n      \"pmids\": [\"18297599\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"SPRY2 acts as a K-Ras effector: the SPRY2 C-terminal fragment (residues 161–315) binds oncogenic RasG12V more strongly than full-length SPRY2; both forms localize to the plasma membrane in a K-Ras activity-dependent manner. Mutations at the predicted K-Ras/SPRY2 interface disrupt the interaction. SPRY2 also homo- and hetero-oligomerizes with SPRY4. Active K-Ras is proposed to recruit SPRY2 dimers to the membrane where they block Ras effector access.\",\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 1/2 — proximity proteomics plus BRET plus Co-IP plus mutagenesis at interface plus functional differentiation readout; single rigorous study with multiple orthogonal methods\",\n      \"pmids\": [\"41362608\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Paclitaxel increases binding of Spry2 to tubulin, sequestering Spry2 away from the plasma membrane; reduced membrane Spry2 relieves its negative regulation of Raf/ERK signaling, upregulating p-c-Raf and p-ERK and aggravating pulmonary fibrosis. Microtubule depolymerizing agents abolish PTX's inhibition of Spry2 membrane distribution.\",\n      \"method\": \"Co-immunoprecipitation (Spry2-tubulin), immunofluorescence colocalization, plasma membrane fractionation, Spry2 knockdown in fibroblasts, microtubule depolymerization rescue, ERK/Raf phosphorylation assay, in vivo fibrosis model\",\n      \"journal\": \"The Journal of pharmacology and experimental therapeutics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — co-IP and fractionation with pharmacological rescue; single lab but multiple orthogonal methods\",\n      \"pmids\": [\"37918858\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Homocysteine alters the Spry2/Dyrk1A/PP2A triad in neural progenitor cells: elevated Hcy causes Spry2 promoter demethylation and sustained upregulation of Spry2 mRNA/protein, while Dyrk1A (Spry2-activating kinase) levels decrease and PP2A (Spry2-inactivating phosphatase) levels increase, altering Spry2 phosphorylation status and inhibiting FGFR-ERK-cyclin E-dependent proliferation.\",\n      \"method\": \"Spry2 promoter methylation analysis, Dyrk1A/PP2A protein expression, Spry2 phosphorylation assay, neural progenitor proliferation assay, methyltransferase inhibitor treatment\",\n      \"journal\": \"Biochimica et biophysica acta\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2/3 — multi-component pathway analysis with promoter methylation and phosphorylation readouts; single lab\",\n      \"pmids\": [\"27686255\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Spry2-deficient mice develop cleft palate due to excessive cell proliferation and abnormal FGF-responsive transcription factor expression (Etv5, Msx1, Barx1) and disorganized Shh in the palate; Spry2 dosage is critical as hypomorphic BAC transgenic rescue shows intermediate phenotypes, establishing Spry2 as a negative regulator of FGF signaling in palate development.\",\n      \"method\": \"Spry2 knockout mouse, Spry2-BAC transgene rescue, in situ hybridization, immunohistochemistry, cell proliferation assay\",\n      \"journal\": \"Mechanisms of development\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — clean KO with defined cellular phenotype, dosage-sensitivity shown by BAC transgene rescue, epistasis with FGF pathway markers\",\n      \"pmids\": [\"17693063\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Spry1 and Spry2 function downstream of FGF signaling in the lens (FGF is necessary and sufficient for their induction) and, via negative modulation of ERK, allow lens vesicle separation from the overlying ectoderm. Combined loss causes persistent keratolenticular stalks with inhibited apoptosis of stalk cells and concomitant ERK activation.\",\n      \"method\": \"Conditional double knockout (Le-Cre), in situ hybridization, immunohistochemistry, ERK phosphorylation assay, apoptosis assay\",\n      \"journal\": \"Investigative ophthalmology & visual science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — clean conditional KO with defined developmental and ERK phenotype; FGF induction confirmed\",\n      \"pmids\": [\"21743007\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"SPRY2 is an inducible, membrane-associated intracellular antagonist of receptor tyrosine kinase (RTK) signaling that, upon growth factor stimulation, is tyrosine-phosphorylated (by Src and other kinases), translocates to membrane ruffles, and inhibits the Ras/Raf/MEK/ERK cascade by binding Grb2, scaffolding Raf-1, and interacting with oncogenic K-Ras at the plasma membrane; it is regulated by c-Cbl–mediated ubiquitination and proteasomal degradation (promoted by EGF/FGF stimulation), by PKD-mediated phosphorylation at Ser112 through the COP9/Signalosome, by Tesk1-dependent relocalization to endosomes (abrogating Grb2 interaction and PP2A-mediated reactivation), and by JMJD1A-controlled H3K9 demethylation at its promoter; in addition, SPRY2 modulates EGFR/ErbB endosomal trafficking via a Cbl-Shp2-intersectin axis, stabilizes MET receptor signaling in rhabdomyosarcoma, suppresses LDHA/SRC-driven glycolysis in fibroblasts, and links FGFR1 signaling to restraint of stem-cell proliferation in the airway epithelium and to ER calcium homeostasis and insulin transcription in pancreatic β-cells.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"SPRY2 is an inducible intracellular antagonist of receptor tyrosine kinase (RTK) signaling that constrains the Ras/Raf/MEK/ERK cascade across diverse developmental and homeostatic contexts. Upon growth factor stimulation, SPRY2 is tyrosine-phosphorylated, translocates to the plasma membrane, and suppresses ERK activation by binding GRB2, scaffolding Raf-1, and interacting directly with oncogenic K-Ras to block effector access [PMID:10940627, PMID:15313890, PMID:20356462, PMID:41362608]. SPRY2 abundance and activity are tightly regulated: c-Cbl–mediated ubiquitination targets it for proteasomal degradation upon EGF/FGF stimulation, PKD phosphorylation at Ser112 cooperates with the COP9 signalosome to promote its turnover, Tesk1 relocalizes it to endosomes abrogating GRB2 binding, and an intersectin-1/Shp2 axis dephosphorylates SPRY2 to relieve its inhibition of Cbl-dependent EGFR ubiquitylation and lysosomal trafficking [PMID:12593796, PMID:37045830, PMID:17974561, PMID:24216759]. Beyond canonical RTK-ERK feedback, SPRY2 modulates EGFR/HER2 endosomal signaling in prostate cancer, stabilizes MET receptor in rhabdomyosarcoma, suppresses SRC-dependent LDHA phosphorylation and glycolysis in fibroblasts, maintains ER calcium homeostasis and insulin secretion in pancreatic β-cells, and is required for palatal closure during development [PMID:22649008, PMID:29445192, PMID:37507768, PMID:28246293, PMID:17693063].\",\n  \"teleology\": [\n    {\n      \"year\": 2000,\n      \"claim\": \"Identifying SPRY2 as a GRB2-binding intracellular protein that inhibits FGF-driven proliferation established it as a candidate negative regulator of RTK signaling in human cells.\",\n      \"evidence\": \"Co-immunoprecipitation of SPRY2-GRB2, GFP fusion localization, and conditioned-medium proliferation assay in endothelial cells\",\n      \"pmids\": [\"10940627\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Non-cell-autonomous inhibition mechanism not fully reconstituted\", \"Direct 3′UTR or signaling node target not identified\", \"No in vivo loss-of-function data\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Demonstrating that c-Cbl ubiquitinates SPRY2 in an EGF-dependent manner and targets it for proteasomal degradation revealed the first post-translational mechanism limiting the duration of SPRY2 activity.\",\n      \"evidence\": \"Co-IP, ubiquitination assay, and proteasome inhibitor treatment in EGF/FGF-stimulated cells\",\n      \"pmids\": [\"12593796\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of Cbl-SPRY2 interaction unknown\", \"Whether other E3 ligases contribute not tested\"]\n    },\n    {\n      \"year\": 2004,\n      \"claim\": \"Showing that SPRY2 binds wild-type BRAF but not oncogenic V599E BRAF to inhibit ERK signaling pinpointed the Raf level of the cascade as a direct node of SPRY2 action and explained why mutant-BRAF tumors escape SPRY2 inhibition.\",\n      \"evidence\": \"Co-IP/binding assay with BRAF mutant panel and siRNA knockdown with ERK phosphorylation readout in melanocytes\",\n      \"pmids\": [\"15313890\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"SPRY2-BRAF binding interface not mapped\", \"Whether SPRY2 affects BRAF dimerization not addressed\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Two studies resolved distinct regulatory layers: Tesk1 relocalizes SPRY2 to endosomes to abrogate GRB2 binding and PP2A-mediated reactivation, while Spry2-knockout mice develop cleft palate from excess FGF-driven proliferation, confirming SPRY2 as an essential dosage-sensitive negative-feedback regulator of FGF signaling in vivo.\",\n      \"evidence\": \"Endogenous co-IP, kinase-dead mutant analysis, and neurite outgrowth assay (Tesk1); Spry2 KO and BAC-transgene rescue with proliferation and FGF-target gene analysis (palate)\",\n      \"pmids\": [\"17974561\", \"17693063\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Tesk1 interaction domain on SPRY2 not mapped\", \"Downstream FGF target selectivity in palate not fully dissected\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Multiple discoveries expanded the SPRY2 mechanism: SPRY2 scaffolds Raf-1 without directly regulating its kinase activity, a Spry2-cCbl-TSC2 complex links FGF-10 signaling to mTORC1 activation in fetal lung, and JMJD1A-dependent H3K9 demethylation at the SPRY2 promoter was identified as a transcriptional on-switch.\",\n      \"evidence\": \"Raf-1 phosphorylation-site mutant panel with Co-IP and kinase assay; Spry2-cCbl-TSC2 Co-IP and GTPase assay in lung explants; ChIP-on-chip for H3K9me2 at SPRY2 promoter with functional rescue\",\n      \"pmids\": [\"20356462\", \"20622121\", \"20881000\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Raf-1 scaffolding model lacks structural detail\", \"TSC2-SPRY2 interaction stoichiometry unclear\", \"JMJD1A regulation confirmed only in nickel/hypoxia context\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"miR-21 was identified as a post-transcriptional repressor of SPRY2 in glioma and mesenchymal stem cells, revealing how the SPRY2 feedback loop can be hijacked to sustain ERK signaling in tumor and differentiation contexts.\",\n      \"evidence\": \"miR-21 overexpression/knockdown with SPRY2 protein quantification, invasion assay in glioma, and differentiation assay in MSCs\",\n      \"pmids\": [\"21278789\", \"23239100\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct 3′UTR luciferase validation not provided in these studies\", \"Other miRNAs targeting SPRY2 not systematically evaluated\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Conditional double knockout of Spry1/Spry2 in the lens demonstrated that FGF-induced Sprouty proteins restrain ERK to permit lens vesicle separation, establishing a non-redundant developmental requirement.\",\n      \"evidence\": \"Le-Cre conditional double KO with ERK phosphorylation and apoptosis assays\",\n      \"pmids\": [\"21743007\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Individual contributions of Spry1 vs. Spry2 in this tissue not fully resolved\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Two studies placed SPRY2 at the intersection of RTK trafficking and cancer: SPRY2 loss enhanced EGFR/HER2 endosomal signaling and PI3K/AKT-driven invasion in prostate cancer, while dominant-negative SPRY2Y55F accelerated AKT-driven hepatocarcinogenesis via MAPK and PKM2.\",\n      \"evidence\": \"siRNA/shRNA with receptor internalization and endosome fractionation (prostate); hydrodynamic gene delivery of Spry2Y55F in mouse liver with pharmacological rescue (liver)\",\n      \"pmids\": [\"22649008\", \"22617155\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether SPRY2 directly contacts endosomal machinery not shown\", \"PKM2 regulation mechanism downstream of SPRY2 not dissected\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Reconstitution of an ITSN1–Shp2–Cbl–SPRY2 complex showed that Shp2-mediated dephosphorylation of SPRY2 relieves its sequestration of Cbl, thereby promoting EGFR ubiquitylation and lysosomal degradation—mechanistically connecting SPRY2 phosphorylation status to receptor downregulation.\",\n      \"evidence\": \"Co-IP of quaternary complex, catalytic-dead Shp2 mutant, Spry2 ITSN1-binding site mutant, EGFR ubiquitylation assay\",\n      \"pmids\": [\"24216759\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo relevance of ITSN1-Shp2 axis not validated\", \"Whether other phosphatases substitute in ITSN1-null cells unknown\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Conditional knockout studies in tracheal basal cells and neural progenitors revealed that FGFR1-dependent post-translational modification of SPRY2 restrains stem-cell proliferation, and that homocysteine disrupts the Spry2/Dyrk1A/PP2A triad to alter SPRY2 phosphorylation and neural progenitor proliferation.\",\n      \"evidence\": \"Fgfr1 and Spry2 conditional KO in adult mouse trachea with BrdU assay; Spry2 promoter methylation, Dyrk1A/PP2A expression, and proliferation assays in neural progenitors\",\n      \"pmids\": [\"27046834\", \"27686255\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Identity of FGFR1-induced post-translational modification not determined\", \"Dyrk1A phosphorylation site on SPRY2 not mapped\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Discovery that SPRY2 is a PERK-dependent UPR target in β-cells, required for SERCA2 expression and ER calcium homeostasis, expanded its function beyond RTK-ERK feedback to organelle stress management and insulin secretion.\",\n      \"evidence\": \"Whole-genome RNAi screen, PERK pathway analysis, SERCA2/calcium measurements, conditional β-cell-specific Spry2 KO mouse\",\n      \"pmids\": [\"28246293\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism by which SPRY2 maintains SERCA2 levels unknown\", \"Whether SPRY2-ER calcium link exists outside β-cells not tested\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"SPRY2 was found to physically interact with and stabilize the MET receptor in rhabdomyosarcoma, demonstrating a context where SPRY2 sustains rather than inhibits RTK signaling.\",\n      \"evidence\": \"Co-IP and colocalization of SPRY2-MET, siRNA knockdown leading to MET degradation, ERK and differentiation assays\",\n      \"pmids\": [\"29445192\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism of SPRY2-mediated MET stabilization not defined\", \"Whether this is unique to rhabdomyosarcoma or generalizable unclear\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"In glioblastoma, NF-κB–driven autocrine FGFR signaling was shown to reactivate ERK, which induces SPRY2 transcription; paradoxically, SPRY2 itself promotes resistance to EGFR/MET inhibitors, revealing a context-dependent pro-survival role.\",\n      \"evidence\": \"Pharmacological epistasis (EGFR, MET, FGFR, NF-κB inhibitors), ERK-bioluminescent reporter, shRNA knockdown, xenograft model\",\n      \"pmids\": [\"32160544\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism by which SPRY2 promotes drug resistance not defined at the molecular level\", \"Generalizability across GBM subtypes not tested\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Two studies uncovered novel regulatory inputs: PKD phosphorylates SPRY2 at Ser112 cooperating with the COP9 signalosome to promote proteasomal turnover, and SPRY2 suppresses SRC-mediated LDHA Y10 phosphorylation to inhibit glycolysis in fibroblasts.\",\n      \"evidence\": \"In vitro kinase assay, Ser112Ala mutagenesis, CSN3 co-IP and stability assay (PKD); co-IP of LDHA-SRC disrupted by SPRY2, LDHA activity assay, in vivo tumor/metastasis model (LDHA)\",\n      \"pmids\": [\"37045830\", \"37507768\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether PKD-Ser112 and c-Cbl ubiquitination are sequential or parallel degradation pathways unknown\", \"SPRY2-SRC interaction interface not mapped\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Paclitaxel was shown to sequester SPRY2 on tubulin, depleting it from the plasma membrane and thereby de-repressing Raf/ERK signaling—a drug-induced redistribution mechanism aggravating pulmonary fibrosis.\",\n      \"evidence\": \"Co-IP of SPRY2-tubulin, membrane fractionation, microtubule depolymerization rescue, in vivo fibrosis model\",\n      \"pmids\": [\"37918858\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Tubulin-binding domain on SPRY2 not identified\", \"Whether this mechanism applies to other microtubule-targeting agents not tested\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Proximity proteomics and BRET studies established that SPRY2 is a direct K-Ras effector: the C-terminal fragment binds active K-Ras at the plasma membrane, homo/hetero-oligomerizes with SPRY4, and blocks Ras effector access, providing the first structural-level model of SPRY2 inhibition at the Ras node.\",\n      \"evidence\": \"TurboID proximity proteomics, BRET assay, interface mutagenesis, K-Ras membrane anchorage inhibitors, C2C12 differentiation assay\",\n      \"pmids\": [\"41362608\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Full atomic-resolution structure of K-Ras–SPRY2 complex not yet available\", \"Stoichiometry and dynamics of SPRY2 oligomers at the membrane not determined\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"Key unresolved questions include the structural basis of SPRY2 interactions with K-Ras/Raf/GRB2, how SPRY2 switches between tumor-suppressive and pro-survival roles in different cancer contexts, and the mechanism linking SPRY2 to ER calcium homeostasis and SERCA2 expression.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No atomic-resolution structure of any SPRY2 signaling complex\", \"Molecular determinants of context-dependent pro-tumorigenic vs. tumor-suppressive function uncharacterized\", \"SPRY2-SERCA2 regulatory mechanism undefined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [1, 3, 6, 7, 9, 15, 19]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [2, 8, 15]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [19, 20]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [2]},\n      {\"term_id\": \"GO:0005768\", \"supporting_discovery_ids\": [3, 6]},\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [20]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [1, 2, 3, 6, 7, 8, 9, 12, 15, 19]},\n      {\"term_id\": \"R-HSA-1266738\", \"supporting_discovery_ids\": [22, 23]},\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [0, 8, 17]},\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [16]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\n      \"GRB2\",\n      \"BRAF\",\n      \"CBL\",\n      \"KRAS\",\n      \"TESK1\",\n      \"ITSN1\",\n      \"MET\",\n      \"RAF1\"\n    ],\n    \"other_free_text\": []\n  }\n}\n```"}