{"gene":"SH3KBP1","run_date":"2026-04-28T20:42:07","timeline":{"discoveries":[{"year":2002,"finding":"CIN85 forms a constitutive complex with endophilins via their SH3 domains, and is recruited by Cbl to activated EGF receptors upon EGF stimulation; disruption of the Cbl–CIN85–endophilin complex blocks EGFR internalization and delays receptor degradation without affecting Cbl-directed receptor ubiquitination, demonstrating that this is a mechanistically distinct pathway from ubiquitin ligase activity.","method":"Co-immunoprecipitation, dominant-negative interference, internalization assays","journal":"Nature","confidence":"High","confidence_rationale":"Tier 2 — reciprocal Co-IP and functional block of internalization; replicated in same issue by independent lab (PMID:11894096)","pmids":["11894095","11894096"],"is_preprint":false},{"year":2002,"finding":"CIN85 binds to Cbl/Cbl-b via a novel polyproline-arginine motif (PXXXPR) in the C-terminus of Cbl; this interaction is enhanced by ligand-induced phosphorylation of Cbl and is required for Cbl-b-mediated internalization of EGFR and PDGFR but is dispensable for receptor polyubiquitination.","method":"Co-immunoprecipitation, deletion/mutation analysis, internalization assays","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 — clean epistasis between binding mutant and internalization vs. ubiquitination, replicated across Cbl family members","pmids":["12177062","12874286"],"is_preprint":false},{"year":2000,"finding":"CIN85 was identified as a novel c-Cbl-interacting protein; the second SH3 domain of CIN85 mediates constitutive association with c-Cbl, which is enhanced after EGF stimulation correlating with c-Cbl tyrosine phosphorylation.","method":"Co-immunoprecipitation, domain mapping","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 3 — single Co-IP/domain mapping study; foundational identification of the interaction","pmids":["10679202"],"is_preprint":false},{"year":2000,"finding":"CIN85 associates with BLNK/SLP65 through its SH3 domains; CIN85 also self-associates through its coiled-coil domain to form tetramers; both the coiled-coil and SH3 domains are required for subcellular localization.","method":"Yeast two-hybrid, co-immunoprecipitation, immunostaining","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 3 — single study with yeast two-hybrid confirmed by Co-IP and imaging","pmids":["11071869"],"is_preprint":false},{"year":2002,"finding":"Cbl and Cbl-b monoubiquitinate CIN85 after EGF stimulation; monoubiquitination requires direct CIN85–Cbl interaction, an intact RING finger domain, and a ubiquitin acceptor site in the CIN85 C-terminus; monoubiquitinated CIN85 is degraded in the lysosome together with polyubiquitinated EGFR.","method":"In vivo ubiquitination assays, mutagenesis of RING domain and ubiquitin acceptor site, lysosomal inhibitor experiments","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1-2 — mutagenesis of both writer (RING) and acceptor site combined with degradation assays; single lab but multiple orthogonal methods","pmids":["12218189"],"is_preprint":false},{"year":2003,"finding":"CIN85 SH3 domains specifically recognize the PXXXPR motif; individual SH3 domains bind with micromolar affinity while tandem SH3 domains bind with higher stoichiometry; full-length CIN85 simultaneously clusters multiple Cbl molecules, which is essential for ligand-induced stabilization of CIN85–Cbl–EGFR complexes and EGFR lysosomal degradation.","method":"Binding assays, peptide competition, affinity measurements, clustering assays in mammalian cells","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1-2 — in vitro binding with affinity quantification plus functional cell-based validation with domain mutants","pmids":["12874286"],"is_preprint":false},{"year":2003,"finding":"The atypical SH3-domain recognition consensus of CIN85 is PX(P/A)XXR; this consensus was validated by mutagenesis and in vitro binding, and allowed identification of novel binding partners synaptojanin 1 and PAK2 confirmed by GST pulldown and Far Western blotting.","method":"Target-assisted iterative peptide library screening, mutagenesis, in vitro binding, GST pulldown","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 — reconstituted in vitro with mutagenesis confirming binding determinants","pmids":["12829691"],"is_preprint":false},{"year":2004,"finding":"CIN85 associates with multiple endocytic effectors (SHIP-1, synaptojanin 2B1, ASAP1, ARAP3, Hip1R, STAP1, p115RhoGEF) all via PXXXPR motifs; CIN85 acts as a scaffold clustering these effectors into high-molecular-weight complexes; ASAP1 overexpression promotes EGFR recycling in a CIN85-binding-dependent manner.","method":"Co-immunoprecipitation, GST pulldown, functional recycling assay with PXXXPR mutants","journal":"Molecular biology of the cell","confidence":"High","confidence_rationale":"Tier 2 — multiple binding partners identified and functionally validated with binding-deficient mutants","pmids":["15090612"],"is_preprint":false},{"year":2004,"finding":"Alix/AIP1, which binds CIN85 and endophilins, antagonizes the Cbl–CIN85 complex by weakening CIN85–Cbl interaction and reducing EGFR ubiquitination; Alix overexpression reduces EGFR internalization while siRNA knockdown of Alix promotes internalization and downregulation.","method":"Co-immunoprecipitation, siRNA knockdown, internalization assays","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 — reciprocal gain- and loss-of-function with mechanistic Co-IP showing disrupted Cbl–CIN85 interaction","pmids":["15456872"],"is_preprint":false},{"year":2003,"finding":"CIN85 and CMS link the T cell surface protein CD2 (via its proline-rich cytoplasmic tail) to the actin-capping protein CAPZ; N-terminal SH3 domains of CIN85 bind CD2 (KD ~100 µM) and CAPZ binds the C-terminal half of CIN85/CMS, providing a direct connection to the actin cytoskeleton.","method":"Peptide pulldown, BIAcore binding analysis, overexpression functional assays","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1-2 — quantitative in vitro binding (BIAcore) combined with functional overexpression assays","pmids":["12690097"],"is_preprint":false},{"year":2003,"finding":"CIN85 (SETA) co-localizes with actin in microfilaments and focal adhesions and with microtubules; it interacts with focal adhesion kinase (FAK) and PYK-2 and promotes cell adhesion in ECIS assays; SETA dimerizes to interact with FAKs.","method":"Co-immunoprecipitation, ECIS cell adhesion assay, co-localization microscopy","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 2-3 — localization tied to functional adhesion readout; indirect interactions, single lab","pmids":["12771190"],"is_preprint":false},{"year":2004,"finding":"The yeast orthologue of CIN85, Sla1, directly binds the endophilin-like Rvs167; the ubiquitin ligase Rsp5 binds both Sla1 and Rvs167 and monoubiquitinates Rvs167 on Lys481 via its SH3 domain, providing conservation of the CIN85–endophilin–ubiquitin ligase axis.","method":"In vitro binding reconstitution, ubiquitination assay, mutagenesis (K481R)","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 — direct reconstitution of ubiquitination with mutagenesis confirmation in yeast ortholog","pmids":["14761940"],"is_preprint":false},{"year":2005,"finding":"Sprouty2 binds CIN85 SH3 domains A and C via PXXXPR motifs and acts at the Cbl–CIN85 interface to inhibit EGFR endocytosis; intact Sprouty2–Cbl–CIN85 ternary complex is required for inhibition; Sprouty4, which lacks CIN85-binding motifs, does not inhibit EGFR downregulation.","method":"Co-immunoprecipitation, domain mutagenesis, EGFR internalization assays","journal":"EMBO reports","confidence":"High","confidence_rationale":"Tier 2 — mechanistic dissection using binding-deficient mutants with clear functional readout","pmids":["15962011"],"is_preprint":false},{"year":2005,"finding":"CIN85 is recruited to the FcεRI signaling complex via Cbl after receptor engagement on mast cells and promotes ligand-induced receptor internalization, accelerated sorting into early endosomes and lysosomal delivery; CIN85-overexpressing mast cells are severely impaired in degranulation.","method":"Co-immunoprecipitation, confocal microscopy, degranulation assay","journal":"Journal of immunology","confidence":"Medium","confidence_rationale":"Tier 2-3 — functional gain-of-function with internalization and sorting readouts; single lab","pmids":["16177060"],"is_preprint":false},{"year":2005,"finding":"CIN85 binds MEKK4 via three PXXXPR motifs in MEKK4; disrupting this interaction impairs CIN85-enhanced activation of MKK6 and p38 MAP kinase upon oxidative stress/growth factor stimulation; CIN85 also promotes multi-ubiquitination of MEKK4.","method":"Co-immunoprecipitation, kinase assays, ubiquitination assay, dominant-negative mutants","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2-3 — Co-IP with functional kinase readout; single lab","pmids":["16256071"],"is_preprint":false},{"year":2007,"finding":"CIN85 co-localizes with AMAP1 at invadopodia in breast cancer cells; CIN85–AMAP1 binding is required for invasive activity; Cbl monoubiquitinates AMAP1, and this monoubiquitination is required for invasion; siRNA knockdown of CIN85 or Cbl inhibits invasion.","method":"Co-immunoprecipitation, siRNA knockdown, invasion assays, ubiquitination assays","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 — multiple orthogonal methods (Co-IP, siRNA, ubiquitination) with defined invasion phenotype","pmids":["17255943"],"is_preprint":false},{"year":2008,"finding":"All three SH3 domains of CIN85 bind ubiquitin; ubiquitin binding is at the canonical proline-rich ligand interaction surface and competes with PXXXPR ligands; disruption of ubiquitin binding leads to constitutive CIN85 ubiquitination and elevated basal EGFR ubiquitination, suggesting ubiquitin–SH3 competition regulates Cbl function.","method":"NMR structural model, binding assays, mutagenesis, ubiquitination assays","journal":"Biochemistry","confidence":"High","confidence_rationale":"Tier 1 — NMR-based structural model with mutagenesis and functional ubiquitination assays","pmids":["18680311"],"is_preprint":false},{"year":2008,"finding":"Drosophila Cindr (sole CD2AP/CIN85 family member) links E-cadherin and the adhesion receptor Roughest with actin-capping proteins (CPα and CPβ); reducing cindr causes defective E-cadherin and Roughest localization, impaired local cell movement, and tissue patterning defects.","method":"Genetic loss-of-function, immunofluorescence, protein interaction assays in Drosophila","journal":"The Journal of cell biology","confidence":"High","confidence_rationale":"Tier 2 — genetic epistasis in Drosophila ortholog with clear cellular phenotype and physical interaction data","pmids":["18362180"],"is_preprint":false},{"year":2008,"finding":"Endogenous CIN85/Ruk localizes predominantly to COPI-coated vesicles of the Golgi complex in a manner dependent on Golgi integrity and intact microtubules; only a small fraction is at clathrin-mediated endocytic compartments.","method":"Subcellular fractionation, immunofluorescence with organelle markers","journal":"Traffic (Copenhagen, Denmark)","confidence":"Medium","confidence_rationale":"Tier 2-3 — localization by imaging and fractionation; no functional consequence directly established","pmids":["18266907"],"is_preprint":false},{"year":2009,"finding":"CIN85 associates with endosomal membranes through binding of its positively charged C-terminal coiled-coil domain to the fusogenic lipid phosphatidic acid; deletion of the coiled-coil domain ablates membrane association and CIN85–c-Cbl interaction, blocking EGFR downregulation.","method":"Lipid-binding assay, deletion mutants, EGFR downregulation assays, confocal microscopy","journal":"Cell research","confidence":"Medium","confidence_rationale":"Tier 2-3 — protein-lipid interaction mapped to coiled-coil domain with functional EGFR trafficking readout; single lab","pmids":["19417776"],"is_preprint":false},{"year":2010,"finding":"CIN85 directly binds the coiled-coil domain of nephrin and podocin; in podocytes CIN85 promotes FGF-4-stimulated nephrin ubiquitination and internalization; co-expression of CD2AP reduces CIN85 binding to nephrin/podocin, indicating functional competition.","method":"Binding assays with CIN85 domain mutants, co-immunoprecipitation, internalization assays","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 — domain mapping with functional internalization assay and competition experiments","pmids":["20457601"],"is_preprint":false},{"year":2010,"finding":"CIN85 interacts with the HEV ORF3 protein, competing with formation of the Cbl–CIN85 complex at growth factor receptors; this reduces CIN85 ubiquitination and delays receptor trafficking to late endosomes/lysosomes, prolonging endomembrane signaling.","method":"Co-immunoprecipitation, competitive binding assays, EGFR/c-Met trafficking assays","journal":"Journal of virology","confidence":"Medium","confidence_rationale":"Tier 2-3 — competition binding confirmed with trafficking readout; single lab","pmids":["20130058"],"is_preprint":false},{"year":2010,"finding":"CIN85 in striatal neurons co-clusters with D2 dopamine receptors and interacts with endocytic regulators dynamin and endophilins; CIN85-deficient (exon 2 knockout) mice show decreased D2DR endocytosis in striatal neurons, elevated striatal dopamine and D2DR levels, and hyperactivity.","method":"Conditional knockout mouse model, receptor internalization assay, co-immunoprecipitation, behavioral analysis","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 — in vivo genetic model with mechanistic biochemical data linking CIN85 to dopamine receptor endocytosis","pmids":["20551902"],"is_preprint":false},{"year":2010,"finding":"The SH3 domains of CIN85 bind a PXXXPR motif near the C-terminus of tristetraprolin (TTP/hTTP); CIN85 co-expression leads to increased TTP phosphorylation at Ser66 and Ser93, possibly through co-recruitment of MEKK4.","method":"Yeast two-hybrid, co-immunoprecipitation, co-localization, phosphorylation analysis","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 3 — Co-IP and phosphorylation readout; functional consequences not fully defined","pmids":["20221403"],"is_preprint":false},{"year":2011,"finding":"CIN85 is constitutively associated with SLP65 in resting B cells; absence of the SLP65–CIN85 complex abrogates SLP65 phosphorylation, plasma membrane translocation, BCR-induced Ca2+ and NF-κB responses; live cell imaging confirmed both are components of the BCR primary transducer module.","method":"Mass spectrometry interactome, co-immunoprecipitation, live cell imaging, RNAi knockdown with functional signaling assays","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 — quantitative MS interactome plus functional signaling assays with clear loss-of-function phenotype","pmids":["21822214"],"is_preprint":false},{"year":2011,"finding":"B cell-specific deletion of CIN85 impairs IKK-β activation and T cell-independent type II antibody responses; introduction of constitutively active IKK-β corrects the defect, placing CIN85 upstream of IKK-β in BCR-to-NF-κB signaling.","method":"Conditional knockout (Mb1-cre), in vivo immunization, in vitro IKK-β kinase assay, rescue experiment","journal":"The Journal of experimental medicine","confidence":"High","confidence_rationale":"Tier 2 — genetic epistasis via constitutively active IKK-β rescue in conditional KO; robust in vivo phenotype","pmids":["21708930"],"is_preprint":false},{"year":2011,"finding":"CIN85 depletion decreases EGF-induced EGFR ubiquitination and delays degradation while increasing EGF recycling; CIN85 is constitutively associated with Hrs; overexpression of a dominant-negative CIN85 (three SH3 domains) inhibits EGF internalization, whereas RNAi knockdown of CIN85 does not, suggesting CIN85 functions primarily in endosomal sorting rather than initial internalization.","method":"RNAi knockdown, dominant-negative overexpression, ubiquitination assays, recycling/degradation assays, co-immunoprecipitation","journal":"Experimental cell research","confidence":"High","confidence_rationale":"Tier 2 — complementary gain- and loss-of-function with multiple trafficking readouts distinguishing internalization from sorting","pmids":["21635887"],"is_preprint":false},{"year":2011,"finding":"CIN85 overexpression inhibits FcεRI-induced Syk signaling and reduces Syk protein levels via the ubiquitin-proteasome pathway through c-Cbl; CIN85 overexpression limits Cbl binding to the negative regulator Sts1, while CIN85 knockdown promotes Cbl–Sts1 complex formation.","method":"Overexpression/knockdown, proteasome inhibitor rescue, co-immunoprecipitation","journal":"Journal of immunology","confidence":"Medium","confidence_rationale":"Tier 2-3 — mechanistic pathway defined by proteasome inhibitor rescue and Sts1 competition; single lab","pmids":["17675467"],"is_preprint":false},{"year":2011,"finding":"CIN85 modulates FcγRIIa downregulation in neutrophils in concert with c-Cbl; siRNA knockdown of CIN85 prevents FcγRIIa ubiquitination and degradation and increases IgG-mediated phagocytosis; CIN85 is a PKC substrate and classical PKCs positively regulate FcγRIIa ubiquitination and degradation.","method":"siRNA knockdown, co-immunoprecipitation, confocal microscopy, PKC inhibitor experiments","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2-3 — loss-of-function with defined receptor degradation phenotype and PKC mechanistic link; single lab","pmids":["21372129"],"is_preprint":false},{"year":2011,"finding":"CD2AP regulates SUMOylation of full-length CIN85 in podocytes; CIN85 is SUMOylated by SUMO-1, -2, and -3; SUMOylation is enhanced by CD2AP; mutation of Lys598 abolishes SUMOylation and increases CIN85 binding to nephrin.","method":"SUMO modification assays, mutagenesis (K598R), co-immunoprecipitation","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 1-2 — site-specific mutagenesis of SUMO acceptor combined with functional binding assay; establishes CD2AP as a SUMO-writer regulator","pmids":["22203040"],"is_preprint":false},{"year":2011,"finding":"SHKBP1 constitutively binds the SH3 domains of CIN85 via PXXXPR motifs, competing with c-Cbl; SHKBP1 binding prevents CIN85 translocation to EGFR-containing vesicles and reduces EGFR degradation, enhancing EGF-induced signaling.","method":"Co-immunoprecipitation, domain competition, EGFR trafficking/degradation assays","journal":"Cell biochemistry and function","confidence":"Medium","confidence_rationale":"Tier 3 — single lab, Co-IP with functional trafficking readout","pmids":["21830225"],"is_preprint":false},{"year":2012,"finding":"EGF stimulation induces Src-mediated tyrosine phosphorylation of CIN85; phospho-CIN85 interacts with Rab5-positive early endosomes and mediates EGFR sequestration into multivesicular bodies for degradation; disruption of Src-dependent CIN85 phosphorylation impairs EGFR ubiquitination and MVB sorting.","method":"Phosphorylation assays, co-immunoprecipitation with Rab5, dominant-negative and kinase inhibitor experiments, MVB sorting assays","journal":"Molecular biology of the cell","confidence":"High","confidence_rationale":"Tier 2 — identifies Src as the writer kinase phosphorylating CIN85 and links this to endosomal sorting with subcellular localization data","pmids":["22833562"],"is_preprint":false},{"year":2010,"finding":"CIN85 interacts with the CIN85-interacting protein dynamin 2 (Dyn2) in an EGF-stimulation-dependent manner; disruption of the CIN85–Dyn2 interaction causes internalized EGFR to accumulate in aberrantly elongated late endosomal tubules and sustains downstream signaling, identifying a novel late-endocytic role for CIN85.","method":"Co-immunoprecipitation, dominant-negative disruption, confocal microscopy, signaling assays","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 — direct interaction confirmed, disruption gives specific morphological and signaling phenotype","pmids":["20711168"],"is_preprint":false},{"year":2007,"finding":"CIN85/CIN85 and CMS can crosslink F-actin into bundles via their proline-rich and coiled-coil domains; removal of these domains reduces cell migration; CMS and CIN85 form heterotypic complexes via their coiled-coil domains.","method":"F-actin binding/bundling assays, domain deletion mutants, migration assays","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 2-3 — in vitro F-actin assay with functional migration readout; single lab","pmids":["17606992"],"is_preprint":false},{"year":2013,"finding":"MUC1 associates with CIN85 at invadopodia-like structures; siRNA-mediated silencing of CIN85 reduces breast cancer cell migration and invasion in vitro and lung metastasis in vivo; MUC1 overexpression partially rescues CIN85-depletion-reduced invasion.","method":"Co-immunoprecipitation, siRNA knockdown, invasion/migration assays, in vivo metastasis model","journal":"Oncotarget","confidence":"Medium","confidence_rationale":"Tier 2-3 — Co-IP with functional loss-of-function invasion and in vivo metastasis data; single lab","pmids":["24072600"],"is_preprint":false},{"year":2015,"finding":"TβRI interacts with CIN85 SH3 domains in a TGFβ- and TRAF6-dependent manner; CIN85 knockdown causes TβRI accumulation in intracellular compartments and diminishes Smad2 phosphorylation; CIN85 overexpression increases TβRI at the cell surface via a Rab11-dependent recycling pathway.","method":"Co-immunoprecipitation, siRNA knockdown, Rab11 dominant-negative, Smad2 phosphorylation assays, surface biotinylation","journal":"The Journal of cell biology","confidence":"High","confidence_rationale":"Tier 2 — mechanistic dissection with dominant-negative Rab11 placing CIN85 in recycling pathway; multiple orthogonal methods","pmids":["26169354"],"is_preprint":false},{"year":2016,"finding":"CIN85 trimerizes via its C-terminal coiled-coil domain; trimeric CIN85 oligomerizes SLP65 via its multiple SH3 domains interacting with SLP65 PRMs, propagating condensate formation; this oligomeric signaling complex pre-assembles in resting B cells and is required for efficient BCR signal initiation.","method":"NMR structural analysis, biochemical oligomerization assays, B cell signaling functional assays","journal":"Science signaling","confidence":"High","confidence_rationale":"Tier 1-2 — structural (NMR) plus biochemical reconstitution with functional B cell validation","pmids":["27353366"],"is_preprint":false},{"year":2016,"finding":"CIN85/RukL knockout in diabetic mice prevents nephrin internalization, preserves nephrin surface expression on the slit diaphragm, and reduces proteinuria; conversely, CIN85 overexpression in zebrafish induces edema and filtration barrier disruption.","method":"Knockout mouse model (diabetic), zebrafish overexpression, nephrin surface expression, proteinuria measurement","journal":"Diabetes","confidence":"High","confidence_rationale":"Tier 2 — in vivo genetic models (mouse KO and zebrafish OE) with clear functional readout","pmids":["27531950"],"is_preprint":false},{"year":2018,"finding":"Germline deletion of CIN85 in humans causes X-linked antibody deficiency with intact immune cell development but intrinsic B cell defects in NF-κB activation and CD86 upregulation downstream of BCR, establishing non-redundant in vivo roles of CIN85 in humoral immunity.","method":"Human germline genetics, B cell functional assays (NF-κB activation, CD86 expression)","journal":"The Journal of experimental medicine","confidence":"High","confidence_rationale":"Tier 2 — human loss-of-function genetics with mechanistic cell-type-specific signaling readouts","pmids":["29636373"],"is_preprint":false},{"year":2019,"finding":"CIN85 is a novel binding partner of PHD2 (but not PHD1 or PHD3); the N-terminal SH3 domains of CIN85 interact with the proline-arginine-rich N-terminus of PHD2, inhibiting PHD2 hydroxylase activity and thereby stabilizing HIF-1α; CRISPR disruption of the CIN85–PHD2 interaction in cells impairs tumor growth and migration.","method":"Co-immunoprecipitation, PHD2 enzymatic activity assay, CRISPR/Cas9 editing, in vivo xenograft","journal":"Cancer research","confidence":"High","confidence_rationale":"Tier 1-2 — enzymatic inhibition assay combined with CRISPR editing and in vivo functional validation","pmids":["31142511"],"is_preprint":false},{"year":2019,"finding":"After TCR stimulation, CIN85 is recruited to the TCR signaling complex and limits T cell activation; CIN85-deficient T cells show enhanced Zap70, SLP76, and ERK phosphorylation; the inhibitory function requires CIN85 SH3 and PR domains and involves association with the phosphatase Sts-2.","method":"T cell-specific conditional knockout, signaling analysis (phospho-western), co-immunoprecipitation, domain mutagenesis","journal":"Science signaling","confidence":"High","confidence_rationale":"Tier 2 — conditional KO combined with domain mutants identifying Sts-2 as the effector","pmids":["30723173"],"is_preprint":false},{"year":2022,"finding":"CIN85 localizes to the intercellular bridge during cytokinesis; CIN85 directly interacts with the N-terminal region of anillin and SEPT9, facilitating SEPT9-containing filament localization to the ICB plasma membrane; loss of CIN85 impairs cytokinesis timing and robustness.","method":"siRNA knockdown, co-immunoprecipitation, live cell imaging, direct in vitro binding","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 1-2 — direct binding reconstitution combined with live imaging and loss-of-function cytokinesis phenotype","pmids":["36044846"],"is_preprint":false},{"year":2023,"finding":"ASAP2 directly binds CIN85, disrupting its interaction with c-MET and thereby antagonizing CIN85-induced c-MET internalization and lysosomal degradation; CIN85 knockdown rescues the inhibitory effects of ASAP2 knockdown on HGF/c-MET signaling.","method":"Co-immunoprecipitation, cycloheximide chase, siRNA epistasis knockdown experiments","journal":"Experimental hematology & oncology","confidence":"Medium","confidence_rationale":"Tier 2-3 — epistasis by double knockdown with receptor degradation/signaling readout; single lab","pmids":["37061723"],"is_preprint":false},{"year":2023,"finding":"An intramolecular SH3C:PRM interaction within CIN85 (between C-terminal SH3 domain and adjacent PRM) autoinhibits CIN85 valency toward SLP65; phosphorylation of a serine residue adjacent to the PRM modulates this intramolecular interaction; disrupting it impairs SLP65/CIN85 condensate formation, membrane recruitment of CIN85, and BCR-induced Ca2+ mobilization.","method":"High-resolution NMR, mutagenesis, B cell functional assays (Ca2+ flux, membrane recruitment)","journal":"Journal of the American Chemical Society","confidence":"High","confidence_rationale":"Tier 1 — NMR structural validation of intramolecular interaction with phosphorylation-regulatory mechanism confirmed by B cell functional assays","pmids":["38111344"],"is_preprint":false},{"year":2024,"finding":"SH3KBP1 enhances RIG-I signal transduction by increasing K63-linked polyubiquitination through interaction with E3 ligase TRIM25; PRRSV NSP2 induces autophagic degradation of SH3KBP1 via the third PVPAPR motif in NSP2; Sh3kbp1−/− mice are more susceptible to VSV infection with reduced serum IFN-β.","method":"Co-immunoprecipitation, ubiquitination assays, knockout mouse model, VSV infection, autophagy assays","journal":"PLoS pathogens","confidence":"High","confidence_rationale":"Tier 2 — knockout mouse with infection phenotype, ubiquitination assay identifying TRIM25 as writer, and mechanism of viral antagonism","pmids":["39466846"],"is_preprint":false},{"year":2025,"finding":"SH3KBP1 N-terminus binds dynamin-2 and C-terminus associates with ER through calnexin; SH3KBP1 is required for myoblast fusion, myonuclear positioning, and myotube elongation; SH3KBP1 contributes to triad formation and excitation-contraction coupling in mature muscle fibers.","method":"siRNA screen, co-immunoprecipitation, domain mapping, excitation-contraction coupling assays, Dnm2 mutant mouse model","journal":"EMBO reports","confidence":"High","confidence_rationale":"Tier 2 — multiple binding partners identified by domain mapping combined with functional siRNA phenotypes and in vivo mouse model","pmids":["40065183"],"is_preprint":false},{"year":2025,"finding":"CIN85 is recruited to tubular recycling endosomes (TREs) via SH3-domain interaction with MICAL-L1; depletion of CIN85 impairs recycling of CD98 cargo; CIN85 likely regulates TRE function through effects on the actin cytoskeleton.","method":"Co-immunoprecipitation, siRNA knockdown, fluorescence live imaging, recycling assays","journal":"Traffic (Copenhagen, Denmark)","confidence":"Medium","confidence_rationale":"Tier 2-3 — localization via Co-IP and imaging tied to recycling functional assay; single lab","pmids":["40740057"],"is_preprint":false},{"year":2025,"finding":"SH3KBP1 is recruited to bleb necks where it interacts with septins and recruits PI3K via liquid-liquid phase separation; the Septin–SH3KBP1–PI3K axis establishes differential PI(3,4,5)P3 enrichment at bleb membranes and confers anoikis resistance.","method":"Fluorescence microscopy, co-immunoprecipitation, phase-separation assays, PI assays, anoikis resistance functional assays","journal":"American journal of physiology. Cell physiology","confidence":"Medium","confidence_rationale":"Tier 2-3 — phase-separation behavior and PI distribution linked to functional anoikis readout; single recent lab","pmids":["40445717"],"is_preprint":false},{"year":2003,"finding":"CIN85 associates with disabled-2 (Dab2) via all three SH3 domains binding the PKPAPR motif in Dab2; this association recruits CIN85 to clathrin-coated structures; upon growth factor stimulation, Dab2 and clathrin dissociate from CIN85, enabling Cbl binding to CIN85.","method":"Co-immunoprecipitation, domain mapping, clathrin association assays","journal":"FEBS letters","confidence":"Medium","confidence_rationale":"Tier 3 — single Co-IP study demonstrating dynamic switching of CIN85 binding partners during endocytosis","pmids":["14596919"],"is_preprint":false},{"year":2004,"finding":"Multiple domains of CIN85 (SH3 domains and coiled-coil) interact with the p85α regulatory subunit of PI3K; the SH3 domain of p85α is required to outcompete intramolecular SH3–PRR interaction within CIN85; homodimerization via coiled-coil affects both intra- and intermolecular CIN85 interactions.","method":"GST pulldown, domain deletion/mutation analyses, binding competition assays","journal":"Journal of molecular biology","confidence":"Medium","confidence_rationale":"Tier 3 — pulldown study with domain mapping; single lab","pmids":["15476827"],"is_preprint":false},{"year":2006,"finding":"CIN85 is localized at synapses; its first SH3 domain and C-terminal region bind the proline-rich region and N-terminal region of dendrin, respectively; CIN85, dendrin, and S-SCAM form a ternary complex in vitro that facilitates recruitment of dendrin and S-SCAM to CIN85-positive vesicle-like structures.","method":"Yeast two-hybrid, co-immunoprecipitation, immunocytochemistry, subcellular fractionation, in vitro complex reconstitution","journal":"Journal of biochemistry","confidence":"Medium","confidence_rationale":"Tier 2-3 — ternary complex reconstituted in vitro with localization data; single lab","pmids":["16751601"],"is_preprint":false},{"year":2009,"finding":"Intersectin 1 (ITSN1) constitutively forms a complex with CIN85/Ruk independent of EGF stimulation; the interaction is mediated by ITSN1 SH3A domain and the third/fourth proline-rich blocks of CIN85; both proteins co-localize with c-Cbl in MCF-7 cells.","method":"Co-immunoprecipitation, domain mapping, confocal co-localization","journal":"Cellular signalling","confidence":"Low","confidence_rationale":"Tier 3 — single Co-IP; functional consequence not defined","pmids":["19166927"],"is_preprint":false},{"year":2014,"finding":"Basic amino acids K645, K646, R648 and R650 in the CIN85 coiled-coil domain are required for interaction with both c-Cbl and phosphatidic acid; their mutation dissociates CIN85 from endosomes and prevents ESCRT assembly on endosomal membranes, blocking EGFR sorting for degradation.","method":"Mutagenesis, co-immunoprecipitation, phosphatidic acid binding assay, ESCRT co-IP, EGFR degradation assay","journal":"BMC biochemistry","confidence":"Medium","confidence_rationale":"Tier 2-3 — site-specific mutagenesis with multiple functional readouts; single lab","pmids":["25005938"],"is_preprint":false},{"year":2016,"finding":"CIN85 interacts with the catalytic subunit of PP2A via a PPKKPRP motif (P3 block) in its proline-rich region; purified PP2Ac-CIN85 complex exhibits suppressed phosphatase activity; disruption of PP2Ac-CIN85 interaction by cell-permeable P3 peptide reduces platelet spreading on fibrinogen and thrombus-related functions.","method":"Yeast two-hybrid, mutagenesis, in vitro PP2A phosphatase activity assay, cell-permeable peptide, ECIS cell adhesion","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1-2 — in vitro enzymatic assay shows CIN85 suppresses PP2Ac activity; domain mutagenesis confirmed; functional platelet assay","pmids":["27334924"],"is_preprint":false},{"year":2011,"finding":"SOX10 binds a highly conserved element within an alternative promoter of the Sh3kbp1 locus in Schwann cells; mutation of the SOX10 binding site ablates promoter activity; ectopic SOX10 expression drives endogenous Sh3kbp1 expression in SOX10-negative cells.","method":"Reporter gene assay, electrophoretic mobility shift/ChIP-equivalent, ectopic SOX10 expression, mutagenesis","journal":"Molecular and cellular neurosciences","confidence":"Medium","confidence_rationale":"Tier 2-3 — transcriptional regulation (not post-translational mechanistic finding) but establishes SOX10 as direct upstream regulator of Sh3kbp1 expression in Schwann cells","pmids":["22037207"],"is_preprint":false},{"year":2025,"finding":"SH3KBP1 co-localizes with MYO1F, CD2AP, ASAP1, and SH3BP2 at podosomes and phagocytic cups in macrophages/microglia; structural modelling and mutagenesis confirmed multivalent proline-rich motif interactions between SH3KBP1 and the MYO1F SH3 domain.","method":"Proximity labelling proteomics, structural modelling, mutagenesis, immunofluorescence","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 2-3 — proximity labelling combined with mutagenesis and localization; functional consequence not fully resolved","pmids":["41208482"],"is_preprint":false}],"current_model":"SH3KBP1 (CIN85) is a multidomain scaffold/adaptor protein whose three SH3 domains recognize atypical PXXXPR motifs to assemble multiprotein complexes that direct clathrin-mediated endocytosis, endosomal sorting, and lysosomal degradation of receptor tyrosine kinases (EGFR, c-MET, D2DR, FcεRI, FcγRIIa, TβRI) via constitutive association with endophilins and ligand-stimulated recruitment by Cbl/Cbl-b, which also monoubiquitinates CIN85; additionally, CIN85 scaffolds B cell antigen receptor signaling by oligomerizing SLP65 into pre-formed condensates that drive NF-κB and Ca2+ responses, inhibits PHD2 prolyl hydroxylase activity to stabilize HIF-1α, facilitates cytokinesis via anillin–SEPT9 interactions, supports myofiber integrity through dynamin-2 and calnexin binding, and regulates actin dynamics, cell adhesion, platelet function, and innate immune signaling through diverse binding partners."},"narrative":{"teleology":[{"year":2000,"claim":"Establishing CIN85 as a Cbl-interacting adaptor resolved the question of how Cbl's endocytic function is coupled to downstream effectors beyond its E3 ligase activity.","evidence":"Co-immunoprecipitation and domain mapping in mammalian cells identifying SH3-domain-mediated Cbl binding","pmids":["10679202"],"confidence":"Medium","gaps":["Functional consequence of CIN85–Cbl interaction on receptor trafficking not yet tested","No structural detail of binding interface"]},{"year":2002,"claim":"Two concurrent studies demonstrated that CIN85 constitutively complexes with endophilins and is recruited to activated EGFR via Cbl, revealing a mechanistically distinct internalization pathway separable from Cbl's ubiquitin ligase activity.","evidence":"Reciprocal Co-IPs, dominant-negative interference, and EGFR internalization assays in mammalian cells by two independent laboratories","pmids":["11894095","11894096","12177062"],"confidence":"High","gaps":["Whether CIN85 functions in initial internalization versus endosomal sorting was unresolved","Structural basis of PXXXPR recognition unknown"]},{"year":2002,"claim":"Discovery that Cbl monoubiquitinates CIN85 and that monoubiquitinated CIN85 is co-degraded with polyubiquitinated EGFR in lysosomes established a feedback mechanism linking the scaffold to receptor fate.","evidence":"In vivo ubiquitination assays with RING-domain and ubiquitin-acceptor-site mutagenesis, lysosomal inhibitor rescue","pmids":["12218189"],"confidence":"High","gaps":["Identity of the monoubiquitinated lysine residue in CIN85 not mapped","Physiological relevance of CIN85 co-degradation unclear"]},{"year":2003,"claim":"Definition of the PX(P/A)XXR consensus for CIN85 SH3 domains and demonstration that tandem SH3 domains cluster multiple Cbl molecules explained how CIN85 valency drives ligand-induced complex stabilization and receptor downregulation.","evidence":"Iterative peptide library screening, in vitro affinity measurements, mutagenesis, and functional clustering assays in mammalian cells; identification of new partners synaptojanin-1, PAK2, and Dab2","pmids":["12874286","12829691","14596919"],"confidence":"High","gaps":["High-resolution structure of SH3:PXXXPR complex not determined","How ligand-gated switching between Dab2 and Cbl binding is regulated remained unclear"]},{"year":2003,"claim":"Linking CIN85 to the actin cytoskeleton through direct CAPZ binding and to focal adhesions through FAK/PYK-2 expanded its role beyond endocytosis to cell adhesion and actin dynamics.","evidence":"BIAcore quantitative binding to CD2/CAPZ, F-actin bundling assays, ECIS adhesion assays, and co-localization with focal adhesion markers","pmids":["12690097","12771190"],"confidence":"Medium","gaps":["In vivo relevance of CIN85–CAPZ axis to T cell function not tested","Whether F-actin bundling is CIN85-specific or shared with CMS not resolved"]},{"year":2004,"claim":"Identification of a broad spectrum of PXXXPR-containing endocytic effectors (SHIP-1, ASAP1, ARAP3, Hip1R) binding CIN85, and conservation of the CIN85–endophilin–ubiquitin ligase axis in yeast (Sla1–Rvs167–Rsp5), established CIN85 as an evolutionarily conserved endocytic scaffold hub.","evidence":"Co-IPs and pulldowns with PXXXPR-mutant controls in mammalian cells; in vitro reconstitution of Rsp5-mediated Rvs167 ubiquitination in yeast","pmids":["15090612","14761940"],"confidence":"High","gaps":["Hierarchy and competition among multiple PXXXPR partners for CIN85 SH3 occupancy undefined","Whether yeast Sla1 and mammalian CIN85 are true functional orthologs debated"]},{"year":2005,"claim":"Sprouty2 and Alix were identified as negative regulators that compete at the Cbl–CIN85 interface, providing a mechanistic basis for fine-tuning EGFR downregulation through antagonistic complex formation.","evidence":"Domain mutagenesis, ternary complex Co-IPs, and reciprocal gain/loss-of-function EGFR internalization assays","pmids":["15962011","15456872"],"confidence":"High","gaps":["Physiological contexts in which Sprouty2 versus Alix dominate are unknown","Whether antagonism extends to non-EGFR receptors not tested"]},{"year":2005,"claim":"Demonstrating CIN85 recruitment to FcεRI via Cbl in mast cells and its role in receptor internalization/sorting extended the Cbl–CIN85 trafficking paradigm to immune receptors beyond RTKs.","evidence":"Co-IP, confocal sorting assays, and degranulation functional readout in mast cells","pmids":["16177060"],"confidence":"Medium","gaps":["Whether CIN85 also controls FcεRI signaling amplitude independently of internalization unclear","No genetic loss-of-function in mast cells"]},{"year":2008,"claim":"NMR-based discovery that CIN85 SH3 domains bind ubiquitin at the same surface used for PXXXPR ligands revealed a competition mechanism that auto-regulates Cbl-mediated ubiquitination, and Drosophila Cindr genetics confirmed conserved roles in adhesion receptor localization.","evidence":"NMR structural modeling with mutagenesis and ubiquitination assays; genetic loss-of-function in Drosophila with E-cadherin/Roughest localization phenotypes","pmids":["18680311","18362180"],"confidence":"High","gaps":["Relative affinities of ubiquitin versus PXXXPR for each SH3 domain not quantified in cells","How ubiquitin competition is resolved temporally during trafficking unknown"]},{"year":2010,"claim":"CIN85-knockout mice showed impaired D2 dopamine receptor endocytosis, hyperactivity, and elevated striatal dopamine, providing the first in vivo genetic evidence that CIN85 is a non-redundant endocytic regulator in the brain.","evidence":"Conditional exon-2 knockout mouse with receptor internalization assays, neurochemistry, and behavioral analysis","pmids":["20551902"],"confidence":"High","gaps":["Whether CD2AP compensates partially in other tissues not addressed","Molecular mechanism linking CIN85 loss to elevated dopamine release undefined"]},{"year":2010,"claim":"Binding of CIN85 to dynamin-2 at late endosomes was shown to prevent aberrant tubulation and sustained signaling, identifying a distinct late-endocytic membrane-scission role for CIN85 beyond early internalization.","evidence":"Co-IP, dominant-negative disruption, confocal imaging of elongated endosomal tubules, and signaling assays","pmids":["20711168"],"confidence":"High","gaps":["Whether CIN85 directly stimulates dynamin-2 GTPase activity or acts as a scaffold not resolved","Relative contribution of late-endosomal versus early-endocytic CIN85 to EGFR fate unclear"]},{"year":2011,"claim":"CIN85 was established as a core component of the BCR primary transducer module: it constitutively associates with SLP65, and B-cell-specific CIN85 deletion abrogates IKK-β activation and T-cell-independent antibody responses, placing CIN85 upstream of NF-κB in humoral immunity.","evidence":"Quantitative MS interactome, live-cell imaging, conditional Mb1-cre knockout with IKK-β rescue, and in vivo immunization","pmids":["21822214","21708930"],"confidence":"High","gaps":["How CIN85 specifically promotes IKK-β activation mechanistically undefined","Contribution of CIN85 to germinal center reactions not examined"]},{"year":2011,"claim":"Refined trafficking analysis showed CIN85 depletion delays EGFR degradation and increases recycling without blocking initial internalization, repositioning CIN85's primary function to endosomal sorting rather than plasma-membrane uptake; concurrently, CIN85 was linked to FcγRIIa and Syk degradation in neutrophils and mast cells.","evidence":"RNAi versus dominant-negative comparison with EGFR recycling/degradation assays; siRNA in neutrophils with phagocytosis readout","pmids":["21635887","21372129","17675467"],"confidence":"High","gaps":["Whether the sorting versus internalization distinction applies to all CIN85-regulated receptors not tested","Mechanism by which CIN85 recruits ESCRT machinery not defined"]},{"year":2011,"claim":"Discovery that CD2AP promotes CIN85 SUMOylation at K598 and that SUMOylation reduces CIN85 binding to nephrin established a post-translational switch controlling slit-diaphragm receptor availability in podocytes.","evidence":"SUMO modification assays with K598R mutagenesis and Co-IP with nephrin","pmids":["22203040"],"confidence":"High","gaps":["Identity of the SUMO E3 ligase not determined","In vivo relevance of K598 SUMOylation to podocyte function not tested"]},{"year":2012,"claim":"Src-mediated tyrosine phosphorylation of CIN85 was identified as a signal directing CIN85 to Rab5-positive early endosomes for EGFR MVB sorting, linking kinase signaling to CIN85's endosomal sorting function.","evidence":"Phosphorylation assays, Rab5 Co-IP, Src inhibitor experiments, and MVB sorting assays","pmids":["22833562"],"confidence":"High","gaps":["Specific tyrosine residue(s) phosphorylated by Src not mapped","Whether Src phosphorylation of CIN85 affects non-EGFR cargoes unknown"]},{"year":2015,"claim":"CIN85 was shown to promote TβRI surface recycling via Rab11 rather than degradation, revealing that CIN85 can direct receptors toward recycling in a context-dependent manner, opposite to its role in EGFR downregulation.","evidence":"Co-IP, siRNA, dominant-negative Rab11, surface biotinylation, and Smad2 phosphorylation assays","pmids":["26169354"],"confidence":"High","gaps":["What determines whether CIN85 routes a receptor for degradation versus recycling unresolved","Role of TRAF6 in CIN85 recruitment to TβRI not fully dissected"]},{"year":2016,"claim":"Structural and biochemical demonstration that CIN85 trimerizes via its coiled-coil domain and oligomerizes SLP65 into condensates through multivalent SH3:PRM interactions explained how pre-formed signaling clusters enable rapid BCR signal initiation; separately, CIN85 was shown to inhibit PP2A catalytic activity in platelets, affecting platelet spreading.","evidence":"NMR structural analysis, oligomerization reconstitution, B cell signaling assays; yeast two-hybrid, in vitro PP2A activity assay, cell-permeable peptide disruption","pmids":["27353366","27334924"],"confidence":"High","gaps":["Whether CIN85-driven condensates undergo true liquid-liquid phase separation in vivo not proven","In vivo platelet phenotype of CIN85 loss not characterized"]},{"year":2016,"claim":"CIN85 knockout in diabetic mice preserved nephrin surface expression and reduced proteinuria, while zebrafish overexpression disrupted the filtration barrier, providing direct in vivo evidence for CIN85 as a driver of nephrin internalization and diabetic nephropathy pathogenesis.","evidence":"Knockout mouse model (diabetic), zebrafish overexpression, nephrin surface quantification, proteinuria measurement","pmids":["27531950"],"confidence":"High","gaps":["Whether CIN85-mediated nephrin internalization is Cbl-dependent in vivo not determined","Applicability to non-diabetic nephropathies untested"]},{"year":2018,"claim":"Identification of germline SH3KBP1 loss in humans causing X-linked antibody deficiency with defective NF-κB activation established a non-redundant, disease-causing role for CIN85 in human humoral immunity.","evidence":"Human genetic analysis with B cell functional assays (NF-κB, CD86) from affected individuals","pmids":["29636373"],"confidence":"High","gaps":["Precise mutation(s) and their structural consequences not fully characterized","Whether T cell or myeloid defects contribute to clinical phenotype not fully resolved"]},{"year":2019,"claim":"CIN85 was shown to directly inhibit PHD2 hydroxylase activity, stabilizing HIF-1α, and to limit T cell activation via the phosphatase Sts-2, revealing non-trafficking functions of CIN85 as an enzyme inhibitor and signaling attenuator.","evidence":"PHD2 enzymatic activity assay, CRISPR editing, xenograft tumor model; T cell conditional KO with Zap70/SLP76/ERK phosphorylation and Sts-2 Co-IP","pmids":["31142511","30723173"],"confidence":"High","gaps":["Structural basis of PHD2 inhibition by CIN85 SH3 domains not determined","How CIN85 recruits Sts-2 to the TCR complex mechanistically undefined"]},{"year":2022,"claim":"Discovery that CIN85 localizes to the intercellular bridge during cytokinesis and directly binds anillin and SEPT9 to facilitate septin filament membrane recruitment revealed a cell-division role entirely distinct from endocytic trafficking.","evidence":"Direct in vitro binding reconstitution, live cell imaging of cytokinesis, siRNA loss-of-function","pmids":["36044846"],"confidence":"High","gaps":["Whether CIN85 condensate/oligomerization properties contribute to septin assembly unknown","Upstream signal recruiting CIN85 to the ICB not identified"]},{"year":2023,"claim":"NMR resolution of an intramolecular SH3C:PRM autoinhibitory interaction within CIN85, regulated by serine phosphorylation, explained how CIN85 valency toward SLP65 is gated, controlling condensate formation and BCR-induced Ca²⁺ signaling.","evidence":"High-resolution NMR, phospho-mimetic mutagenesis, B cell Ca²⁺ flux and membrane recruitment assays","pmids":["38111344"],"confidence":"High","gaps":["Identity of the kinase phosphorylating the regulatory serine not established","Whether autoinhibition regulates CIN85 interactions with non-SLP65 partners untested"]},{"year":2024,"claim":"SH3KBP1 was shown to enhance RIG-I antiviral signaling by promoting TRIM25-mediated K63-ubiquitination, and Sh3kbp1-knockout mice showed increased susceptibility to VSV, expanding CIN85's role to innate antiviral immunity.","evidence":"Co-IP, ubiquitination assays, knockout mouse VSV infection model, IFN-β measurement","pmids":["39466846"],"confidence":"High","gaps":["Whether CIN85 scaffolds TRIM25–RIG-I or allosterically activates TRIM25 not distinguished","Relevance to RNA viruses beyond VSV/PRRSV unknown"]},{"year":2025,"claim":"Recent studies expanded CIN85 functions to muscle biology (myoblast fusion, triad formation via dynamin-2/calnexin), tubular recycling endosomes (CD98 recycling via MICAL-L1), podosome assembly in macrophages (via MYO1F), and bleb-neck signaling (septin–PI3K phase separation conferring anoikis resistance).","evidence":"siRNA screens in myotubes with Dnm2-mutant mouse; live imaging of TRE recycling; proximity labelling proteomics in macrophages; phase-separation and PI assays with anoikis readout","pmids":["40065183","40740057","41208482","40445717"],"confidence":"Medium","gaps":["In vivo muscle phenotype of CIN85 loss not fully characterized","Whether TRE and bleb-neck functions depend on CIN85 oligomerization unknown","Phase-separation behavior awaits reconstitution with purified components"]},{"year":null,"claim":"Key unresolved questions include the structural basis of CIN85 SH3 selectivity among dozens of PXXXPR partners, the kinase(s) controlling autoinhibitory phosphorylation, how CIN85 switches between degradative and recycling sorting of different cargoes, and the full physiological spectrum of CIN85 loss in humans beyond antibody deficiency.","evidence":"","pmids":[],"confidence":"High","gaps":["No high-resolution crystal/cryo-EM structure of full-length CIN85 or CIN85–Cbl complex","Kinase phosphorylating the autoinhibitory serine unknown","Decision logic routing receptors to degradation versus recycling not defined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[0,1,5,7,24,36]},{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[9,10,17,33]},{"term_id":"GO:0008289","term_label":"lipid binding","supporting_discovery_ids":[19,52]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[39,53]}],"localization":[{"term_id":"GO:0005768","term_label":"endosome","supporting_discovery_ids":[19,26,31,32,52]},{"term_id":"GO:0005794","term_label":"Golgi apparatus","supporting_discovery_ids":[18]},{"term_id":"GO:0031410","term_label":"cytoplasmic vesicle","supporting_discovery_ids":[0,13,48]},{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[35,47]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[3,36]},{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[10,33,41]}],"pathway":[{"term_id":"R-HSA-5653656","term_label":"Vesicle-mediated transport","supporting_discovery_ids":[0,1,4,7,22,26,32]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[24,25,38,40,44]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[35,39,14]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[41]},{"term_id":"R-HSA-9609507","term_label":"Protein localization","supporting_discovery_ids":[13,20,37,46]}],"complexes":["CIN85–endophilin complex","Cbl–CIN85–endophilin ternary complex","CIN85–SLP65 BCR signalosome"],"partners":["CBL","CBLB","SH3GL2","SLP65","DNM2","EGFR","PHD2","ANLN"],"other_free_text":[]},"mechanistic_narrative":"SH3KBP1 (CIN85) is a multivalent scaffold protein that coordinates receptor trafficking, immune signaling, cytoskeletal remodeling, and cytokinesis by assembling multiprotein complexes through its three SH3 domains, proline-rich region, and coiled-coil oligomerization domain. Its SH3 domains recognize atypical PX(P/A)XXR motifs to constitutively associate with endophilins, Hrs, and Dab2, and upon ligand stimulation it is recruited by Cbl/Cbl-b to activated receptor tyrosine kinases (EGFR, c-MET) and immune receptors (FcεRI, FcγRIIa, D2DR, TβRI), promoting their clathrin-mediated internalization, ESCRT-dependent endosomal sorting, and lysosomal degradation [PMID:11894095, PMID:20551902, PMID:21635887, PMID:26169354]. CIN85 trimerizes via its coiled-coil domain to oligomerize the B cell adaptor SLP65 into pre-formed signalosome condensates that are essential for BCR-induced NF-κB and Ca²⁺ responses — an autoinhibitory intramolecular SH3C:PRM interaction gates this oligomerization and is regulated by serine phosphorylation — and germline SH3KBP1 loss in humans causes X-linked antibody deficiency [PMID:27353366, PMID:38111344, PMID:29636373]. Beyond trafficking and immune signaling, CIN85 inhibits PHD2 prolyl hydroxylase to stabilize HIF-1α, enhances RIG-I antiviral signaling via TRIM25-mediated K63-ubiquitination, facilitates cytokinesis through anillin–SEPT9 interactions, and supports myofiber integrity through dynamin-2 and calnexin binding [PMID:31142511, PMID:39466846, PMID:36044846, PMID:40065183]."},"prefetch_data":{"uniprot":{"accession":"Q96B97","full_name":"SH3 domain-containing kinase-binding protein 1","aliases":["CD2-binding protein 3","CD2BP3","Cbl-interacting protein of 85 kDa","Human Src family kinase-binding protein 1","HSB-1"],"length_aa":665,"mass_kda":73.1,"function":"Adapter protein involved in regulating diverse signal transduction pathways. Involved in the regulation of endocytosis and lysosomal degradation of ligand-induced receptor tyrosine kinases, including EGFR and MET/hepatocyte growth factor receptor, through an association with CBL and endophilins. The association with CBL, and thus the receptor internalization, may be inhibited by an interaction with PDCD6IP and/or SPRY2. Involved in regulation of ligand-dependent endocytosis of the IgE receptor. Attenuates phosphatidylinositol 3-kinase activity by interaction with its regulatory subunit (By similarity). May be involved in regulation of cell adhesion; promotes the interaction between TTK2B and PDCD6IP. May be involved in the regulation of cellular stress response via the MAPK pathways through its interaction with MAP3K4. Is involved in modulation of tumor necrosis factor mediated apoptosis. Plays a role in the regulation of cell morphology and cytoskeletal organization. Required in the control of cell shape and migration. Has an essential role in the stimulation of B cell activation (PubMed:29636373)","subcellular_location":"Cytoplasm; Cytoplasm, cytoskeleton; Cytoplasmic vesicle membrane; Synapse, synaptosome; Cell junction, focal adhesion","url":"https://www.uniprot.org/uniprotkb/Q96B97/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/SH3KBP1","classification":"Not Classified","n_dependent_lines":0,"n_total_lines":1208,"dependency_fraction":0.0},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"CAPZB","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/SH3KBP1","total_profiled":1310},"omim":[{"mim_id":"621453","title":"MULTIVESICULAR BODY SUBUNIT 12A; MVB12A","url":"https://www.omim.org/entry/621453"},{"mim_id":"619583","title":"EH DOMAIN-BINDING PROTEIN 1-LIKE 1; EHBP1L1","url":"https://www.omim.org/entry/619583"},{"mim_id":"617322","title":"SH3KBP1-BINDING PROTEIN 1; SHKBP1","url":"https://www.omim.org/entry/617322"},{"mim_id":"610591","title":"RHO GTPase-ACTIVATING PROTEIN 27; ARHGAP27","url":"https://www.omim.org/entry/610591"},{"mim_id":"604241","title":"CD2-ASSOCIATED PROTEIN; CD2AP","url":"https://www.omim.org/entry/604241"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Cytosol","reliability":"Supported"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/SH3KBP1"},"hgnc":{"alias_symbol":["CIN85"],"prev_symbol":[]},"alphafold":{"accession":"Q96B97","domains":[{"cath_id":"2.30.30.40","chopping":"3-57","consensus_level":"high","plddt":92.1855,"start":3,"end":57},{"cath_id":"2.30.30.40","chopping":"103-155","consensus_level":"high","plddt":92.3677,"start":103,"end":155},{"cath_id":"2.30.30.40","chopping":"272-334","consensus_level":"high","plddt":89.7308,"start":272,"end":334}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q96B97","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q96B97-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q96B97-F1-predicted_aligned_error_v6.png","plddt_mean":62.19},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=SH3KBP1","jax_strain_url":"https://www.jax.org/strain/search?query=SH3KBP1"},"sequence":{"accession":"Q96B97","fasta_url":"https://rest.uniprot.org/uniprotkb/Q96B97.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q96B97/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q96B97"}},"corpus_meta":[{"pmid":"11894095","id":"PMC_11894095","title":"Cbl-CIN85-endophilin complex mediates ligand-induced downregulation of EGF receptors.","date":"2002","source":"Nature","url":"https://pubmed.ncbi.nlm.nih.gov/11894095","citation_count":470,"is_preprint":false},{"pmid":"11894096","id":"PMC_11894096","title":"The endophilin-CIN85-Cbl complex mediates ligand-dependent downregulation of c-Met.","date":"2002","source":"Nature","url":"https://pubmed.ncbi.nlm.nih.gov/11894096","citation_count":370,"is_preprint":false},{"pmid":"12354621","id":"PMC_12354621","title":"CIN85/CMS family of adaptor molecules.","date":"2002","source":"FEBS letters","url":"https://pubmed.ncbi.nlm.nih.gov/12354621","citation_count":163,"is_preprint":false},{"pmid":"10679202","id":"PMC_10679202","title":"Cloning and characterization of a novel adaptor protein, CIN85, that interacts with c-Cbl.","date":"2000","source":"Biochemical and biophysical research 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Cell physiology","url":"https://pubmed.ncbi.nlm.nih.gov/40445717","citation_count":2,"is_preprint":false},{"pmid":"41208482","id":"PMC_41208482","title":"The MYO1F interactome reveals ASAP1, CD2AP and SH3KBP1 as novel adaptor proteins in podosomes and phagosomes.","date":"2025","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/41208482","citation_count":2,"is_preprint":false},{"pmid":"39955954","id":"PMC_39955954","title":"Feature gene selection and functional validation of SH3KBP1 in infantile hemangioma using machine learning.","date":"2025","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/39955954","citation_count":2,"is_preprint":false},{"pmid":"21400643","id":"PMC_21400643","title":"Lentiviral vector-mediated siRNA knockdown and concurrent rescue of Murine CIN85.","date":"2011","source":"Journal of biochemical and molecular toxicology","url":"https://pubmed.ncbi.nlm.nih.gov/21400643","citation_count":1,"is_preprint":false},{"pmid":"25816594","id":"PMC_25816594","title":"Multiple molecular forms of adaptor protein Ruk/CIN85 specifically associate with different subcellular compartments in human breast adenocarcinoma MCF-7 cells.","date":"2014","source":"Ukrainian biochemical journal","url":"https://pubmed.ncbi.nlm.nih.gov/25816594","citation_count":1,"is_preprint":false},{"pmid":"33672244","id":"PMC_33672244","title":"Exploring Molecular Contacts of MUC1 at CIN85 Binding Interface to Address Future Drug Design Efforts.","date":"2021","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/33672244","citation_count":1,"is_preprint":false},{"pmid":"40740057","id":"PMC_40740057","title":"CIN85 and CD2AP Are Novel Constituents of Dynamic Tubular Recycling Endosomes That Regulate Recycling Upon Recruitment by MICAL-L1.","date":"2025","source":"Traffic (Copenhagen, Denmark)","url":"https://pubmed.ncbi.nlm.nih.gov/40740057","citation_count":1,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":47696,"output_tokens":12402,"usd":0.164559},"stage2":{"model":"claude-opus-4-6","input_tokens":16754,"output_tokens":6091,"usd":0.354067},"total_usd":0.518626,"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\": 2002,\n      \"finding\": \"CIN85 forms a constitutive complex with endophilins via their SH3 domains, and is recruited by Cbl to activated EGF receptors upon EGF stimulation; disruption of the Cbl–CIN85–endophilin complex blocks EGFR internalization and delays receptor degradation without affecting Cbl-directed receptor ubiquitination, demonstrating that this is a mechanistically distinct pathway from ubiquitin ligase activity.\",\n      \"method\": \"Co-immunoprecipitation, dominant-negative interference, internalization assays\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — reciprocal Co-IP and functional block of internalization; replicated in same issue by independent lab (PMID:11894096)\",\n      \"pmids\": [\"11894095\", \"11894096\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"CIN85 binds to Cbl/Cbl-b via a novel polyproline-arginine motif (PXXXPR) in the C-terminus of Cbl; this interaction is enhanced by ligand-induced phosphorylation of Cbl and is required for Cbl-b-mediated internalization of EGFR and PDGFR but is dispensable for receptor polyubiquitination.\",\n      \"method\": \"Co-immunoprecipitation, deletion/mutation analysis, internalization assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — clean epistasis between binding mutant and internalization vs. ubiquitination, replicated across Cbl family members\",\n      \"pmids\": [\"12177062\", \"12874286\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"CIN85 was identified as a novel c-Cbl-interacting protein; the second SH3 domain of CIN85 mediates constitutive association with c-Cbl, which is enhanced after EGF stimulation correlating with c-Cbl tyrosine phosphorylation.\",\n      \"method\": \"Co-immunoprecipitation, domain mapping\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — single Co-IP/domain mapping study; foundational identification of the interaction\",\n      \"pmids\": [\"10679202\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"CIN85 associates with BLNK/SLP65 through its SH3 domains; CIN85 also self-associates through its coiled-coil domain to form tetramers; both the coiled-coil and SH3 domains are required for subcellular localization.\",\n      \"method\": \"Yeast two-hybrid, co-immunoprecipitation, immunostaining\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — single study with yeast two-hybrid confirmed by Co-IP and imaging\",\n      \"pmids\": [\"11071869\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Cbl and Cbl-b monoubiquitinate CIN85 after EGF stimulation; monoubiquitination requires direct CIN85–Cbl interaction, an intact RING finger domain, and a ubiquitin acceptor site in the CIN85 C-terminus; monoubiquitinated CIN85 is degraded in the lysosome together with polyubiquitinated EGFR.\",\n      \"method\": \"In vivo ubiquitination assays, mutagenesis of RING domain and ubiquitin acceptor site, lysosomal inhibitor experiments\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — mutagenesis of both writer (RING) and acceptor site combined with degradation assays; single lab but multiple orthogonal methods\",\n      \"pmids\": [\"12218189\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"CIN85 SH3 domains specifically recognize the PXXXPR motif; individual SH3 domains bind with micromolar affinity while tandem SH3 domains bind with higher stoichiometry; full-length CIN85 simultaneously clusters multiple Cbl molecules, which is essential for ligand-induced stabilization of CIN85–Cbl–EGFR complexes and EGFR lysosomal degradation.\",\n      \"method\": \"Binding assays, peptide competition, affinity measurements, clustering assays in mammalian cells\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — in vitro binding with affinity quantification plus functional cell-based validation with domain mutants\",\n      \"pmids\": [\"12874286\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"The atypical SH3-domain recognition consensus of CIN85 is PX(P/A)XXR; this consensus was validated by mutagenesis and in vitro binding, and allowed identification of novel binding partners synaptojanin 1 and PAK2 confirmed by GST pulldown and Far Western blotting.\",\n      \"method\": \"Target-assisted iterative peptide library screening, mutagenesis, in vitro binding, GST pulldown\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — reconstituted in vitro with mutagenesis confirming binding determinants\",\n      \"pmids\": [\"12829691\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"CIN85 associates with multiple endocytic effectors (SHIP-1, synaptojanin 2B1, ASAP1, ARAP3, Hip1R, STAP1, p115RhoGEF) all via PXXXPR motifs; CIN85 acts as a scaffold clustering these effectors into high-molecular-weight complexes; ASAP1 overexpression promotes EGFR recycling in a CIN85-binding-dependent manner.\",\n      \"method\": \"Co-immunoprecipitation, GST pulldown, functional recycling assay with PXXXPR mutants\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — multiple binding partners identified and functionally validated with binding-deficient mutants\",\n      \"pmids\": [\"15090612\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"Alix/AIP1, which binds CIN85 and endophilins, antagonizes the Cbl–CIN85 complex by weakening CIN85–Cbl interaction and reducing EGFR ubiquitination; Alix overexpression reduces EGFR internalization while siRNA knockdown of Alix promotes internalization and downregulation.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, internalization assays\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — reciprocal gain- and loss-of-function with mechanistic Co-IP showing disrupted Cbl–CIN85 interaction\",\n      \"pmids\": [\"15456872\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"CIN85 and CMS link the T cell surface protein CD2 (via its proline-rich cytoplasmic tail) to the actin-capping protein CAPZ; N-terminal SH3 domains of CIN85 bind CD2 (KD ~100 µM) and CAPZ binds the C-terminal half of CIN85/CMS, providing a direct connection to the actin cytoskeleton.\",\n      \"method\": \"Peptide pulldown, BIAcore binding analysis, overexpression functional assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — quantitative in vitro binding (BIAcore) combined with functional overexpression assays\",\n      \"pmids\": [\"12690097\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"CIN85 (SETA) co-localizes with actin in microfilaments and focal adhesions and with microtubules; it interacts with focal adhesion kinase (FAK) and PYK-2 and promotes cell adhesion in ECIS assays; SETA dimerizes to interact with FAKs.\",\n      \"method\": \"Co-immunoprecipitation, ECIS cell adhesion assay, co-localization microscopy\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — localization tied to functional adhesion readout; indirect interactions, single lab\",\n      \"pmids\": [\"12771190\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"The yeast orthologue of CIN85, Sla1, directly binds the endophilin-like Rvs167; the ubiquitin ligase Rsp5 binds both Sla1 and Rvs167 and monoubiquitinates Rvs167 on Lys481 via its SH3 domain, providing conservation of the CIN85–endophilin–ubiquitin ligase axis.\",\n      \"method\": \"In vitro binding reconstitution, ubiquitination assay, mutagenesis (K481R)\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — direct reconstitution of ubiquitination with mutagenesis confirmation in yeast ortholog\",\n      \"pmids\": [\"14761940\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Sprouty2 binds CIN85 SH3 domains A and C via PXXXPR motifs and acts at the Cbl–CIN85 interface to inhibit EGFR endocytosis; intact Sprouty2–Cbl–CIN85 ternary complex is required for inhibition; Sprouty4, which lacks CIN85-binding motifs, does not inhibit EGFR downregulation.\",\n      \"method\": \"Co-immunoprecipitation, domain mutagenesis, EGFR internalization assays\",\n      \"journal\": \"EMBO reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — mechanistic dissection using binding-deficient mutants with clear functional readout\",\n      \"pmids\": [\"15962011\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"CIN85 is recruited to the FcεRI signaling complex via Cbl after receptor engagement on mast cells and promotes ligand-induced receptor internalization, accelerated sorting into early endosomes and lysosomal delivery; CIN85-overexpressing mast cells are severely impaired in degranulation.\",\n      \"method\": \"Co-immunoprecipitation, confocal microscopy, degranulation assay\",\n      \"journal\": \"Journal of immunology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — functional gain-of-function with internalization and sorting readouts; single lab\",\n      \"pmids\": [\"16177060\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"CIN85 binds MEKK4 via three PXXXPR motifs in MEKK4; disrupting this interaction impairs CIN85-enhanced activation of MKK6 and p38 MAP kinase upon oxidative stress/growth factor stimulation; CIN85 also promotes multi-ubiquitination of MEKK4.\",\n      \"method\": \"Co-immunoprecipitation, kinase assays, ubiquitination assay, dominant-negative mutants\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — Co-IP with functional kinase readout; single lab\",\n      \"pmids\": [\"16256071\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"CIN85 co-localizes with AMAP1 at invadopodia in breast cancer cells; CIN85–AMAP1 binding is required for invasive activity; Cbl monoubiquitinates AMAP1, and this monoubiquitination is required for invasion; siRNA knockdown of CIN85 or Cbl inhibits invasion.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, invasion assays, ubiquitination assays\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — multiple orthogonal methods (Co-IP, siRNA, ubiquitination) with defined invasion phenotype\",\n      \"pmids\": [\"17255943\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"All three SH3 domains of CIN85 bind ubiquitin; ubiquitin binding is at the canonical proline-rich ligand interaction surface and competes with PXXXPR ligands; disruption of ubiquitin binding leads to constitutive CIN85 ubiquitination and elevated basal EGFR ubiquitination, suggesting ubiquitin–SH3 competition regulates Cbl function.\",\n      \"method\": \"NMR structural model, binding assays, mutagenesis, ubiquitination assays\",\n      \"journal\": \"Biochemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — NMR-based structural model with mutagenesis and functional ubiquitination assays\",\n      \"pmids\": [\"18680311\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Drosophila Cindr (sole CD2AP/CIN85 family member) links E-cadherin and the adhesion receptor Roughest with actin-capping proteins (CPα and CPβ); reducing cindr causes defective E-cadherin and Roughest localization, impaired local cell movement, and tissue patterning defects.\",\n      \"method\": \"Genetic loss-of-function, immunofluorescence, protein interaction assays in Drosophila\",\n      \"journal\": \"The Journal of cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — genetic epistasis in Drosophila ortholog with clear cellular phenotype and physical interaction data\",\n      \"pmids\": [\"18362180\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Endogenous CIN85/Ruk localizes predominantly to COPI-coated vesicles of the Golgi complex in a manner dependent on Golgi integrity and intact microtubules; only a small fraction is at clathrin-mediated endocytic compartments.\",\n      \"method\": \"Subcellular fractionation, immunofluorescence with organelle markers\",\n      \"journal\": \"Traffic (Copenhagen, Denmark)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — localization by imaging and fractionation; no functional consequence directly established\",\n      \"pmids\": [\"18266907\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"CIN85 associates with endosomal membranes through binding of its positively charged C-terminal coiled-coil domain to the fusogenic lipid phosphatidic acid; deletion of the coiled-coil domain ablates membrane association and CIN85–c-Cbl interaction, blocking EGFR downregulation.\",\n      \"method\": \"Lipid-binding assay, deletion mutants, EGFR downregulation assays, confocal microscopy\",\n      \"journal\": \"Cell research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — protein-lipid interaction mapped to coiled-coil domain with functional EGFR trafficking readout; single lab\",\n      \"pmids\": [\"19417776\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"CIN85 directly binds the coiled-coil domain of nephrin and podocin; in podocytes CIN85 promotes FGF-4-stimulated nephrin ubiquitination and internalization; co-expression of CD2AP reduces CIN85 binding to nephrin/podocin, indicating functional competition.\",\n      \"method\": \"Binding assays with CIN85 domain mutants, co-immunoprecipitation, internalization assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — domain mapping with functional internalization assay and competition experiments\",\n      \"pmids\": [\"20457601\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"CIN85 interacts with the HEV ORF3 protein, competing with formation of the Cbl–CIN85 complex at growth factor receptors; this reduces CIN85 ubiquitination and delays receptor trafficking to late endosomes/lysosomes, prolonging endomembrane signaling.\",\n      \"method\": \"Co-immunoprecipitation, competitive binding assays, EGFR/c-Met trafficking assays\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — competition binding confirmed with trafficking readout; single lab\",\n      \"pmids\": [\"20130058\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"CIN85 in striatal neurons co-clusters with D2 dopamine receptors and interacts with endocytic regulators dynamin and endophilins; CIN85-deficient (exon 2 knockout) mice show decreased D2DR endocytosis in striatal neurons, elevated striatal dopamine and D2DR levels, and hyperactivity.\",\n      \"method\": \"Conditional knockout mouse model, receptor internalization assay, co-immunoprecipitation, behavioral analysis\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — in vivo genetic model with mechanistic biochemical data linking CIN85 to dopamine receptor endocytosis\",\n      \"pmids\": [\"20551902\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"The SH3 domains of CIN85 bind a PXXXPR motif near the C-terminus of tristetraprolin (TTP/hTTP); CIN85 co-expression leads to increased TTP phosphorylation at Ser66 and Ser93, possibly through co-recruitment of MEKK4.\",\n      \"method\": \"Yeast two-hybrid, co-immunoprecipitation, co-localization, phosphorylation analysis\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — Co-IP and phosphorylation readout; functional consequences not fully defined\",\n      \"pmids\": [\"20221403\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"CIN85 is constitutively associated with SLP65 in resting B cells; absence of the SLP65–CIN85 complex abrogates SLP65 phosphorylation, plasma membrane translocation, BCR-induced Ca2+ and NF-κB responses; live cell imaging confirmed both are components of the BCR primary transducer module.\",\n      \"method\": \"Mass spectrometry interactome, co-immunoprecipitation, live cell imaging, RNAi knockdown with functional signaling assays\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — quantitative MS interactome plus functional signaling assays with clear loss-of-function phenotype\",\n      \"pmids\": [\"21822214\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"B cell-specific deletion of CIN85 impairs IKK-β activation and T cell-independent type II antibody responses; introduction of constitutively active IKK-β corrects the defect, placing CIN85 upstream of IKK-β in BCR-to-NF-κB signaling.\",\n      \"method\": \"Conditional knockout (Mb1-cre), in vivo immunization, in vitro IKK-β kinase assay, rescue experiment\",\n      \"journal\": \"The Journal of experimental medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — genetic epistasis via constitutively active IKK-β rescue in conditional KO; robust in vivo phenotype\",\n      \"pmids\": [\"21708930\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"CIN85 depletion decreases EGF-induced EGFR ubiquitination and delays degradation while increasing EGF recycling; CIN85 is constitutively associated with Hrs; overexpression of a dominant-negative CIN85 (three SH3 domains) inhibits EGF internalization, whereas RNAi knockdown of CIN85 does not, suggesting CIN85 functions primarily in endosomal sorting rather than initial internalization.\",\n      \"method\": \"RNAi knockdown, dominant-negative overexpression, ubiquitination assays, recycling/degradation assays, co-immunoprecipitation\",\n      \"journal\": \"Experimental cell research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — complementary gain- and loss-of-function with multiple trafficking readouts distinguishing internalization from sorting\",\n      \"pmids\": [\"21635887\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"CIN85 overexpression inhibits FcεRI-induced Syk signaling and reduces Syk protein levels via the ubiquitin-proteasome pathway through c-Cbl; CIN85 overexpression limits Cbl binding to the negative regulator Sts1, while CIN85 knockdown promotes Cbl–Sts1 complex formation.\",\n      \"method\": \"Overexpression/knockdown, proteasome inhibitor rescue, co-immunoprecipitation\",\n      \"journal\": \"Journal of immunology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — mechanistic pathway defined by proteasome inhibitor rescue and Sts1 competition; single lab\",\n      \"pmids\": [\"17675467\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"CIN85 modulates FcγRIIa downregulation in neutrophils in concert with c-Cbl; siRNA knockdown of CIN85 prevents FcγRIIa ubiquitination and degradation and increases IgG-mediated phagocytosis; CIN85 is a PKC substrate and classical PKCs positively regulate FcγRIIa ubiquitination and degradation.\",\n      \"method\": \"siRNA knockdown, co-immunoprecipitation, confocal microscopy, PKC inhibitor experiments\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — loss-of-function with defined receptor degradation phenotype and PKC mechanistic link; single lab\",\n      \"pmids\": [\"21372129\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"CD2AP regulates SUMOylation of full-length CIN85 in podocytes; CIN85 is SUMOylated by SUMO-1, -2, and -3; SUMOylation is enhanced by CD2AP; mutation of Lys598 abolishes SUMOylation and increases CIN85 binding to nephrin.\",\n      \"method\": \"SUMO modification assays, mutagenesis (K598R), co-immunoprecipitation\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — site-specific mutagenesis of SUMO acceptor combined with functional binding assay; establishes CD2AP as a SUMO-writer regulator\",\n      \"pmids\": [\"22203040\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"SHKBP1 constitutively binds the SH3 domains of CIN85 via PXXXPR motifs, competing with c-Cbl; SHKBP1 binding prevents CIN85 translocation to EGFR-containing vesicles and reduces EGFR degradation, enhancing EGF-induced signaling.\",\n      \"method\": \"Co-immunoprecipitation, domain competition, EGFR trafficking/degradation assays\",\n      \"journal\": \"Cell biochemistry and function\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — single lab, Co-IP with functional trafficking readout\",\n      \"pmids\": [\"21830225\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"EGF stimulation induces Src-mediated tyrosine phosphorylation of CIN85; phospho-CIN85 interacts with Rab5-positive early endosomes and mediates EGFR sequestration into multivesicular bodies for degradation; disruption of Src-dependent CIN85 phosphorylation impairs EGFR ubiquitination and MVB sorting.\",\n      \"method\": \"Phosphorylation assays, co-immunoprecipitation with Rab5, dominant-negative and kinase inhibitor experiments, MVB sorting assays\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — identifies Src as the writer kinase phosphorylating CIN85 and links this to endosomal sorting with subcellular localization data\",\n      \"pmids\": [\"22833562\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"CIN85 interacts with the CIN85-interacting protein dynamin 2 (Dyn2) in an EGF-stimulation-dependent manner; disruption of the CIN85–Dyn2 interaction causes internalized EGFR to accumulate in aberrantly elongated late endosomal tubules and sustains downstream signaling, identifying a novel late-endocytic role for CIN85.\",\n      \"method\": \"Co-immunoprecipitation, dominant-negative disruption, confocal microscopy, signaling assays\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — direct interaction confirmed, disruption gives specific morphological and signaling phenotype\",\n      \"pmids\": [\"20711168\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"CIN85/CIN85 and CMS can crosslink F-actin into bundles via their proline-rich and coiled-coil domains; removal of these domains reduces cell migration; CMS and CIN85 form heterotypic complexes via their coiled-coil domains.\",\n      \"method\": \"F-actin binding/bundling assays, domain deletion mutants, migration assays\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — in vitro F-actin assay with functional migration readout; single lab\",\n      \"pmids\": [\"17606992\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"MUC1 associates with CIN85 at invadopodia-like structures; siRNA-mediated silencing of CIN85 reduces breast cancer cell migration and invasion in vitro and lung metastasis in vivo; MUC1 overexpression partially rescues CIN85-depletion-reduced invasion.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, invasion/migration assays, in vivo metastasis model\",\n      \"journal\": \"Oncotarget\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — Co-IP with functional loss-of-function invasion and in vivo metastasis data; single lab\",\n      \"pmids\": [\"24072600\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"TβRI interacts with CIN85 SH3 domains in a TGFβ- and TRAF6-dependent manner; CIN85 knockdown causes TβRI accumulation in intracellular compartments and diminishes Smad2 phosphorylation; CIN85 overexpression increases TβRI at the cell surface via a Rab11-dependent recycling pathway.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, Rab11 dominant-negative, Smad2 phosphorylation assays, surface biotinylation\",\n      \"journal\": \"The Journal of cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — mechanistic dissection with dominant-negative Rab11 placing CIN85 in recycling pathway; multiple orthogonal methods\",\n      \"pmids\": [\"26169354\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"CIN85 trimerizes via its C-terminal coiled-coil domain; trimeric CIN85 oligomerizes SLP65 via its multiple SH3 domains interacting with SLP65 PRMs, propagating condensate formation; this oligomeric signaling complex pre-assembles in resting B cells and is required for efficient BCR signal initiation.\",\n      \"method\": \"NMR structural analysis, biochemical oligomerization assays, B cell signaling functional assays\",\n      \"journal\": \"Science signaling\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — structural (NMR) plus biochemical reconstitution with functional B cell validation\",\n      \"pmids\": [\"27353366\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"CIN85/RukL knockout in diabetic mice prevents nephrin internalization, preserves nephrin surface expression on the slit diaphragm, and reduces proteinuria; conversely, CIN85 overexpression in zebrafish induces edema and filtration barrier disruption.\",\n      \"method\": \"Knockout mouse model (diabetic), zebrafish overexpression, nephrin surface expression, proteinuria measurement\",\n      \"journal\": \"Diabetes\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — in vivo genetic models (mouse KO and zebrafish OE) with clear functional readout\",\n      \"pmids\": [\"27531950\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Germline deletion of CIN85 in humans causes X-linked antibody deficiency with intact immune cell development but intrinsic B cell defects in NF-κB activation and CD86 upregulation downstream of BCR, establishing non-redundant in vivo roles of CIN85 in humoral immunity.\",\n      \"method\": \"Human germline genetics, B cell functional assays (NF-κB activation, CD86 expression)\",\n      \"journal\": \"The Journal of experimental medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — human loss-of-function genetics with mechanistic cell-type-specific signaling readouts\",\n      \"pmids\": [\"29636373\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"CIN85 is a novel binding partner of PHD2 (but not PHD1 or PHD3); the N-terminal SH3 domains of CIN85 interact with the proline-arginine-rich N-terminus of PHD2, inhibiting PHD2 hydroxylase activity and thereby stabilizing HIF-1α; CRISPR disruption of the CIN85–PHD2 interaction in cells impairs tumor growth and migration.\",\n      \"method\": \"Co-immunoprecipitation, PHD2 enzymatic activity assay, CRISPR/Cas9 editing, in vivo xenograft\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — enzymatic inhibition assay combined with CRISPR editing and in vivo functional validation\",\n      \"pmids\": [\"31142511\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"After TCR stimulation, CIN85 is recruited to the TCR signaling complex and limits T cell activation; CIN85-deficient T cells show enhanced Zap70, SLP76, and ERK phosphorylation; the inhibitory function requires CIN85 SH3 and PR domains and involves association with the phosphatase Sts-2.\",\n      \"method\": \"T cell-specific conditional knockout, signaling analysis (phospho-western), co-immunoprecipitation, domain mutagenesis\",\n      \"journal\": \"Science signaling\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — conditional KO combined with domain mutants identifying Sts-2 as the effector\",\n      \"pmids\": [\"30723173\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"CIN85 localizes to the intercellular bridge during cytokinesis; CIN85 directly interacts with the N-terminal region of anillin and SEPT9, facilitating SEPT9-containing filament localization to the ICB plasma membrane; loss of CIN85 impairs cytokinesis timing and robustness.\",\n      \"method\": \"siRNA knockdown, co-immunoprecipitation, live cell imaging, direct in vitro binding\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — direct binding reconstitution combined with live imaging and loss-of-function cytokinesis phenotype\",\n      \"pmids\": [\"36044846\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"ASAP2 directly binds CIN85, disrupting its interaction with c-MET and thereby antagonizing CIN85-induced c-MET internalization and lysosomal degradation; CIN85 knockdown rescues the inhibitory effects of ASAP2 knockdown on HGF/c-MET signaling.\",\n      \"method\": \"Co-immunoprecipitation, cycloheximide chase, siRNA epistasis knockdown experiments\",\n      \"journal\": \"Experimental hematology & oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — epistasis by double knockdown with receptor degradation/signaling readout; single lab\",\n      \"pmids\": [\"37061723\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"An intramolecular SH3C:PRM interaction within CIN85 (between C-terminal SH3 domain and adjacent PRM) autoinhibits CIN85 valency toward SLP65; phosphorylation of a serine residue adjacent to the PRM modulates this intramolecular interaction; disrupting it impairs SLP65/CIN85 condensate formation, membrane recruitment of CIN85, and BCR-induced Ca2+ mobilization.\",\n      \"method\": \"High-resolution NMR, mutagenesis, B cell functional assays (Ca2+ flux, membrane recruitment)\",\n      \"journal\": \"Journal of the American Chemical Society\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — NMR structural validation of intramolecular interaction with phosphorylation-regulatory mechanism confirmed by B cell functional assays\",\n      \"pmids\": [\"38111344\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"SH3KBP1 enhances RIG-I signal transduction by increasing K63-linked polyubiquitination through interaction with E3 ligase TRIM25; PRRSV NSP2 induces autophagic degradation of SH3KBP1 via the third PVPAPR motif in NSP2; Sh3kbp1−/− mice are more susceptible to VSV infection with reduced serum IFN-β.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assays, knockout mouse model, VSV infection, autophagy assays\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — knockout mouse with infection phenotype, ubiquitination assay identifying TRIM25 as writer, and mechanism of viral antagonism\",\n      \"pmids\": [\"39466846\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"SH3KBP1 N-terminus binds dynamin-2 and C-terminus associates with ER through calnexin; SH3KBP1 is required for myoblast fusion, myonuclear positioning, and myotube elongation; SH3KBP1 contributes to triad formation and excitation-contraction coupling in mature muscle fibers.\",\n      \"method\": \"siRNA screen, co-immunoprecipitation, domain mapping, excitation-contraction coupling assays, Dnm2 mutant mouse model\",\n      \"journal\": \"EMBO reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — multiple binding partners identified by domain mapping combined with functional siRNA phenotypes and in vivo mouse model\",\n      \"pmids\": [\"40065183\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"CIN85 is recruited to tubular recycling endosomes (TREs) via SH3-domain interaction with MICAL-L1; depletion of CIN85 impairs recycling of CD98 cargo; CIN85 likely regulates TRE function through effects on the actin cytoskeleton.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, fluorescence live imaging, recycling assays\",\n      \"journal\": \"Traffic (Copenhagen, Denmark)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — localization via Co-IP and imaging tied to recycling functional assay; single lab\",\n      \"pmids\": [\"40740057\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"SH3KBP1 is recruited to bleb necks where it interacts with septins and recruits PI3K via liquid-liquid phase separation; the Septin–SH3KBP1–PI3K axis establishes differential PI(3,4,5)P3 enrichment at bleb membranes and confers anoikis resistance.\",\n      \"method\": \"Fluorescence microscopy, co-immunoprecipitation, phase-separation assays, PI assays, anoikis resistance functional assays\",\n      \"journal\": \"American journal of physiology. Cell physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — phase-separation behavior and PI distribution linked to functional anoikis readout; single recent lab\",\n      \"pmids\": [\"40445717\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"CIN85 associates with disabled-2 (Dab2) via all three SH3 domains binding the PKPAPR motif in Dab2; this association recruits CIN85 to clathrin-coated structures; upon growth factor stimulation, Dab2 and clathrin dissociate from CIN85, enabling Cbl binding to CIN85.\",\n      \"method\": \"Co-immunoprecipitation, domain mapping, clathrin association assays\",\n      \"journal\": \"FEBS letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — single Co-IP study demonstrating dynamic switching of CIN85 binding partners during endocytosis\",\n      \"pmids\": [\"14596919\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"Multiple domains of CIN85 (SH3 domains and coiled-coil) interact with the p85α regulatory subunit of PI3K; the SH3 domain of p85α is required to outcompete intramolecular SH3–PRR interaction within CIN85; homodimerization via coiled-coil affects both intra- and intermolecular CIN85 interactions.\",\n      \"method\": \"GST pulldown, domain deletion/mutation analyses, binding competition assays\",\n      \"journal\": \"Journal of molecular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — pulldown study with domain mapping; single lab\",\n      \"pmids\": [\"15476827\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"CIN85 is localized at synapses; its first SH3 domain and C-terminal region bind the proline-rich region and N-terminal region of dendrin, respectively; CIN85, dendrin, and S-SCAM form a ternary complex in vitro that facilitates recruitment of dendrin and S-SCAM to CIN85-positive vesicle-like structures.\",\n      \"method\": \"Yeast two-hybrid, co-immunoprecipitation, immunocytochemistry, subcellular fractionation, in vitro complex reconstitution\",\n      \"journal\": \"Journal of biochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — ternary complex reconstituted in vitro with localization data; single lab\",\n      \"pmids\": [\"16751601\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Intersectin 1 (ITSN1) constitutively forms a complex with CIN85/Ruk independent of EGF stimulation; the interaction is mediated by ITSN1 SH3A domain and the third/fourth proline-rich blocks of CIN85; both proteins co-localize with c-Cbl in MCF-7 cells.\",\n      \"method\": \"Co-immunoprecipitation, domain mapping, confocal co-localization\",\n      \"journal\": \"Cellular signalling\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 — single Co-IP; functional consequence not defined\",\n      \"pmids\": [\"19166927\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Basic amino acids K645, K646, R648 and R650 in the CIN85 coiled-coil domain are required for interaction with both c-Cbl and phosphatidic acid; their mutation dissociates CIN85 from endosomes and prevents ESCRT assembly on endosomal membranes, blocking EGFR sorting for degradation.\",\n      \"method\": \"Mutagenesis, co-immunoprecipitation, phosphatidic acid binding assay, ESCRT co-IP, EGFR degradation assay\",\n      \"journal\": \"BMC biochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — site-specific mutagenesis with multiple functional readouts; single lab\",\n      \"pmids\": [\"25005938\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"CIN85 interacts with the catalytic subunit of PP2A via a PPKKPRP motif (P3 block) in its proline-rich region; purified PP2Ac-CIN85 complex exhibits suppressed phosphatase activity; disruption of PP2Ac-CIN85 interaction by cell-permeable P3 peptide reduces platelet spreading on fibrinogen and thrombus-related functions.\",\n      \"method\": \"Yeast two-hybrid, mutagenesis, in vitro PP2A phosphatase activity assay, cell-permeable peptide, ECIS cell adhesion\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — in vitro enzymatic assay shows CIN85 suppresses PP2Ac activity; domain mutagenesis confirmed; functional platelet assay\",\n      \"pmids\": [\"27334924\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"SOX10 binds a highly conserved element within an alternative promoter of the Sh3kbp1 locus in Schwann cells; mutation of the SOX10 binding site ablates promoter activity; ectopic SOX10 expression drives endogenous Sh3kbp1 expression in SOX10-negative cells.\",\n      \"method\": \"Reporter gene assay, electrophoretic mobility shift/ChIP-equivalent, ectopic SOX10 expression, mutagenesis\",\n      \"journal\": \"Molecular and cellular neurosciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — transcriptional regulation (not post-translational mechanistic finding) but establishes SOX10 as direct upstream regulator of Sh3kbp1 expression in Schwann cells\",\n      \"pmids\": [\"22037207\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"SH3KBP1 co-localizes with MYO1F, CD2AP, ASAP1, and SH3BP2 at podosomes and phagocytic cups in macrophages/microglia; structural modelling and mutagenesis confirmed multivalent proline-rich motif interactions between SH3KBP1 and the MYO1F SH3 domain.\",\n      \"method\": \"Proximity labelling proteomics, structural modelling, mutagenesis, immunofluorescence\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — proximity labelling combined with mutagenesis and localization; functional consequence not fully resolved\",\n      \"pmids\": [\"41208482\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"SH3KBP1 (CIN85) is a multidomain scaffold/adaptor protein whose three SH3 domains recognize atypical PXXXPR motifs to assemble multiprotein complexes that direct clathrin-mediated endocytosis, endosomal sorting, and lysosomal degradation of receptor tyrosine kinases (EGFR, c-MET, D2DR, FcεRI, FcγRIIa, TβRI) via constitutive association with endophilins and ligand-stimulated recruitment by Cbl/Cbl-b, which also monoubiquitinates CIN85; additionally, CIN85 scaffolds B cell antigen receptor signaling by oligomerizing SLP65 into pre-formed condensates that drive NF-κB and Ca2+ responses, inhibits PHD2 prolyl hydroxylase activity to stabilize HIF-1α, facilitates cytokinesis via anillin–SEPT9 interactions, supports myofiber integrity through dynamin-2 and calnexin binding, and regulates actin dynamics, cell adhesion, platelet function, and innate immune signaling through diverse binding partners.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"SH3KBP1 (CIN85) is a multivalent scaffold protein that coordinates receptor trafficking, immune signaling, cytoskeletal remodeling, and cytokinesis by assembling multiprotein complexes through its three SH3 domains, proline-rich region, and coiled-coil oligomerization domain. Its SH3 domains recognize atypical PX(P/A)XXR motifs to constitutively associate with endophilins, Hrs, and Dab2, and upon ligand stimulation it is recruited by Cbl/Cbl-b to activated receptor tyrosine kinases (EGFR, c-MET) and immune receptors (FcεRI, FcγRIIa, D2DR, TβRI), promoting their clathrin-mediated internalization, ESCRT-dependent endosomal sorting, and lysosomal degradation [PMID:11894095, PMID:20551902, PMID:21635887, PMID:26169354]. CIN85 trimerizes via its coiled-coil domain to oligomerize the B cell adaptor SLP65 into pre-formed signalosome condensates that are essential for BCR-induced NF-κB and Ca²⁺ responses — an autoinhibitory intramolecular SH3C:PRM interaction gates this oligomerization and is regulated by serine phosphorylation — and germline SH3KBP1 loss in humans causes X-linked antibody deficiency [PMID:27353366, PMID:38111344, PMID:29636373]. Beyond trafficking and immune signaling, CIN85 inhibits PHD2 prolyl hydroxylase to stabilize HIF-1α, enhances RIG-I antiviral signaling via TRIM25-mediated K63-ubiquitination, facilitates cytokinesis through anillin–SEPT9 interactions, and supports myofiber integrity through dynamin-2 and calnexin binding [PMID:31142511, PMID:39466846, PMID:36044846, PMID:40065183].\",\n  \"teleology\": [\n    {\n      \"year\": 2000,\n      \"claim\": \"Establishing CIN85 as a Cbl-interacting adaptor resolved the question of how Cbl's endocytic function is coupled to downstream effectors beyond its E3 ligase activity.\",\n      \"evidence\": \"Co-immunoprecipitation and domain mapping in mammalian cells identifying SH3-domain-mediated Cbl binding\",\n      \"pmids\": [\"10679202\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Functional consequence of CIN85–Cbl interaction on receptor trafficking not yet tested\", \"No structural detail of binding interface\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Two concurrent studies demonstrated that CIN85 constitutively complexes with endophilins and is recruited to activated EGFR via Cbl, revealing a mechanistically distinct internalization pathway separable from Cbl's ubiquitin ligase activity.\",\n      \"evidence\": \"Reciprocal Co-IPs, dominant-negative interference, and EGFR internalization assays in mammalian cells by two independent laboratories\",\n      \"pmids\": [\"11894095\", \"11894096\", \"12177062\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether CIN85 functions in initial internalization versus endosomal sorting was unresolved\", \"Structural basis of PXXXPR recognition unknown\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Discovery that Cbl monoubiquitinates CIN85 and that monoubiquitinated CIN85 is co-degraded with polyubiquitinated EGFR in lysosomes established a feedback mechanism linking the scaffold to receptor fate.\",\n      \"evidence\": \"In vivo ubiquitination assays with RING-domain and ubiquitin-acceptor-site mutagenesis, lysosomal inhibitor rescue\",\n      \"pmids\": [\"12218189\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Identity of the monoubiquitinated lysine residue in CIN85 not mapped\", \"Physiological relevance of CIN85 co-degradation unclear\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Definition of the PX(P/A)XXR consensus for CIN85 SH3 domains and demonstration that tandem SH3 domains cluster multiple Cbl molecules explained how CIN85 valency drives ligand-induced complex stabilization and receptor downregulation.\",\n      \"evidence\": \"Iterative peptide library screening, in vitro affinity measurements, mutagenesis, and functional clustering assays in mammalian cells; identification of new partners synaptojanin-1, PAK2, and Dab2\",\n      \"pmids\": [\"12874286\", \"12829691\", \"14596919\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"High-resolution structure of SH3:PXXXPR complex not determined\", \"How ligand-gated switching between Dab2 and Cbl binding is regulated remained unclear\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Linking CIN85 to the actin cytoskeleton through direct CAPZ binding and to focal adhesions through FAK/PYK-2 expanded its role beyond endocytosis to cell adhesion and actin dynamics.\",\n      \"evidence\": \"BIAcore quantitative binding to CD2/CAPZ, F-actin bundling assays, ECIS adhesion assays, and co-localization with focal adhesion markers\",\n      \"pmids\": [\"12690097\", \"12771190\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"In vivo relevance of CIN85–CAPZ axis to T cell function not tested\", \"Whether F-actin bundling is CIN85-specific or shared with CMS not resolved\"]\n    },\n    {\n      \"year\": 2004,\n      \"claim\": \"Identification of a broad spectrum of PXXXPR-containing endocytic effectors (SHIP-1, ASAP1, ARAP3, Hip1R) binding CIN85, and conservation of the CIN85–endophilin–ubiquitin ligase axis in yeast (Sla1–Rvs167–Rsp5), established CIN85 as an evolutionarily conserved endocytic scaffold hub.\",\n      \"evidence\": \"Co-IPs and pulldowns with PXXXPR-mutant controls in mammalian cells; in vitro reconstitution of Rsp5-mediated Rvs167 ubiquitination in yeast\",\n      \"pmids\": [\"15090612\", \"14761940\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Hierarchy and competition among multiple PXXXPR partners for CIN85 SH3 occupancy undefined\", \"Whether yeast Sla1 and mammalian CIN85 are true functional orthologs debated\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Sprouty2 and Alix were identified as negative regulators that compete at the Cbl–CIN85 interface, providing a mechanistic basis for fine-tuning EGFR downregulation through antagonistic complex formation.\",\n      \"evidence\": \"Domain mutagenesis, ternary complex Co-IPs, and reciprocal gain/loss-of-function EGFR internalization assays\",\n      \"pmids\": [\"15962011\", \"15456872\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Physiological contexts in which Sprouty2 versus Alix dominate are unknown\", \"Whether antagonism extends to non-EGFR receptors not tested\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Demonstrating CIN85 recruitment to FcεRI via Cbl in mast cells and its role in receptor internalization/sorting extended the Cbl–CIN85 trafficking paradigm to immune receptors beyond RTKs.\",\n      \"evidence\": \"Co-IP, confocal sorting assays, and degranulation functional readout in mast cells\",\n      \"pmids\": [\"16177060\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether CIN85 also controls FcεRI signaling amplitude independently of internalization unclear\", \"No genetic loss-of-function in mast cells\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"NMR-based discovery that CIN85 SH3 domains bind ubiquitin at the same surface used for PXXXPR ligands revealed a competition mechanism that auto-regulates Cbl-mediated ubiquitination, and Drosophila Cindr genetics confirmed conserved roles in adhesion receptor localization.\",\n      \"evidence\": \"NMR structural modeling with mutagenesis and ubiquitination assays; genetic loss-of-function in Drosophila with E-cadherin/Roughest localization phenotypes\",\n      \"pmids\": [\"18680311\", \"18362180\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Relative affinities of ubiquitin versus PXXXPR for each SH3 domain not quantified in cells\", \"How ubiquitin competition is resolved temporally during trafficking unknown\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"CIN85-knockout mice showed impaired D2 dopamine receptor endocytosis, hyperactivity, and elevated striatal dopamine, providing the first in vivo genetic evidence that CIN85 is a non-redundant endocytic regulator in the brain.\",\n      \"evidence\": \"Conditional exon-2 knockout mouse with receptor internalization assays, neurochemistry, and behavioral analysis\",\n      \"pmids\": [\"20551902\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether CD2AP compensates partially in other tissues not addressed\", \"Molecular mechanism linking CIN85 loss to elevated dopamine release undefined\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Binding of CIN85 to dynamin-2 at late endosomes was shown to prevent aberrant tubulation and sustained signaling, identifying a distinct late-endocytic membrane-scission role for CIN85 beyond early internalization.\",\n      \"evidence\": \"Co-IP, dominant-negative disruption, confocal imaging of elongated endosomal tubules, and signaling assays\",\n      \"pmids\": [\"20711168\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether CIN85 directly stimulates dynamin-2 GTPase activity or acts as a scaffold not resolved\", \"Relative contribution of late-endosomal versus early-endocytic CIN85 to EGFR fate unclear\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"CIN85 was established as a core component of the BCR primary transducer module: it constitutively associates with SLP65, and B-cell-specific CIN85 deletion abrogates IKK-β activation and T-cell-independent antibody responses, placing CIN85 upstream of NF-κB in humoral immunity.\",\n      \"evidence\": \"Quantitative MS interactome, live-cell imaging, conditional Mb1-cre knockout with IKK-β rescue, and in vivo immunization\",\n      \"pmids\": [\"21822214\", \"21708930\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How CIN85 specifically promotes IKK-β activation mechanistically undefined\", \"Contribution of CIN85 to germinal center reactions not examined\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Refined trafficking analysis showed CIN85 depletion delays EGFR degradation and increases recycling without blocking initial internalization, repositioning CIN85's primary function to endosomal sorting rather than plasma-membrane uptake; concurrently, CIN85 was linked to FcγRIIa and Syk degradation in neutrophils and mast cells.\",\n      \"evidence\": \"RNAi versus dominant-negative comparison with EGFR recycling/degradation assays; siRNA in neutrophils with phagocytosis readout\",\n      \"pmids\": [\"21635887\", \"21372129\", \"17675467\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether the sorting versus internalization distinction applies to all CIN85-regulated receptors not tested\", \"Mechanism by which CIN85 recruits ESCRT machinery not defined\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Discovery that CD2AP promotes CIN85 SUMOylation at K598 and that SUMOylation reduces CIN85 binding to nephrin established a post-translational switch controlling slit-diaphragm receptor availability in podocytes.\",\n      \"evidence\": \"SUMO modification assays with K598R mutagenesis and Co-IP with nephrin\",\n      \"pmids\": [\"22203040\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Identity of the SUMO E3 ligase not determined\", \"In vivo relevance of K598 SUMOylation to podocyte function not tested\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Src-mediated tyrosine phosphorylation of CIN85 was identified as a signal directing CIN85 to Rab5-positive early endosomes for EGFR MVB sorting, linking kinase signaling to CIN85's endosomal sorting function.\",\n      \"evidence\": \"Phosphorylation assays, Rab5 Co-IP, Src inhibitor experiments, and MVB sorting assays\",\n      \"pmids\": [\"22833562\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Specific tyrosine residue(s) phosphorylated by Src not mapped\", \"Whether Src phosphorylation of CIN85 affects non-EGFR cargoes unknown\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"CIN85 was shown to promote TβRI surface recycling via Rab11 rather than degradation, revealing that CIN85 can direct receptors toward recycling in a context-dependent manner, opposite to its role in EGFR downregulation.\",\n      \"evidence\": \"Co-IP, siRNA, dominant-negative Rab11, surface biotinylation, and Smad2 phosphorylation assays\",\n      \"pmids\": [\"26169354\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"What determines whether CIN85 routes a receptor for degradation versus recycling unresolved\", \"Role of TRAF6 in CIN85 recruitment to TβRI not fully dissected\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Structural and biochemical demonstration that CIN85 trimerizes via its coiled-coil domain and oligomerizes SLP65 into condensates through multivalent SH3:PRM interactions explained how pre-formed signaling clusters enable rapid BCR signal initiation; separately, CIN85 was shown to inhibit PP2A catalytic activity in platelets, affecting platelet spreading.\",\n      \"evidence\": \"NMR structural analysis, oligomerization reconstitution, B cell signaling assays; yeast two-hybrid, in vitro PP2A activity assay, cell-permeable peptide disruption\",\n      \"pmids\": [\"27353366\", \"27334924\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether CIN85-driven condensates undergo true liquid-liquid phase separation in vivo not proven\", \"In vivo platelet phenotype of CIN85 loss not characterized\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"CIN85 knockout in diabetic mice preserved nephrin surface expression and reduced proteinuria, while zebrafish overexpression disrupted the filtration barrier, providing direct in vivo evidence for CIN85 as a driver of nephrin internalization and diabetic nephropathy pathogenesis.\",\n      \"evidence\": \"Knockout mouse model (diabetic), zebrafish overexpression, nephrin surface quantification, proteinuria measurement\",\n      \"pmids\": [\"27531950\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether CIN85-mediated nephrin internalization is Cbl-dependent in vivo not determined\", \"Applicability to non-diabetic nephropathies untested\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Identification of germline SH3KBP1 loss in humans causing X-linked antibody deficiency with defective NF-κB activation established a non-redundant, disease-causing role for CIN85 in human humoral immunity.\",\n      \"evidence\": \"Human genetic analysis with B cell functional assays (NF-κB, CD86) from affected individuals\",\n      \"pmids\": [\"29636373\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Precise mutation(s) and their structural consequences not fully characterized\", \"Whether T cell or myeloid defects contribute to clinical phenotype not fully resolved\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"CIN85 was shown to directly inhibit PHD2 hydroxylase activity, stabilizing HIF-1α, and to limit T cell activation via the phosphatase Sts-2, revealing non-trafficking functions of CIN85 as an enzyme inhibitor and signaling attenuator.\",\n      \"evidence\": \"PHD2 enzymatic activity assay, CRISPR editing, xenograft tumor model; T cell conditional KO with Zap70/SLP76/ERK phosphorylation and Sts-2 Co-IP\",\n      \"pmids\": [\"31142511\", \"30723173\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of PHD2 inhibition by CIN85 SH3 domains not determined\", \"How CIN85 recruits Sts-2 to the TCR complex mechanistically undefined\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Discovery that CIN85 localizes to the intercellular bridge during cytokinesis and directly binds anillin and SEPT9 to facilitate septin filament membrane recruitment revealed a cell-division role entirely distinct from endocytic trafficking.\",\n      \"evidence\": \"Direct in vitro binding reconstitution, live cell imaging of cytokinesis, siRNA loss-of-function\",\n      \"pmids\": [\"36044846\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether CIN85 condensate/oligomerization properties contribute to septin assembly unknown\", \"Upstream signal recruiting CIN85 to the ICB not identified\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"NMR resolution of an intramolecular SH3C:PRM autoinhibitory interaction within CIN85, regulated by serine phosphorylation, explained how CIN85 valency toward SLP65 is gated, controlling condensate formation and BCR-induced Ca²⁺ signaling.\",\n      \"evidence\": \"High-resolution NMR, phospho-mimetic mutagenesis, B cell Ca²⁺ flux and membrane recruitment assays\",\n      \"pmids\": [\"38111344\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Identity of the kinase phosphorylating the regulatory serine not established\", \"Whether autoinhibition regulates CIN85 interactions with non-SLP65 partners untested\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"SH3KBP1 was shown to enhance RIG-I antiviral signaling by promoting TRIM25-mediated K63-ubiquitination, and Sh3kbp1-knockout mice showed increased susceptibility to VSV, expanding CIN85's role to innate antiviral immunity.\",\n      \"evidence\": \"Co-IP, ubiquitination assays, knockout mouse VSV infection model, IFN-β measurement\",\n      \"pmids\": [\"39466846\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether CIN85 scaffolds TRIM25–RIG-I or allosterically activates TRIM25 not distinguished\", \"Relevance to RNA viruses beyond VSV/PRRSV unknown\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Recent studies expanded CIN85 functions to muscle biology (myoblast fusion, triad formation via dynamin-2/calnexin), tubular recycling endosomes (CD98 recycling via MICAL-L1), podosome assembly in macrophages (via MYO1F), and bleb-neck signaling (septin–PI3K phase separation conferring anoikis resistance).\",\n      \"evidence\": \"siRNA screens in myotubes with Dnm2-mutant mouse; live imaging of TRE recycling; proximity labelling proteomics in macrophages; phase-separation and PI assays with anoikis readout\",\n      \"pmids\": [\"40065183\", \"40740057\", \"41208482\", \"40445717\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"In vivo muscle phenotype of CIN85 loss not fully characterized\", \"Whether TRE and bleb-neck functions depend on CIN85 oligomerization unknown\", \"Phase-separation behavior awaits reconstitution with purified components\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"Key unresolved questions include the structural basis of CIN85 SH3 selectivity among dozens of PXXXPR partners, the kinase(s) controlling autoinhibitory phosphorylation, how CIN85 switches between degradative and recycling sorting of different cargoes, and the full physiological spectrum of CIN85 loss in humans beyond antibody deficiency.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No high-resolution crystal/cryo-EM structure of full-length CIN85 or CIN85–Cbl complex\", \"Kinase phosphorylating the autoinhibitory serine unknown\", \"Decision logic routing receptors to degradation versus recycling not defined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [0, 1, 5, 7, 24, 36]},\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [9, 10, 17, 33]},\n      {\"term_id\": \"GO:0008289\", \"supporting_discovery_ids\": [19, 52]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [39, 53]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005768\", \"supporting_discovery_ids\": [19, 26, 31, 32, 52]},\n      {\"term_id\": \"GO:0005794\", \"supporting_discovery_ids\": [18]},\n      {\"term_id\": \"GO:0031410\", \"supporting_discovery_ids\": [0, 13, 48]},\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [35, 47]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [3, 36]},\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [10, 33, 41]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-5653656\", \"supporting_discovery_ids\": [0, 1, 4, 7, 22, 26, 32]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [24, 25, 38, 40, 44]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [35, 39, 14]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [41]},\n      {\"term_id\": \"R-HSA-9609507\", \"supporting_discovery_ids\": [13, 20, 37, 46]}\n    ],\n    \"complexes\": [\n      \"CIN85–endophilin complex\",\n      \"Cbl–CIN85–endophilin ternary complex\",\n      \"CIN85–SLP65 BCR signalosome\"\n    ],\n    \"partners\": [\n      \"CBL\",\n      \"CBLB\",\n      \"SH3GL2\",\n      \"SLP65\",\n      \"DNM2\",\n      \"EGFR\",\n      \"PHD2\",\n      \"ANLN\"\n    ],\n    \"other_free_text\": []\n  }\n}\n```"}