{"gene":"WAS","run_date":"2026-06-11T09:02:06","timeline":{"discoveries":[{"year":1998,"finding":"WASP (and its homolog N-WASP) contains a VCA domain that links upstream signals including Cdc42 (Rho GTPase) to activation of the Arp2/3 complex, leading to actin polymerization; Cdc42 binds WASP/N-WASP to regulate actin dynamics and filopodium formation.","method":"Cell-free system actin depolymerization assay, co-expression in cells, functional domain analysis","journal":"Nature","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro cell-free assay plus cellular co-expression, replicated across multiple subsequent studies","pmids":["9422512"],"is_preprint":false},{"year":1996,"finding":"WASP physically associates with c-Src family protein-tyrosine kinases, particularly Fyn, through SH3 domain-proline-rich region interactions; this interaction was demonstrated both in vitro (affinity chromatography with GST-SH3 fusion proteins) and in vivo (co-immunoprecipitation in human haematopoietic cells).","method":"GST-SH3 affinity chromatography, baculovirus co-expression, co-immunoprecipitation in haematopoietic cells","journal":"Current biology : CB","confidence":"High","confidence_rationale":"Tier 2 / Moderate — reciprocal binding demonstrated in vitro and in vivo with two orthogonal methods in a single study","pmids":["8805332"],"is_preprint":false},{"year":2007,"finding":"WIP (WASP-interacting protein) functions as a chaperone for WASP, protecting it from calpain- and proteasome-mediated degradation; WIP-deficient T cells show severely reduced WASP protein (but not mRNA), and introduction of WIP rescues WASP levels. The WASP-binding domain of WIP is necessary for protection.","method":"WIP knockout mouse T cells, proteasome/calpain inhibitor treatment, WASP protein quantification, WIP domain mapping","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic KO combined with pharmacological inhibitors and domain mapping; replicated in patient cells and mouse cells","pmids":["17213309"],"is_preprint":false},{"year":2008,"finding":"WASP activity is regulated by two hierarchical mechanisms: (1) allosteric autoinhibition where the VCA domain is sequestered, and (2) dimerization/oligomerization that increases Arp2/3 complex affinity by up to 180-fold, greatly enhancing actin assembly. Dimerization explains activation by bacterial effector EspFu and numerous SH3 domain proteins.","method":"In vitro actin assembly assays, biochemical dimerization assays, affinity measurements","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro reconstitution with quantitative biochemical measurements, mechanistic mutagenesis, multiple orthogonal approaches in one study","pmids":["18995840"],"is_preprint":false},{"year":2005,"finding":"Cdc42 and the Src family kinase Lck cooperate synergistically to activate WASP: autoinhibition reduces phosphorylation/dephosphorylation efficiency by 30-40-fold, and Cdc42 reverses this; Cdc42 and the SH3-SH2 module of Lck cooperatively stimulate phosphorylated WASP activity with coupling energy of ~2.4 kcal/mol.","method":"In vitro kinase assays with purified proteins, actin polymerization assays, quantitative biochemical measurements","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — reconstituted in vitro with quantitative mutagenesis and kinetic analysis, multiple orthogonal measurements","pmids":["16293614"],"is_preprint":false},{"year":2007,"finding":"WASp plays an opposing role to PKCtheta in immunological synapse (IS) dynamics: WASp is required for IS reformation after T cell migration, while PKCtheta drives IS symmetry breaking. WASp-deficient T cells display normal initial IS formation but cannot reform the IS after migration unless PKCtheta is inhibited.","method":"WASp-/- and PKCtheta-/- T cell imaging, live cell microscopy of IS formation and reformation in vitro and in vivo","journal":"Cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic KO phenotype with live imaging in vitro and in vivo, double mutant epistasis analysis","pmids":["17512410"],"is_preprint":false},{"year":2007,"finding":"Differential regulation of WASP and N-WASP by upstream activators: Cdc42 is a more effective activator of WASP than N-WASP; Rac1 exclusively stimulates N-WASP with greater potency than Cdc42; PIP2 vesicles improve N-WASP but not WASP actin nucleation; Nck1/2 are the most potent activators of both with distinct effects.","method":"In vitro actin nucleation assays with purified full-length human WASP and N-WASP from mammalian cells, pyrene actin polymerization assays","journal":"Biochemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — reconstituted in vitro with purified full-length proteins, multiple activators tested systematically, single lab","pmids":["17302440"],"is_preprint":false},{"year":2012,"finding":"WASP is ubiquitylated following TCR activation; tyrosine phosphorylation of WASP at Y291 recruits E3 ligases Cbl-b and c-Cbl, which ubiquitylate WASP at lysine residues K76 and K81 in the WH1 domain. Disruption of ubiquitylation causes WASP accumulation and altered actin dynamics.","method":"TCR stimulation of T cells, ubiquitylation assays, site-directed mutagenesis of K76/K81 and Y291, co-immunoprecipitation of Cbl-b/c-Cbl","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — identification of specific ubiquitylation sites with mutagenesis, E3 ligase identification by co-IP, functional actin phenotype readout","pmids":["22665495"],"is_preprint":false},{"year":2014,"finding":"Hck (p61 isoform), a leukocyte-predominant Src family kinase, is the primary tyrosine kinase that phosphorylates WASP in macrophages, regulating phagocytosis, chemotaxis, podosome dynamics, matrix degradation, and transendothelial migration. Hck-/- macrophages show severely reduced WASP tyrosine phosphorylation.","method":"Hck-/- bone marrow-derived macrophages, Hck siRNA knockdown, functional assays (phagocytosis, chemotaxis, matrix degradation), CX3CL1 and FcγR stimulation","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Moderate — genetic KO and RNAi knockdown with multiple functional readouts, kinase-substrate relationship established","pmids":["24482227"],"is_preprint":false},{"year":2014,"finding":"Tyrosine phosphorylation of WIP triggers release of WASP from the WIP-WASP complex, leading to rapid WASP degradation, podosome disruption, and failure to degrade extracellular matrix. Bruton's tyrosine kinase (Btk) was identified as a regulator of WIP tyrosine phosphorylation.","method":"WIP knockdown with phosphomimic expression, WASP stability assays, podosome lifetime measurements, matrix degradation assays, kinase inhibitor screen, Btk identification","journal":"Journal of cell science","confidence":"High","confidence_rationale":"Tier 2 / Moderate — knockdown rescue experiments with phosphomimics, multiple functional readouts, kinase identification by inhibitor screen","pmids":["25413351"],"is_preprint":false},{"year":2018,"finding":"Gain-of-function WASp mutations causing X-linked neutropenia (XLN) result in constitutively open WASp conformation due to phosphorylation of tyrosine-293 combined with plasma membrane localization, rendering WASp activity less dependent on PI3K regulation and producing hyperactive neutrophils with increased actin dynamics.","method":"XLN patient neutrophils, two XLN mouse models, molecular analysis of WASp conformation, tyrosine phosphorylation analysis, actin dynamics measurements, PI3K inhibition","journal":"The Journal of clinical investigation","confidence":"High","confidence_rationale":"Tier 2 / Strong — patient cells plus two mouse models, molecular mechanism of constitutive activation defined, multiple orthogonal methods","pmids":["30124469"],"is_preprint":false},{"year":2018,"finding":"WASP and WIP are tumor suppressors in T cell lymphoma; in their absence, active GTP-bound CDC42 is increased, and deletion of one CDC42 allele is sufficient to impair lymphoma growth. ALK-driven STAT3 and C/EBP-β suppress WASP and WIP expression in ALK+ ALCL.","method":"WASP/WIP-deficient mice, lymphoma acceleration assays, CDC42-GTP pulldown, CDC42 heterozygous deletion rescue, ALK inhibitor experiments","journal":"Nature medicine","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic mouse models with epistasis (CDC42 allele rescue), molecular pathway dissection, multiple orthogonal approaches","pmids":["30510251"],"is_preprint":false},{"year":2021,"finding":"WASP enriches to sites of inward membrane deformation (invaginations) caused by substrate topology, preferentially at their necks, in a front-biased manner set by Cdc42. WASP facilitates Arp2/3 recruitment and local actin assembly at these sites, coupling substrate features to the cytoskeleton; WASP KO cells are defective at topology-directed migration.","method":"WASP KO neutrophils, superresolution imaging, live cell imaging on 3D substrates, Arp2/3 localization assays, Cdc42 manipulation","journal":"The Journal of cell biology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — genetic KO with superresolution imaging, functional migration assay, mechanistic relationship between substrate topology, Cdc42, and WASP established","pmids":["34964841"],"is_preprint":false},{"year":2019,"finding":"WASP contributes to maintenance of front-rear cell polarity by restricting active Rac to the cell front through its CRIB motif; WASP-lacking cells inappropriately activate Rac at the rear. The CRIB motif is required for restricting Rac activity but not for WASP localization to clathrin pits or Arp2/3 activation during endocytosis.","method":"Dictyostelium WASP KO, WASP CRIB mutants, FRET-based Rac activity reporters, live cell imaging, endocytosis assays","journal":"Current biology : CB","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic KO with CRIB mutants and Rac activity reporters; Dictyostelium model, not mammalian","pmids":["31786060"],"is_preprint":false},{"year":2003,"finding":"In Drosophila, Abi protein binds WASP through its C-terminal domain and acts as a potent stimulator of WASP-dependent F-actin formation; reduction of abi function causes bristle loss (similar to wasp mutants); this genetic interaction is wasp-specific and not wave-dependent.","method":"Drosophila genetics (abi and wasp mutants), biochemical binding assay, F-actin formation assay, epistasis analysis","journal":"Development (Cambridge, England)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Drosophila genetic epistasis with biochemical binding confirmation, single lab","pmids":["12900458"],"is_preprint":false},{"year":2005,"finding":"Abi protein activates WASP-mediated actin polymerization in vivo; Abi binds WASP through its C-terminal domain; abi loss-of-function causes bristle defects rescued by wasp but not wave, establishing Abi as a WASP-specific activator in sensory organ development.","method":"Drosophila genetics, biochemical binding, F-actin formation assays, genetic epistasis with wasp and wave mutants","journal":"Nature cell biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic epistasis with biochemical and cell biological validation, multiple orthogonal methods in single study","pmids":["16155589"],"is_preprint":false},{"year":2001,"finding":"WASP (but not N-WASP) is sensitive to cleavage by calpain (a Ca2+-dependent protease) activated in agonist-stimulated platelets; this differential sensitivity suggests calpain-mediated WASP cleavage functions as a Ca2+-dependent switch terminating the surface projection phase of blood cell activation.","method":"Ex vivo broken cell calpain cleavage assays with calpeptin and E64d inhibitors, platelet stimulation experiments","journal":"Blood","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — biochemical reconstitution with specific inhibitors; single lab, single methodology","pmids":["11698281"],"is_preprint":false},{"year":2013,"finding":"Aldolase (fructose-1,6-bisphosphate aldolase) binds WASP and sequesters it, inhibiting WASP/Arp2/3-dependent actin polymerization in vitro; this effect depends on aldolase's actin-binding activity (moonlighting function) rather than its catalytic activity.","method":"In vitro actin polymerization assays, aldolase knockdown in cells, aldolase catalytic and actin-binding mutants, cell motility/spreading assays","journal":"Journal of cellular biochemistry","confidence":"Medium","confidence_rationale":"Tier 1-2 / Moderate — in vitro reconstitution with domain mutants plus cellular knockdown/rescue experiments, single lab","pmids":["23495010"],"is_preprint":false},{"year":2007,"finding":"WIP binds to the N-WASP EVH1 domain through three distinct epitopes (residues 451-461, 461-485, and an extended interface); all three epitopes are required for functional N-WASP binding in cells, with a central polyproline motif occupying the canonical binding site in reversed orientation relative to other EVH1 complexes.","method":"NMR structure of WIP-EVH1 complex, site-directed mutagenesis of WIP epitopes, cellular binding assays","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — NMR structure combined with mutagenesis and cellular validation, single lab but multiple orthogonal methods","pmids":["17229736"],"is_preprint":false},{"year":2007,"finding":"WIP expression is necessary for WASP protein stability and functional expression in human cells; the WIP WASP-binding domain is the minimal region needed to maintain WASP levels, but this minimal domain alone is insufficient to rescue WASP-dependent IL-2 transcriptional activity.","method":"WIP knockdown in T cells, WIP domain deletion experiments, WASP protein quantification, NFAT-IL2 reporter assay","journal":"International immunology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — siRNA knockdown with domain rescue experiments and functional readout; single lab, two orthogonal assays","pmids":["17205972"],"is_preprint":false},{"year":2016,"finding":"WASP expression specifically in FOXP3+ Tregs is required to prevent Th2-mediated food allergy; conditional deletion of Was in Tregs results in more severe Th2 intestinal inflammation than global WASP deficiency. WASP-deficient Tregs fail to restrain Th2 effector responses, associated with increased GATA3 expression in effector memory FOXP3+ Tregs.","method":"Conditional Was knockout in FOXP3+ Tregs (Cre-loxP), comparison with global WAS KO mice, Th1/Th2/Th17 cytokine profiling, GATA3 expression analysis, germ-free housing experiments","journal":"The Journal of clinical investigation","confidence":"High","confidence_rationale":"Tier 2 / Strong — cell-type-specific conditional KO with epistatic comparison to global KO, multiple immune readouts, germ-free controls","pmids":["27643438"],"is_preprint":false},{"year":2007,"finding":"T cell development requires the combined activity of WASP and N-WASP; double knockout mice show thymic hypocellularity, reduced peripheral T cells, impaired DN-to-DP transition, reduced cycling DN3 cells, and impaired SP cell migration. N-WASP single KO T cells are indistinguishable from WT.","method":"RAG2-deficient blastocyst complementation with DKO ES cells; conditional DKO using Cre-loxP; flow cytometry, migration assays","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — two independent genetic systems (blastocyst complementation and conditional Cre-loxP), multiple phenotypic readouts","pmids":["17878299"],"is_preprint":false},{"year":2012,"finding":"The combined activity of WASP and N-WASP is required for peripheral B cell development (follicular and marginal zone B cells), spreading, migration, antigen uptake, and immune responses to T-independent and T-dependent antigens; single WASP KO B cells show partial defects in migration but not development.","method":"Conditional DKO mice (WASP and N-WASP deleted in B cells), flow cytometry, spreading assays, in vivo homing, antigen uptake, antibody response assays","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 / Strong — conditional double KO with multiple orthogonal functional assays comparing single vs. double KO","pmids":["22411869"],"is_preprint":false},{"year":2007,"finding":"Activation of p61Hck triggers WASP- and Arp2/3-dependent actin comet tail formation on lysosomes, accelerating lysosome motility by 35%; this process requires Cdc42 but not Rac or Rho, and was reconstituted in vitro on beads.","method":"In vitro actin comet tail reconstitution on p61Hck-positive lysosomes and latex beads in phagocyte cytosol, kinase-dead mutants, WASP-/- cells, Arp2/3 inhibition, GTPase inhibitors","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution on beads with specific genetic/pharmacological perturbations, quantitative motility measurements","pmids":["17500055"],"is_preprint":false},{"year":2016,"finding":"In C. elegans, WASP functions downstream of MIG-13/SEM-5(Grb2) to activate the Arp2/3 complex during neuroblast migration; purified SEM-5 and MIG-2 synergistically stimulate WASP-Arp2/3 F-actin branching activity in vitro. WASP mutations enhance migration defects in WAVE-deficient cells, indicating partial compensatory roles.","method":"C. elegans genetics, GFP knockin, in vitro F-actin branching assay with purified proteins, epistasis analysis with WAVE mutants","journal":"Developmental cell","confidence":"High","confidence_rationale":"Tier 1-2 / Moderate — in vitro reconstitution with purified proteins plus genetic epistasis in C. elegans, two orthogonal methods","pmids":["27780040"],"is_preprint":false}],"current_model":"WASP is a hematopoietic-cell-restricted actin nucleation-promoting factor that integrates signals from Cdc42/GTPases, phosphoinositides, SH3-domain adaptors, and tyrosine kinases (principally Hck and Lck/Fyn) through relief of VCA-domain autoinhibition and dimerization-dependent Arp2/3 complex activation; its stability depends on constitutive interaction with WIP (which protects it from calpain and proteasome degradation), its activity is terminated by Cbl-b/c-Cbl-mediated ubiquitylation at K76/K81 following Y291 phosphorylation, and its cellular roles include immunological synapse formation and reformation, podosome assembly, lysosome motility, topology-directed neutrophil migration, front-rear polarity maintenance via Rac restriction, T and B cell development, and Treg-mediated suppression of Th2 responses."},"narrative":{"mechanistic_narrative":"WAS encodes WASP, a hematopoietic actin nucleation-promoting factor that couples upstream GTPase and tyrosine-kinase signals to Arp2/3-dependent actin assembly [PMID:9422512]. Activation is governed by relief of VCA-domain autoinhibition followed by dimerization/oligomerization that raises Arp2/3 affinity up to 180-fold [PMID:18995840]; Cdc42 reverses autoinhibition and cooperates synergistically with the Src-family kinase Lck to stimulate phosphorylated WASP [PMID:9422512, PMID:16293614], and Cdc42 is a more effective WASP activator than is the case for the N-WASP paralog, which is preferentially driven by Rac1 and PIP2 [PMID:17302440]. WASP is engaged by Src-family kinases through SH3–proline-rich interactions [PMID:8805332], with Hck serving as the dominant WASP-phosphorylating kinase in macrophages where it controls phagocytosis, chemotaxis, podosome dynamics, and matrix degradation [PMID:24482227]. WASP stability is enforced by constitutive binding to WIP, which shields it from calpain- and proteasome-mediated degradation [PMID:17213309], while tyrosine phosphorylation of WIP releases WASP and triggers its rapid degradation and podosome loss [PMID:25413351]; WASP activity is terminated by Y291-phosphorylation-dependent recruitment of the E3 ligases Cbl-b and c-Cbl, which ubiquitylate K76/K81 in the WH1 domain [PMID:22665495]. Through these mechanisms WASP drives immunological synapse reformation [PMID:17512410], topology-directed neutrophil migration and front-rear polarity via Cdc42-biased Arp2/3 recruitment [PMID:34964841], lysosome actin-comet motility [PMID:17500055], and T- and B-cell development [PMID:17878299, PMID:22411869]. Gain-of-function mutations producing a constitutively open, membrane-localized WASP cause X-linked neutropenia with hyperactive neutrophils [PMID:30124469], and loss of WASP/WIP acts as a tumor suppressor lesion in T-cell lymphoma through elevated CDC42-GTP [PMID:30510251].","teleology":[{"year":1996,"claim":"Established WASP as a physical effector of Src-family tyrosine kinase signaling, linking it to receptor-proximal kinase cascades in hematopoietic cells.","evidence":"GST-SH3 affinity chromatography and co-immunoprecipitation in haematopoietic cells, principally with Fyn","pmids":["8805332"],"confidence":"High","gaps":["Did not define the functional consequence of phosphorylation on actin output","Did not distinguish which kinase is dominant in which cell type"]},{"year":1998,"claim":"Defined the core output mechanism: WASP's VCA domain transmits Cdc42 GTPase signals to Arp2/3-driven actin polymerization.","evidence":"Cell-free actin depolymerization assays, cellular co-expression, and domain analysis of WASP/N-WASP","pmids":["9422512"],"confidence":"High","gaps":["Did not quantify the affinity gains underlying activation","Did not address dimerization or cell-type-specific regulation"]},{"year":2001,"claim":"Identified WASP as a calpain substrate, proposing a Ca2+-dependent proteolytic switch terminating actin-driven projections, distinguishing it from N-WASP.","evidence":"Ex vivo broken-cell calpain cleavage assays with calpeptin/E64d inhibitors in stimulated platelets","pmids":["11698281"],"confidence":"Medium","gaps":["Single-lab biochemical assay without in vivo confirmation of the switch","Cleavage sites not mapped"]},{"year":2005,"claim":"Resolved how two upstream inputs are integrated, showing Cdc42 and Lck cooperate synergistically and quantifying the coupling energy of WASP activation.","evidence":"In vitro kinase and actin polymerization assays with purified proteins and kinetic analysis","pmids":["16293614"],"confidence":"High","gaps":["In vitro coupling energetics not validated in cells","Did not address downstream termination"]},{"year":2007,"claim":"Established WIP as an obligate WASP chaperone, explaining why WASP protein collapses without WIP and defining a degradation axis via calpain and proteasome.","evidence":"WIP-knockout T cells, proteasome/calpain inhibitors, WASP protein quantification, and WIP domain mapping; NMR structure of the WIP-EVH1 interface","pmids":["17213309","17229736","17205972"],"confidence":"High","gaps":["Minimal WIP WASP-binding domain insufficient to restore WASP-dependent IL-2 transcription","Did not define the signal that releases WASP from WIP"]},{"year":2007,"claim":"Distinguished WASP from N-WASP by their differential GTPase and lipid responsiveness, showing the paralogs are not interchangeable in their upstream wiring.","evidence":"In vitro pyrene actin nucleation assays with purified full-length WASP and N-WASP and multiple activators","pmids":["17302440"],"confidence":"High","gaps":["Single-lab in vitro comparison","Did not test combinatorial activator inputs in cells"]},{"year":2007,"claim":"Assigned WASP a specific cellular task in T cells — reforming the immunological synapse after migration — in opposition to PKCtheta-driven symmetry breaking.","evidence":"WASp-/- and PKCtheta-/- T cell live imaging in vitro and in vivo with double-mutant epistasis","pmids":["17512410"],"confidence":"High","gaps":["Molecular link between WASP actin activity and synapse stability not detailed","Did not connect to the WIP/ubiquitylation regulatory layers"]},{"year":2007,"claim":"Extended WASP function to organelle dynamics, showing Hck-triggered, Cdc42-dependent WASP/Arp2/3 actin comet tails propel lysosomes.","evidence":"In vitro actin comet reconstitution on lysosomes and beads with WASP-/- cells, Arp2/3 inhibition, and GTPase inhibitors","pmids":["17500055"],"confidence":"High","gaps":["In vivo physiological role of lysosome comets not established","Did not address other organelle substrates"]},{"year":2007,"claim":"Demonstrated functional redundancy between WASP and N-WASP in T-cell development, revealing combined requirement masked in single knockouts.","evidence":"RAG2 blastocyst complementation and conditional Cre-loxP double knockout with flow cytometry and migration assays","pmids":["17878299"],"confidence":"High","gaps":["Stage-specific molecular targets of the actin defect not defined","Relative contribution of each paralog per developmental step unresolved"]},{"year":2008,"claim":"Refined the activation model by showing WASP uses two hierarchical mechanisms — autoinhibition relief plus dimerization that boosts Arp2/3 affinity up to 180-fold — unifying diverse activators.","evidence":"In vitro actin assembly, dimerization, and affinity measurements with mechanistic mutagenesis","pmids":["18995840"],"confidence":"High","gaps":["In vivo prevalence of dimeric versus monomeric WASP not measured","Did not specify which physiological ligands drive dimerization in cells"]},{"year":2012,"claim":"Identified the termination switch: Y291 phosphorylation recruits Cbl-b/c-Cbl to ubiquitylate WASP at K76/K81, controlling WASP turnover and actin dynamics after TCR engagement.","evidence":"TCR-stimulated T cells, ubiquitylation assays, K76/K81 and Y291 mutagenesis, and Cbl co-immunoprecipitation","pmids":["22665495"],"confidence":"High","gaps":["Interplay between Cbl-mediated degradation and WIP-mediated protection not resolved","Did not quantify how much active WASP this removes physiologically"]},{"year":2012,"claim":"Showed WASP/N-WASP are also jointly required for peripheral B-cell development, spreading, antigen uptake, and antibody responses, broadening the actin requirement across lymphocyte lineages.","evidence":"Conditional B-cell double knockout mice with spreading, homing, antigen uptake, and antibody response assays","pmids":["22411869"],"confidence":"High","gaps":["Molecular distinction between development and migration defects not mapped","Did not identify the receptor inputs driving each phenotype"]},{"year":2014,"claim":"Named Hck as the primary WASP-phosphorylating kinase in macrophages, tying WASP phosphorylation to phagocytosis, podosomes, and matrix degradation.","evidence":"Hck-/- macrophages and siRNA knockdown with phagocytosis, chemotaxis, and matrix degradation assays under CX3CL1/FcgR stimulation","pmids":["24482227"],"confidence":"High","gaps":["Phosphorylated tyrosine residues not individually mapped here","Redundancy with other Src-family kinases in macrophages not excluded"]},{"year":2014,"claim":"Linked WIP phosphorylation to WASP release and degradation, showing Btk-dependent WIP tyrosine phosphorylation destabilizes WASP and disrupts podosomes.","evidence":"WIP knockdown with phosphomimic rescue, WASP stability and podosome assays, and a kinase-inhibitor screen identifying Btk","pmids":["25413351"],"confidence":"High","gaps":["Direct Btk-WIP phosphorylation not biochemically reconstituted","Coordination with Cbl-mediated WASP ubiquitylation unresolved"]},{"year":2016,"claim":"Assigned a cell-intrinsic immunoregulatory role to WASP in Tregs, showing Treg-specific WASP is required to restrain Th2 responses and prevent food allergy.","evidence":"FOXP3-conditional Was knockout versus global knockout with cytokine profiling, GATA3 analysis, and germ-free controls","pmids":["27643438"],"confidence":"High","gaps":["Mechanistic link between actin regulation and GATA3/Th2 restraint not defined","Did not identify the molecular pathway downstream of WASP in Tregs"]},{"year":2016,"claim":"Demonstrated conserved signaling logic in C. elegans, placing WASP downstream of MIG-13/SEM-5(Grb2) with cooperative activation by SEM-5 and MIG-2 during neuroblast migration.","evidence":"C. elegans genetics, GFP knockin, in vitro F-actin branching with purified proteins, and WAVE epistasis","pmids":["27780040"],"confidence":"High","gaps":["Mammalian relevance of the Grb2-WASP axis not tested","Degree of WASP/WAVE compensation not quantified"]},{"year":2018,"claim":"Defined the molecular basis of X-linked neutropenia: gain-of-function mutations produce constitutively open, membrane-localized WASP that is PI3K-independent and hyperactivates actin dynamics.","evidence":"XLN patient neutrophils and two mouse models with conformation, phosphorylation, and actin-dynamics analysis under PI3K inhibition","pmids":["30124469"],"confidence":"High","gaps":["Long-term consequences of constitutive activation on hematopoiesis not detailed","Relationship to Y291/Y293 termination signaling not fully integrated"]},{"year":2018,"claim":"Established WASP/WIP as tumor suppressors in T-cell lymphoma acting by restraining CDC42-GTP, with ALK-STAT3-C/EBPbeta suppressing their expression.","evidence":"WASP/WIP-deficient mice, CDC42-GTP pulldown, CDC42 heterozygous rescue, and ALK inhibitor experiments","pmids":["30510251"],"confidence":"High","gaps":["Mechanism linking WASP loss to CDC42-GTP accumulation not fully defined","Human ALCL therapeutic relevance not directly tested"]},{"year":2019,"claim":"Showed WASP maintains front-rear polarity by restricting active Rac to the cell front through its CRIB motif, separable from its Arp2/3 and endocytic functions.","evidence":"Dictyostelium WASP knockout, CRIB mutants, FRET Rac reporters, and endocytosis assays","pmids":["31786060"],"confidence":"Medium","gaps":["Demonstrated in Dictyostelium, not mammalian cells","Mechanism by which CRIB restricts Rac not molecularly defined"]},{"year":2021,"claim":"Linked WASP to mechanosensing, showing it enriches at substrate-induced membrane invagination necks in a Cdc42-biased manner to direct topology-guided neutrophil migration.","evidence":"WASP-knockout neutrophils with superresolution and live imaging on 3D substrates and Arp2/3 localization assays","pmids":["34964841"],"confidence":"High","gaps":["Sensor that couples membrane curvature to WASP recruitment not identified","In vivo contribution to tissue migration not measured"]},{"year":null,"claim":"How the protective (WIP), terminating (Cbl ubiquitylation, calpain, WIP phosphorylation) and activating (Cdc42/kinase/dimerization) layers are temporally coordinated at a single actin structure remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No integrated kinetic model of WASP lifecycle from activation to degradation","Cell-type-specific balance of these regulators not mapped","Structural state of WASP during dimerization in cells not directly observed"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[0,3,6,23,24]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[0,3,4]},{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[4]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[10,12]},{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[0,12,23]},{"term_id":"GO:0005764","term_label":"lysosome","supporting_discovery_ids":[23]}],"pathway":[{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[5,20,21,22]},{"term_id":"R-HSA-1266738","term_label":"Developmental Biology","supporting_discovery_ids":[21,22,24]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[10,11]},{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[2,7,9]}],"complexes":["WIP-WASP complex","Arp2/3 complex (activator)"],"partners":["WIPF1","CDC42","LCK","FYN","HCK","CBL","CBLB","ABI1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"A0A0C5B5G6","full_name":"Mitochondrial-derived peptide MOTS-c","aliases":["Mitochondrial open reading frame of the 12S rRNA-c"],"length_aa":16,"mass_kda":2.2,"function":"Regulates insulin sensitivity and metabolic homeostasis (PubMed:25738459, PubMed:33468709). Inhibits the folate cycle, thereby reducing de novo purine biosynthesis which leads to the accumulation of the de novo purine synthesis intermediate 5-aminoimidazole-4-carboxamide (AICAR) and the activation of the metabolic regulator 5'-AMP-activated protein kinase (AMPK) (PubMed:25738459). Protects against age-dependent and diet-induced insulin resistance as well as diet-induced obesity (PubMed:25738459). In response to metabolic stress, translocates to the nucleus where it binds to antioxidant response elements (ARE) present in the promoter regions of a number of genes and plays a role in regulating nuclear gene expression in an NFE2L2-dependent manner and increasing cellular resistance to metabolic stress (PubMed:29983246). Increases mitochondrial respiration and levels of CPT1A and cytokines IL1B, IL6, IL8, IL10 and TNF in senescent cells (PubMed:29886458). Increases activity of the serine/threonine protein kinase complex mTORC2 and reduces activity of the PTEN phosphatase, thus promoting phosphorylation of AKT (PubMed:33554779). This promotes AKT-mediated phosphorylation of transcription factor FOXO1 which reduces FOXO1 activity, leading to reduced levels of MSTN and promotion of skeletal muscle growth (PubMed:33554779). Promotes osteogenic differentiation of bone marrow mesenchymal stem cells via the TGFB/SMAD pathway (PubMed:30468456). 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reports","url":"https://pubmed.ncbi.nlm.nih.gov/31110286","citation_count":24,"is_preprint":false},{"pmid":"24482227","id":"PMC_24482227","title":"Tyrosine phosphorylation of Wiskott-Aldrich syndrome protein (WASP) by Hck regulates macrophage function.","date":"2014","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/24482227","citation_count":23,"is_preprint":false},{"pmid":"21699509","id":"PMC_21699509","title":"Combined effects of THC and caffeine on working memory in rats.","date":"2012","source":"British journal of pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/21699509","citation_count":23,"is_preprint":false},{"pmid":"20585499","id":"PMC_20585499","title":"Regulation of WASp by phosphorylation: Activation or other functions?","date":"2010","source":"Communicative & integrative biology","url":"https://pubmed.ncbi.nlm.nih.gov/20585499","citation_count":23,"is_preprint":false},{"pmid":"38702834","id":"PMC_38702834","title":"An emerging trend in Novel Psychoactive Substances (NPSs): designer THC.","date":"2024","source":"Journal of cannabis research","url":"https://pubmed.ncbi.nlm.nih.gov/38702834","citation_count":22,"is_preprint":false},{"pmid":"14697212","id":"PMC_14697212","title":"FBP11 regulates nuclear localization of N-WASP and inhibits N-WASP-dependent microspike formation.","date":"2004","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/14697212","citation_count":20,"is_preprint":false},{"pmid":"35276333","id":"PMC_35276333","title":"Juvenile hormone as a causal factor for maternal regulation of diapause in a wasp.","date":"2022","source":"Insect biochemistry and molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/35276333","citation_count":20,"is_preprint":false},{"pmid":"18823333","id":"PMC_18823333","title":"Transcriptional repression of the IMD2 gene mediated by the transcriptional co-activator Sub1.","date":"2008","source":"Genes to cells : devoted to molecular & cellular mechanisms","url":"https://pubmed.ncbi.nlm.nih.gov/18823333","citation_count":19,"is_preprint":false},{"pmid":"31786060","id":"PMC_31786060","title":"WASP Restricts Active Rac to Maintain Cells' Front-Rear Polarization.","date":"2019","source":"Current biology : CB","url":"https://pubmed.ncbi.nlm.nih.gov/31786060","citation_count":19,"is_preprint":false},{"pmid":"37084981","id":"PMC_37084981","title":"Selected cannabis terpenes synergize with THC to produce increased CB1 receptor activation.","date":"2023","source":"Biochemical pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/37084981","citation_count":19,"is_preprint":false},{"pmid":"31947970","id":"PMC_31947970","title":"Early Exposure to THC Alters M-Cell Development in Zebrafish Embryos.","date":"2020","source":"Biomedicines","url":"https://pubmed.ncbi.nlm.nih.gov/31947970","citation_count":19,"is_preprint":false},{"pmid":"32360153","id":"PMC_32360153","title":"Wasp venom peptide as a new antichagasic agent.","date":"2020","source":"Toxicon : official journal of the International Society on Toxinology","url":"https://pubmed.ncbi.nlm.nih.gov/32360153","citation_count":19,"is_preprint":false},{"pmid":"32852544","id":"PMC_32852544","title":"THC Regulates Tearing via Cannabinoid CB1 Receptors.","date":"2020","source":"Investigative ophthalmology & visual science","url":"https://pubmed.ncbi.nlm.nih.gov/32852544","citation_count":18,"is_preprint":false},{"pmid":"12917347","id":"PMC_12917347","title":"The critical cis-acting element required for IMD2 feedback regulation by GDP is a TATA box located 202 nucleotides upstream of the transcription start site.","date":"2003","source":"Molecular and cellular biology","url":"https://pubmed.ncbi.nlm.nih.gov/12917347","citation_count":18,"is_preprint":false},{"pmid":"17500055","id":"PMC_17500055","title":"Activation of p61Hck triggers WASp- and Arp2/3-dependent actin-comet tail biogenesis and accelerates lysosomes.","date":"2007","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/17500055","citation_count":18,"is_preprint":false},{"pmid":"25413351","id":"PMC_25413351","title":"Tyrosine phosphorylation of WIP releases bound WASP and impairs podosome assembly in macrophages.","date":"2014","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/25413351","citation_count":18,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":43287,"output_tokens":6093,"usd":0.110628,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":14669,"output_tokens":5850,"usd":0.109798,"stage2_stop_reason":"end_turn"},"total_usd":0.220426,"stage1_batch_id":"msgbatch_01UT1AahFPCAyn2uVweS7A9z","stage2_batch_id":"msgbatch_01HuKWtKNT4Pqj1bc8eEP5RV","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1998,\n      \"finding\": \"WASP (and its homolog N-WASP) contains a VCA domain that links upstream signals including Cdc42 (Rho GTPase) to activation of the Arp2/3 complex, leading to actin polymerization; Cdc42 binds WASP/N-WASP to regulate actin dynamics and filopodium formation.\",\n      \"method\": \"Cell-free system actin depolymerization assay, co-expression in cells, functional domain analysis\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro cell-free assay plus cellular co-expression, replicated across multiple subsequent studies\",\n      \"pmids\": [\"9422512\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1996,\n      \"finding\": \"WASP physically associates with c-Src family protein-tyrosine kinases, particularly Fyn, through SH3 domain-proline-rich region interactions; this interaction was demonstrated both in vitro (affinity chromatography with GST-SH3 fusion proteins) and in vivo (co-immunoprecipitation in human haematopoietic cells).\",\n      \"method\": \"GST-SH3 affinity chromatography, baculovirus co-expression, co-immunoprecipitation in haematopoietic cells\",\n      \"journal\": \"Current biology : CB\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal binding demonstrated in vitro and in vivo with two orthogonal methods in a single study\",\n      \"pmids\": [\"8805332\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"WIP (WASP-interacting protein) functions as a chaperone for WASP, protecting it from calpain- and proteasome-mediated degradation; WIP-deficient T cells show severely reduced WASP protein (but not mRNA), and introduction of WIP rescues WASP levels. The WASP-binding domain of WIP is necessary for protection.\",\n      \"method\": \"WIP knockout mouse T cells, proteasome/calpain inhibitor treatment, WASP protein quantification, WIP domain mapping\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic KO combined with pharmacological inhibitors and domain mapping; replicated in patient cells and mouse cells\",\n      \"pmids\": [\"17213309\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"WASP activity is regulated by two hierarchical mechanisms: (1) allosteric autoinhibition where the VCA domain is sequestered, and (2) dimerization/oligomerization that increases Arp2/3 complex affinity by up to 180-fold, greatly enhancing actin assembly. Dimerization explains activation by bacterial effector EspFu and numerous SH3 domain proteins.\",\n      \"method\": \"In vitro actin assembly assays, biochemical dimerization assays, affinity measurements\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro reconstitution with quantitative biochemical measurements, mechanistic mutagenesis, multiple orthogonal approaches in one study\",\n      \"pmids\": [\"18995840\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Cdc42 and the Src family kinase Lck cooperate synergistically to activate WASP: autoinhibition reduces phosphorylation/dephosphorylation efficiency by 30-40-fold, and Cdc42 reverses this; Cdc42 and the SH3-SH2 module of Lck cooperatively stimulate phosphorylated WASP activity with coupling energy of ~2.4 kcal/mol.\",\n      \"method\": \"In vitro kinase assays with purified proteins, actin polymerization assays, quantitative biochemical measurements\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — reconstituted in vitro with quantitative mutagenesis and kinetic analysis, multiple orthogonal measurements\",\n      \"pmids\": [\"16293614\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"WASp plays an opposing role to PKCtheta in immunological synapse (IS) dynamics: WASp is required for IS reformation after T cell migration, while PKCtheta drives IS symmetry breaking. WASp-deficient T cells display normal initial IS formation but cannot reform the IS after migration unless PKCtheta is inhibited.\",\n      \"method\": \"WASp-/- and PKCtheta-/- T cell imaging, live cell microscopy of IS formation and reformation in vitro and in vivo\",\n      \"journal\": \"Cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic KO phenotype with live imaging in vitro and in vivo, double mutant epistasis analysis\",\n      \"pmids\": [\"17512410\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Differential regulation of WASP and N-WASP by upstream activators: Cdc42 is a more effective activator of WASP than N-WASP; Rac1 exclusively stimulates N-WASP with greater potency than Cdc42; PIP2 vesicles improve N-WASP but not WASP actin nucleation; Nck1/2 are the most potent activators of both with distinct effects.\",\n      \"method\": \"In vitro actin nucleation assays with purified full-length human WASP and N-WASP from mammalian cells, pyrene actin polymerization assays\",\n      \"journal\": \"Biochemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — reconstituted in vitro with purified full-length proteins, multiple activators tested systematically, single lab\",\n      \"pmids\": [\"17302440\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"WASP is ubiquitylated following TCR activation; tyrosine phosphorylation of WASP at Y291 recruits E3 ligases Cbl-b and c-Cbl, which ubiquitylate WASP at lysine residues K76 and K81 in the WH1 domain. Disruption of ubiquitylation causes WASP accumulation and altered actin dynamics.\",\n      \"method\": \"TCR stimulation of T cells, ubiquitylation assays, site-directed mutagenesis of K76/K81 and Y291, co-immunoprecipitation of Cbl-b/c-Cbl\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — identification of specific ubiquitylation sites with mutagenesis, E3 ligase identification by co-IP, functional actin phenotype readout\",\n      \"pmids\": [\"22665495\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Hck (p61 isoform), a leukocyte-predominant Src family kinase, is the primary tyrosine kinase that phosphorylates WASP in macrophages, regulating phagocytosis, chemotaxis, podosome dynamics, matrix degradation, and transendothelial migration. Hck-/- macrophages show severely reduced WASP tyrosine phosphorylation.\",\n      \"method\": \"Hck-/- bone marrow-derived macrophages, Hck siRNA knockdown, functional assays (phagocytosis, chemotaxis, matrix degradation), CX3CL1 and FcγR stimulation\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic KO and RNAi knockdown with multiple functional readouts, kinase-substrate relationship established\",\n      \"pmids\": [\"24482227\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Tyrosine phosphorylation of WIP triggers release of WASP from the WIP-WASP complex, leading to rapid WASP degradation, podosome disruption, and failure to degrade extracellular matrix. Bruton's tyrosine kinase (Btk) was identified as a regulator of WIP tyrosine phosphorylation.\",\n      \"method\": \"WIP knockdown with phosphomimic expression, WASP stability assays, podosome lifetime measurements, matrix degradation assays, kinase inhibitor screen, Btk identification\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — knockdown rescue experiments with phosphomimics, multiple functional readouts, kinase identification by inhibitor screen\",\n      \"pmids\": [\"25413351\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Gain-of-function WASp mutations causing X-linked neutropenia (XLN) result in constitutively open WASp conformation due to phosphorylation of tyrosine-293 combined with plasma membrane localization, rendering WASp activity less dependent on PI3K regulation and producing hyperactive neutrophils with increased actin dynamics.\",\n      \"method\": \"XLN patient neutrophils, two XLN mouse models, molecular analysis of WASp conformation, tyrosine phosphorylation analysis, actin dynamics measurements, PI3K inhibition\",\n      \"journal\": \"The Journal of clinical investigation\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — patient cells plus two mouse models, molecular mechanism of constitutive activation defined, multiple orthogonal methods\",\n      \"pmids\": [\"30124469\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"WASP and WIP are tumor suppressors in T cell lymphoma; in their absence, active GTP-bound CDC42 is increased, and deletion of one CDC42 allele is sufficient to impair lymphoma growth. ALK-driven STAT3 and C/EBP-β suppress WASP and WIP expression in ALK+ ALCL.\",\n      \"method\": \"WASP/WIP-deficient mice, lymphoma acceleration assays, CDC42-GTP pulldown, CDC42 heterozygous deletion rescue, ALK inhibitor experiments\",\n      \"journal\": \"Nature medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic mouse models with epistasis (CDC42 allele rescue), molecular pathway dissection, multiple orthogonal approaches\",\n      \"pmids\": [\"30510251\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"WASP enriches to sites of inward membrane deformation (invaginations) caused by substrate topology, preferentially at their necks, in a front-biased manner set by Cdc42. WASP facilitates Arp2/3 recruitment and local actin assembly at these sites, coupling substrate features to the cytoskeleton; WASP KO cells are defective at topology-directed migration.\",\n      \"method\": \"WASP KO neutrophils, superresolution imaging, live cell imaging on 3D substrates, Arp2/3 localization assays, Cdc42 manipulation\",\n      \"journal\": \"The Journal of cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic KO with superresolution imaging, functional migration assay, mechanistic relationship between substrate topology, Cdc42, and WASP established\",\n      \"pmids\": [\"34964841\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"WASP contributes to maintenance of front-rear cell polarity by restricting active Rac to the cell front through its CRIB motif; WASP-lacking cells inappropriately activate Rac at the rear. The CRIB motif is required for restricting Rac activity but not for WASP localization to clathrin pits or Arp2/3 activation during endocytosis.\",\n      \"method\": \"Dictyostelium WASP KO, WASP CRIB mutants, FRET-based Rac activity reporters, live cell imaging, endocytosis assays\",\n      \"journal\": \"Current biology : CB\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic KO with CRIB mutants and Rac activity reporters; Dictyostelium model, not mammalian\",\n      \"pmids\": [\"31786060\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"In Drosophila, Abi protein binds WASP through its C-terminal domain and acts as a potent stimulator of WASP-dependent F-actin formation; reduction of abi function causes bristle loss (similar to wasp mutants); this genetic interaction is wasp-specific and not wave-dependent.\",\n      \"method\": \"Drosophila genetics (abi and wasp mutants), biochemical binding assay, F-actin formation assay, epistasis analysis\",\n      \"journal\": \"Development (Cambridge, England)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Drosophila genetic epistasis with biochemical binding confirmation, single lab\",\n      \"pmids\": [\"12900458\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Abi protein activates WASP-mediated actin polymerization in vivo; Abi binds WASP through its C-terminal domain; abi loss-of-function causes bristle defects rescued by wasp but not wave, establishing Abi as a WASP-specific activator in sensory organ development.\",\n      \"method\": \"Drosophila genetics, biochemical binding, F-actin formation assays, genetic epistasis with wasp and wave mutants\",\n      \"journal\": \"Nature cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic epistasis with biochemical and cell biological validation, multiple orthogonal methods in single study\",\n      \"pmids\": [\"16155589\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"WASP (but not N-WASP) is sensitive to cleavage by calpain (a Ca2+-dependent protease) activated in agonist-stimulated platelets; this differential sensitivity suggests calpain-mediated WASP cleavage functions as a Ca2+-dependent switch terminating the surface projection phase of blood cell activation.\",\n      \"method\": \"Ex vivo broken cell calpain cleavage assays with calpeptin and E64d inhibitors, platelet stimulation experiments\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — biochemical reconstitution with specific inhibitors; single lab, single methodology\",\n      \"pmids\": [\"11698281\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Aldolase (fructose-1,6-bisphosphate aldolase) binds WASP and sequesters it, inhibiting WASP/Arp2/3-dependent actin polymerization in vitro; this effect depends on aldolase's actin-binding activity (moonlighting function) rather than its catalytic activity.\",\n      \"method\": \"In vitro actin polymerization assays, aldolase knockdown in cells, aldolase catalytic and actin-binding mutants, cell motility/spreading assays\",\n      \"journal\": \"Journal of cellular biochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1-2 / Moderate — in vitro reconstitution with domain mutants plus cellular knockdown/rescue experiments, single lab\",\n      \"pmids\": [\"23495010\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"WIP binds to the N-WASP EVH1 domain through three distinct epitopes (residues 451-461, 461-485, and an extended interface); all three epitopes are required for functional N-WASP binding in cells, with a central polyproline motif occupying the canonical binding site in reversed orientation relative to other EVH1 complexes.\",\n      \"method\": \"NMR structure of WIP-EVH1 complex, site-directed mutagenesis of WIP epitopes, cellular binding assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — NMR structure combined with mutagenesis and cellular validation, single lab but multiple orthogonal methods\",\n      \"pmids\": [\"17229736\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"WIP expression is necessary for WASP protein stability and functional expression in human cells; the WIP WASP-binding domain is the minimal region needed to maintain WASP levels, but this minimal domain alone is insufficient to rescue WASP-dependent IL-2 transcriptional activity.\",\n      \"method\": \"WIP knockdown in T cells, WIP domain deletion experiments, WASP protein quantification, NFAT-IL2 reporter assay\",\n      \"journal\": \"International immunology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — siRNA knockdown with domain rescue experiments and functional readout; single lab, two orthogonal assays\",\n      \"pmids\": [\"17205972\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"WASP expression specifically in FOXP3+ Tregs is required to prevent Th2-mediated food allergy; conditional deletion of Was in Tregs results in more severe Th2 intestinal inflammation than global WASP deficiency. WASP-deficient Tregs fail to restrain Th2 effector responses, associated with increased GATA3 expression in effector memory FOXP3+ Tregs.\",\n      \"method\": \"Conditional Was knockout in FOXP3+ Tregs (Cre-loxP), comparison with global WAS KO mice, Th1/Th2/Th17 cytokine profiling, GATA3 expression analysis, germ-free housing experiments\",\n      \"journal\": \"The Journal of clinical investigation\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — cell-type-specific conditional KO with epistatic comparison to global KO, multiple immune readouts, germ-free controls\",\n      \"pmids\": [\"27643438\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"T cell development requires the combined activity of WASP and N-WASP; double knockout mice show thymic hypocellularity, reduced peripheral T cells, impaired DN-to-DP transition, reduced cycling DN3 cells, and impaired SP cell migration. N-WASP single KO T cells are indistinguishable from WT.\",\n      \"method\": \"RAG2-deficient blastocyst complementation with DKO ES cells; conditional DKO using Cre-loxP; flow cytometry, migration assays\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — two independent genetic systems (blastocyst complementation and conditional Cre-loxP), multiple phenotypic readouts\",\n      \"pmids\": [\"17878299\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"The combined activity of WASP and N-WASP is required for peripheral B cell development (follicular and marginal zone B cells), spreading, migration, antigen uptake, and immune responses to T-independent and T-dependent antigens; single WASP KO B cells show partial defects in migration but not development.\",\n      \"method\": \"Conditional DKO mice (WASP and N-WASP deleted in B cells), flow cytometry, spreading assays, in vivo homing, antigen uptake, antibody response assays\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — conditional double KO with multiple orthogonal functional assays comparing single vs. double KO\",\n      \"pmids\": [\"22411869\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Activation of p61Hck triggers WASP- and Arp2/3-dependent actin comet tail formation on lysosomes, accelerating lysosome motility by 35%; this process requires Cdc42 but not Rac or Rho, and was reconstituted in vitro on beads.\",\n      \"method\": \"In vitro actin comet tail reconstitution on p61Hck-positive lysosomes and latex beads in phagocyte cytosol, kinase-dead mutants, WASP-/- cells, Arp2/3 inhibition, GTPase inhibitors\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution on beads with specific genetic/pharmacological perturbations, quantitative motility measurements\",\n      \"pmids\": [\"17500055\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"In C. elegans, WASP functions downstream of MIG-13/SEM-5(Grb2) to activate the Arp2/3 complex during neuroblast migration; purified SEM-5 and MIG-2 synergistically stimulate WASP-Arp2/3 F-actin branching activity in vitro. WASP mutations enhance migration defects in WAVE-deficient cells, indicating partial compensatory roles.\",\n      \"method\": \"C. elegans genetics, GFP knockin, in vitro F-actin branching assay with purified proteins, epistasis analysis with WAVE mutants\",\n      \"journal\": \"Developmental cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Moderate — in vitro reconstitution with purified proteins plus genetic epistasis in C. elegans, two orthogonal methods\",\n      \"pmids\": [\"27780040\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"WASP is a hematopoietic-cell-restricted actin nucleation-promoting factor that integrates signals from Cdc42/GTPases, phosphoinositides, SH3-domain adaptors, and tyrosine kinases (principally Hck and Lck/Fyn) through relief of VCA-domain autoinhibition and dimerization-dependent Arp2/3 complex activation; its stability depends on constitutive interaction with WIP (which protects it from calpain and proteasome degradation), its activity is terminated by Cbl-b/c-Cbl-mediated ubiquitylation at K76/K81 following Y291 phosphorylation, and its cellular roles include immunological synapse formation and reformation, podosome assembly, lysosome motility, topology-directed neutrophil migration, front-rear polarity maintenance via Rac restriction, T and B cell development, and Treg-mediated suppression of Th2 responses.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"WAS encodes WASP, a hematopoietic actin nucleation-promoting factor that couples upstream GTPase and tyrosine-kinase signals to Arp2/3-dependent actin assembly [#0]. Activation is governed by relief of VCA-domain autoinhibition followed by dimerization/oligomerization that raises Arp2/3 affinity up to 180-fold [#3]; Cdc42 reverses autoinhibition and cooperates synergistically with the Src-family kinase Lck to stimulate phosphorylated WASP [#0, #4], and Cdc42 is a more effective WASP activator than is the case for the N-WASP paralog, which is preferentially driven by Rac1 and PIP2 [#6]. WASP is engaged by Src-family kinases through SH3–proline-rich interactions [#1], with Hck serving as the dominant WASP-phosphorylating kinase in macrophages where it controls phagocytosis, chemotaxis, podosome dynamics, and matrix degradation [#8]. WASP stability is enforced by constitutive binding to WIP, which shields it from calpain- and proteasome-mediated degradation [#2], while tyrosine phosphorylation of WIP releases WASP and triggers its rapid degradation and podosome loss [#9]; WASP activity is terminated by Y291-phosphorylation-dependent recruitment of the E3 ligases Cbl-b and c-Cbl, which ubiquitylate K76/K81 in the WH1 domain [#7]. Through these mechanisms WASP drives immunological synapse reformation [#5], topology-directed neutrophil migration and front-rear polarity via Cdc42-biased Arp2/3 recruitment [#12], lysosome actin-comet motility [#23], and T- and B-cell development [#21, #22]. Gain-of-function mutations producing a constitutively open, membrane-localized WASP cause X-linked neutropenia with hyperactive neutrophils [#10], and loss of WASP/WIP acts as a tumor suppressor lesion in T-cell lymphoma through elevated CDC42-GTP [#11].\",\n  \"teleology\": [\n    {\n      \"year\": 1996,\n      \"claim\": \"Established WASP as a physical effector of Src-family tyrosine kinase signaling, linking it to receptor-proximal kinase cascades in hematopoietic cells.\",\n      \"evidence\": \"GST-SH3 affinity chromatography and co-immunoprecipitation in haematopoietic cells, principally with Fyn\",\n      \"pmids\": [\"8805332\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define the functional consequence of phosphorylation on actin output\", \"Did not distinguish which kinase is dominant in which cell type\"]\n    },\n    {\n      \"year\": 1998,\n      \"claim\": \"Defined the core output mechanism: WASP's VCA domain transmits Cdc42 GTPase signals to Arp2/3-driven actin polymerization.\",\n      \"evidence\": \"Cell-free actin depolymerization assays, cellular co-expression, and domain analysis of WASP/N-WASP\",\n      \"pmids\": [\"9422512\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not quantify the affinity gains underlying activation\", \"Did not address dimerization or cell-type-specific regulation\"]\n    },\n    {\n      \"year\": 2001,\n      \"claim\": \"Identified WASP as a calpain substrate, proposing a Ca2+-dependent proteolytic switch terminating actin-driven projections, distinguishing it from N-WASP.\",\n      \"evidence\": \"Ex vivo broken-cell calpain cleavage assays with calpeptin/E64d inhibitors in stimulated platelets\",\n      \"pmids\": [\"11698281\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-lab biochemical assay without in vivo confirmation of the switch\", \"Cleavage sites not mapped\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Resolved how two upstream inputs are integrated, showing Cdc42 and Lck cooperate synergistically and quantifying the coupling energy of WASP activation.\",\n      \"evidence\": \"In vitro kinase and actin polymerization assays with purified proteins and kinetic analysis\",\n      \"pmids\": [\"16293614\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vitro coupling energetics not validated in cells\", \"Did not address downstream termination\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Established WIP as an obligate WASP chaperone, explaining why WASP protein collapses without WIP and defining a degradation axis via calpain and proteasome.\",\n      \"evidence\": \"WIP-knockout T cells, proteasome/calpain inhibitors, WASP protein quantification, and WIP domain mapping; NMR structure of the WIP-EVH1 interface\",\n      \"pmids\": [\"17213309\", \"17229736\", \"17205972\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Minimal WIP WASP-binding domain insufficient to restore WASP-dependent IL-2 transcription\", \"Did not define the signal that releases WASP from WIP\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Distinguished WASP from N-WASP by their differential GTPase and lipid responsiveness, showing the paralogs are not interchangeable in their upstream wiring.\",\n      \"evidence\": \"In vitro pyrene actin nucleation assays with purified full-length WASP and N-WASP and multiple activators\",\n      \"pmids\": [\"17302440\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Single-lab in vitro comparison\", \"Did not test combinatorial activator inputs in cells\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Assigned WASP a specific cellular task in T cells — reforming the immunological synapse after migration — in opposition to PKCtheta-driven symmetry breaking.\",\n      \"evidence\": \"WASp-/- and PKCtheta-/- T cell live imaging in vitro and in vivo with double-mutant epistasis\",\n      \"pmids\": [\"17512410\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular link between WASP actin activity and synapse stability not detailed\", \"Did not connect to the WIP/ubiquitylation regulatory layers\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Extended WASP function to organelle dynamics, showing Hck-triggered, Cdc42-dependent WASP/Arp2/3 actin comet tails propel lysosomes.\",\n      \"evidence\": \"In vitro actin comet reconstitution on lysosomes and beads with WASP-/- cells, Arp2/3 inhibition, and GTPase inhibitors\",\n      \"pmids\": [\"17500055\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo physiological role of lysosome comets not established\", \"Did not address other organelle substrates\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Demonstrated functional redundancy between WASP and N-WASP in T-cell development, revealing combined requirement masked in single knockouts.\",\n      \"evidence\": \"RAG2 blastocyst complementation and conditional Cre-loxP double knockout with flow cytometry and migration assays\",\n      \"pmids\": [\"17878299\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Stage-specific molecular targets of the actin defect not defined\", \"Relative contribution of each paralog per developmental step unresolved\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Refined the activation model by showing WASP uses two hierarchical mechanisms — autoinhibition relief plus dimerization that boosts Arp2/3 affinity up to 180-fold — unifying diverse activators.\",\n      \"evidence\": \"In vitro actin assembly, dimerization, and affinity measurements with mechanistic mutagenesis\",\n      \"pmids\": [\"18995840\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo prevalence of dimeric versus monomeric WASP not measured\", \"Did not specify which physiological ligands drive dimerization in cells\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Identified the termination switch: Y291 phosphorylation recruits Cbl-b/c-Cbl to ubiquitylate WASP at K76/K81, controlling WASP turnover and actin dynamics after TCR engagement.\",\n      \"evidence\": \"TCR-stimulated T cells, ubiquitylation assays, K76/K81 and Y291 mutagenesis, and Cbl co-immunoprecipitation\",\n      \"pmids\": [\"22665495\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Interplay between Cbl-mediated degradation and WIP-mediated protection not resolved\", \"Did not quantify how much active WASP this removes physiologically\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Showed WASP/N-WASP are also jointly required for peripheral B-cell development, spreading, antigen uptake, and antibody responses, broadening the actin requirement across lymphocyte lineages.\",\n      \"evidence\": \"Conditional B-cell double knockout mice with spreading, homing, antigen uptake, and antibody response assays\",\n      \"pmids\": [\"22411869\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular distinction between development and migration defects not mapped\", \"Did not identify the receptor inputs driving each phenotype\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Named Hck as the primary WASP-phosphorylating kinase in macrophages, tying WASP phosphorylation to phagocytosis, podosomes, and matrix degradation.\",\n      \"evidence\": \"Hck-/- macrophages and siRNA knockdown with phagocytosis, chemotaxis, and matrix degradation assays under CX3CL1/FcgR stimulation\",\n      \"pmids\": [\"24482227\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Phosphorylated tyrosine residues not individually mapped here\", \"Redundancy with other Src-family kinases in macrophages not excluded\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Linked WIP phosphorylation to WASP release and degradation, showing Btk-dependent WIP tyrosine phosphorylation destabilizes WASP and disrupts podosomes.\",\n      \"evidence\": \"WIP knockdown with phosphomimic rescue, WASP stability and podosome assays, and a kinase-inhibitor screen identifying Btk\",\n      \"pmids\": [\"25413351\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct Btk-WIP phosphorylation not biochemically reconstituted\", \"Coordination with Cbl-mediated WASP ubiquitylation unresolved\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Assigned a cell-intrinsic immunoregulatory role to WASP in Tregs, showing Treg-specific WASP is required to restrain Th2 responses and prevent food allergy.\",\n      \"evidence\": \"FOXP3-conditional Was knockout versus global knockout with cytokine profiling, GATA3 analysis, and germ-free controls\",\n      \"pmids\": [\"27643438\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanistic link between actin regulation and GATA3/Th2 restraint not defined\", \"Did not identify the molecular pathway downstream of WASP in Tregs\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Demonstrated conserved signaling logic in C. elegans, placing WASP downstream of MIG-13/SEM-5(Grb2) with cooperative activation by SEM-5 and MIG-2 during neuroblast migration.\",\n      \"evidence\": \"C. elegans genetics, GFP knockin, in vitro F-actin branching with purified proteins, and WAVE epistasis\",\n      \"pmids\": [\"27780040\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mammalian relevance of the Grb2-WASP axis not tested\", \"Degree of WASP/WAVE compensation not quantified\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Defined the molecular basis of X-linked neutropenia: gain-of-function mutations produce constitutively open, membrane-localized WASP that is PI3K-independent and hyperactivates actin dynamics.\",\n      \"evidence\": \"XLN patient neutrophils and two mouse models with conformation, phosphorylation, and actin-dynamics analysis under PI3K inhibition\",\n      \"pmids\": [\"30124469\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Long-term consequences of constitutive activation on hematopoiesis not detailed\", \"Relationship to Y291/Y293 termination signaling not fully integrated\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Established WASP/WIP as tumor suppressors in T-cell lymphoma acting by restraining CDC42-GTP, with ALK-STAT3-C/EBPbeta suppressing their expression.\",\n      \"evidence\": \"WASP/WIP-deficient mice, CDC42-GTP pulldown, CDC42 heterozygous rescue, and ALK inhibitor experiments\",\n      \"pmids\": [\"30510251\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism linking WASP loss to CDC42-GTP accumulation not fully defined\", \"Human ALCL therapeutic relevance not directly tested\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Showed WASP maintains front-rear polarity by restricting active Rac to the cell front through its CRIB motif, separable from its Arp2/3 and endocytic functions.\",\n      \"evidence\": \"Dictyostelium WASP knockout, CRIB mutants, FRET Rac reporters, and endocytosis assays\",\n      \"pmids\": [\"31786060\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Demonstrated in Dictyostelium, not mammalian cells\", \"Mechanism by which CRIB restricts Rac not molecularly defined\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Linked WASP to mechanosensing, showing it enriches at substrate-induced membrane invagination necks in a Cdc42-biased manner to direct topology-guided neutrophil migration.\",\n      \"evidence\": \"WASP-knockout neutrophils with superresolution and live imaging on 3D substrates and Arp2/3 localization assays\",\n      \"pmids\": [\"34964841\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Sensor that couples membrane curvature to WASP recruitment not identified\", \"In vivo contribution to tissue migration not measured\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How the protective (WIP), terminating (Cbl ubiquitylation, calpain, WIP phosphorylation) and activating (Cdc42/kinase/dimerization) layers are temporally coordinated at a single actin structure remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No integrated kinetic model of WASP lifecycle from activation to degradation\", \"Cell-type-specific balance of these regulators not mapped\", \"Structural state of WASP during dimerization in cells not directly observed\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [0, 3, 6, 23, 24]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [0, 3, 4]},\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [4]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [10, 12]},\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [0, 12, 23]},\n      {\"term_id\": \"GO:0005764\", \"supporting_discovery_ids\": [23]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [5, 20, 21, 22]},\n      {\"term_id\": \"R-HSA-1266738\", \"supporting_discovery_ids\": [21, 22, 24]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [10, 11]},\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [2, 7, 9]}\n    ],\n    \"complexes\": [\"WIP-WASP complex\", \"Arp2/3 complex (activator)\"],\n    \"partners\": [\"WIPF1\", \"CDC42\", \"LCK\", \"FYN\", \"HCK\", \"CBL\", \"CBLB\", \"ABI1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}