{"gene":"WASF3","run_date":"2026-06-11T09:02:06","timeline":{"discoveries":[{"year":2005,"finding":"WAVE3/WASF3 localizes to lamellipodia at the leading edge of migrating cells, and its knockdown prevents PDGF-induced lamellipodia formation and cell migration. WAVE3 interacts physically with the PI3K regulatory subunit p85 (mediated by the N-terminal region of WAVE3 and the C-terminal SH2 domain of p85), placing WAVE3 downstream of PI3K signaling. Treatment with PI3K inhibitor LY294002 also abrogated PDGF-induced lamellipodia formation.","method":"Subcellular localization by microscopy, RNA interference knockdown, yeast two-hybrid screen confirmed by co-immunoprecipitation, pharmacological inhibition","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Moderate — reciprocal interaction confirmed by yeast two-hybrid and Co-IP with domain mapping, localization by microscopy, functional KD with defined phenotype","pmids":["15826941"],"is_preprint":false},{"year":2005,"finding":"WAVE3/WASF3 knockdown decreases phospho-p38 MAPK levels (but not phospho-AKT, phospho-ERK, or phospho-JNK) and inhibits expression of MMP-1, MMP-3, and MMP-9 (but not MMP-2), leading to inhibition of cell motility and invasion with increased actin stress fiber formation and reorganization of focal adhesion complexes. WAVE1 and WAVE2 expression levels were not affected by loss of WAVE3.","method":"RNA interference knockdown, Western blotting, invasion assay, actin/focal adhesion staining","journal":"Experimental cell research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — siRNA KD with multiple downstream readouts (kinase phosphorylation, MMP expression, cell morphology), single lab","pmids":["15907837"],"is_preprint":false},{"year":2005,"finding":"WAVE3/WASF3 includes the Scar/WAVE family proteins in the same kinds of protein complexes as WAVE1 and WAVE2, all three isoforms interacting with previously described binding partners, suggesting participation in the same regulatory complexes for actin assembly.","method":"Co-immunoprecipitation, interaction assays with multiple binding partners across isoforms","journal":"BMC cell biology","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Co-IP-based interaction data replicated across three isoforms in single study","pmids":["15752430"],"is_preprint":false},{"year":2005,"finding":"WAVE3 binds directly to LDOC1 through the verprolin homology (VH) domain of WAVE3. WAVE3 expression induces translocation of LDOC1 from the nucleus to the cytoplasm, inhibiting LDOC1-induced apoptosis (which requires nuclear LDOC1 and p53 stabilization). Thus WAVE3 negatively regulates LDOC1 function.","method":"Direct binding assay, ectopic expression, subcellular localization (nuclear/cytoplasmic fractionation and microscopy), apoptosis assay","journal":"Journal of biochemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct binding established with domain mapping (VH domain), functional consequence (apoptosis suppression) demonstrated by localization change; single lab","pmids":["16272576"],"is_preprint":false},{"year":2007,"finding":"c-Abl tyrosine kinase interacts with WAVE3/WASF3 upon PDGF stimulation, and phosphorylates four tyrosine residues on WAVE3. Abl-mediated phosphorylation of WAVE3 is required for stimulation of lamellipodia formation and cell migration. The Abl inhibitor STI-571 abrogates Abl-mediated phosphorylation of WAVE3.","method":"Co-immunoprecipitation, in vitro kinase assay, site-directed mutagenesis of tyrosine residues, pharmacological inhibition (STI-571), lamellipodia/migration assays","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro kinase assay with mutagenesis of phosphorylation sites, co-IP, pharmacological inhibition, and functional migration readout in one study","pmids":["17623672"],"is_preprint":false},{"year":2007,"finding":"Stable shRNA-mediated knockdown of WAVE3 in MDA-MB-231 breast cancer cells reduces Matrigel invasion, lung colony formation after tail-vein injection, and primary tumor growth in orthotopic xenograft models. Suppression of p38 MAPK activity by dominant-negative p38 produces comparable phenotypes, establishing the WAVE3-p38 pathway in metastasis.","method":"shRNA stable knockdown, Matrigel invasion assay, xenograft/orthotopic mouse models, dominant-negative p38 construct","journal":"The American journal of pathology","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic KD with defined phenotype confirmed in vitro and in vivo, pathway placement by dominant-negative epistasis","pmids":["17525277"],"is_preprint":false},{"year":2009,"finding":"miR-200 family microRNAs directly target the 3'-UTR of WAVE3 mRNA and inhibit its expression. miR-200-mediated downregulation of WAVE3 reduces cancer cell invasiveness and causes morphological changes resembling mesenchymal-to-epithelial transition. Re-expression of a miR-200-resistant WAVE3 reverses these effects, confirming specificity.","method":"Luciferase 3'-UTR reporter assay, Western blotting, invasion assay, rescue experiment with miR-200-resistant WAVE3","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Moderate — direct 3'-UTR targeting confirmed by reporter assay, functional rescue with resistant construct, multiple orthogonal methods","pmids":["19801681"],"is_preprint":false},{"year":2011,"finding":"WAVE3/WASF3 knockdown leads to upregulation of the KISS1 metastasis suppressor gene, elevated IκBα levels in the cytoplasm, and reduced nuclear NF-κB (p65/50). Knockdown of KISS1 in WASF3-silenced cells recovers the invasion phenotype. TNF-α treatment has no effect on invasion or NF-κB nuclear translocation in WASF3 knockdown cells, placing WASF3 upstream of NF-κB/KISS1 in the regulation of MMP-9 production.","method":"shRNA knockdown, oligonucleotide arrays, luciferase reporter (KISS1 transcription), Western blotting (IκBα/NF-κB), rescue knockdown of KISS1, TNF-α treatment","journal":"International journal of cancer","confidence":"High","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal methods (transcriptomics, reporter assay, rescue KD, pathway protein analysis), epistasis established by double-KD rescue","pmids":["21544801"],"is_preprint":false},{"year":2011,"finding":"miR-31 directly targets the 3'-UTR of WAVE3 mRNA and inhibits its expression. Loss of miR-31 correlates with increased WAVE3 in invasive breast cancer. Re-expression of miR-31-resistant WAVE3 reverses miR-31-mediated inhibition of cancer cell invasion.","method":"3'-UTR targeting validation, Western blotting, invasion assay, rescue with miR-31-resistant WAVE3","journal":"International journal of cancer","confidence":"High","confidence_rationale":"Tier 2 / Moderate — direct 3'-UTR targeting, functional rescue with resistant construct, correlation in cell lines and human tumors","pmids":["21105030"],"is_preprint":false},{"year":2012,"finding":"HSP90 is present in the WASF3/WAVE3 immunocomplex from prostate cancer cells. Inactivation of HSP90 does not affect WASF3 stability but prevents its phosphoactivation by destabilizing ABL kinase. HSP70 is also present in the WASF3 immunocomplex; inactivation of HSP70 leads to WASF3 destabilization through proteasome degradation. Overexpression of HSP70 in WASF3-null cells does not enhance invasion.","method":"Mass spectrometry, co-immunoprecipitation, pharmacological inhibition of HSP90/70, shRNA knockdown, proteasome inhibition assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1-2 / Moderate — mass spectrometry identification, Co-IP, multiple functional validations (stability vs phospho-activation distinction), negative control (HSP70 OE in null cells)","pmids":["22315230"],"is_preprint":false},{"year":2012,"finding":"HIF1A binds to hypoxia response elements (HRE) in the WASF3 promoter under hypoxic conditions, as shown by ChIP assay, and induces WASF3 transcription. Hypoxia also increases WASF3 phosphoactivation. WASF3 knockdown cells show no motility response to hypoxia.","method":"Chromatin immunoprecipitation (ChIP), luciferase reporter assay, scratch wound motility assay, Western blotting for phospho-WASF3","journal":"International journal of cancer","confidence":"High","confidence_rationale":"Tier 2 / Moderate — ChIP demonstrating direct HIF1A binding, luciferase reporter, functional validation with KD; multiple orthogonal methods in single study","pmids":["22581642"],"is_preprint":false},{"year":2012,"finding":"Loss of WAVE3 in cancer cells leads to inhibition of NFκB signaling through decreased nuclear translocation of NFκB, resulting in loss of activation of NFκB target genes (including MMP9), inhibition of invadopodia formation and ECM degradation, and sensitization of cancer cells to TNFα-induced apoptosis through inhibition of the AKT pro-survival pathway. Conversely, overexpression of WAVE3 enhances NFκB activity.","method":"WAVE3 knockdown/overexpression, NFκB nuclear translocation assay, invadopodia formation assay, ECM degradation assay, MMP9 expression analysis, apoptosis assay","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple functional readouts with both loss- and gain-of-function approaches; single lab","pmids":["25329315"],"is_preprint":false},{"year":2013,"finding":"WASF3/WAVE3 overexpression increases ZEB1/2 levels, which suppress the miR-200a/200b/429 cluster. This occurs through WASF3-mediated downregulation of KISS1, which releases IκBα inhibition of NFκB, and ZEB1 expression is regulated by NFκB. Knockdown of WASF3 leads to reduced ZEB1 levels, increased miR-200 and E-cadherin levels, and loss of invasion.","method":"WASF3 overexpression/knockdown, Western blotting, luciferase reporter, NFκB pathway analysis","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — pathway dissection by gain/loss-of-function with multiple molecular readouts; single lab","pmids":["23318438"],"is_preprint":false},{"year":2013,"finding":"IL-6 induces WASF3 expression and phosphoactivation through the JAK2/STAT3 pathway in two ways: (1) STAT3 directly binds the WASF3 promoter and increases transcription; (2) JAK2 interacts with WASF3 and directly activates (phosphorylates) it. Inhibition of STAT3 (shRNA, dominant negative, or S3I-201) reduces WASF3 levels and migration. Inhibition of JAK2 (shRNA or AG490) reduces WASF3 activation and prevents its membrane localization.","method":"ChIP demonstrating STAT3 binding to WASF3 promoter, Co-IP (JAK2-WASF3), shRNA, dominant-negative constructs, pharmacological inhibition, migration assay, membrane localization assay","journal":"Carcinogenesis","confidence":"High","confidence_rationale":"Tier 2 / Moderate — ChIP, Co-IP, pharmacological and genetic inhibition with functional readouts; multiple orthogonal methods","pmids":["23677069"],"is_preprint":false},{"year":2013,"finding":"TGF-β selectively and robustly induces WAVE3 expression in metastatic breast cancer cells through a Smad2- and β3 integrin-dependent mechanism. WAVE3 is required for TGF-β-induced EMT: stable depletion of WAVE3 prevents TGF-β from inducing EMT programs, lamellipodia formation, and cell migration.","method":"TGF-β treatment with downstream Western blotting, stable WAVE3 knockdown, EMT marker analysis, lamellipodia assay, 3D organotypic culture, in vivo syngeneic mouse model","journal":"Breast cancer research and treatment","confidence":"High","confidence_rationale":"Tier 2 / Strong — pathway placement by genetic KD with multiple readouts (EMT markers, lamellipodia, migration), in vitro and in vivo validation","pmids":["24197660"],"is_preprint":false},{"year":2015,"finding":"ATAD3A (mitochondrial membrane protein) interacts with WASF3/WAVE3 as demonstrated by mass spectrometry. ATAD3A knockdown decreases WASF3 protein levels. HSP70 stabilizes WASF3 in the cytoplasm, but inactivation of HSP70 does not destabilize WASF3 at the mitochondrial membrane where ATAD3A protects it. GRP78 upregulation (during ER stress) increases WASF3 levels, and ATAD3A is present in a WASF3-GRP78 complex. Suppression of GRP78 leads to ATAD3A-dependent destabilization of WASF3 at the mitochondrial membrane. The N-terminal end of WASF3 is within the mitochondria and is the interaction site with the N-terminal end of ATAD3A.","method":"Mass spectrometry, co-immunoprecipitation, shRNA knockdown, mitochondrial fractionation with proteolysis protection assay, in vivo xenograft","journal":"Oncogene","confidence":"High","confidence_rationale":"Tier 1-2 / Moderate — mass spectrometry interaction, mitochondrial proteolysis domain mapping, multiple KD validations, Co-IP","pmids":["25823022"],"is_preprint":false},{"year":2015,"finding":"Genetic knockdown of CYFIP1 in cancer cells destabilizes the WASF3 complex, causes loss of WASF3 function, and suppresses invasion. Stapled peptides (WAHM) targeting the α-helical interface between WASF3 and CYFIP1 suppress motility and invasion in breast and prostate cancer cells, suppress Rac interaction with the WASF3 complex, and dysregulate downstream targets MMP-9 and KISS1. Depletion of WASF1 and WASF2, which also bind CYFIP1, did not affect invasion, demonstrating specificity for WASF3.","method":"shRNA knockdown of CYFIP1, stapled peptide treatment, invasion/motility assay, Rac interaction assay (Co-IP), MMP-9 and KISS1 expression analysis","journal":"Cancer research","confidence":"High","confidence_rationale":"Tier 2 / Moderate — genetic and pharmacologic disruption of defined protein-protein interface with multiple functional readouts including RAC binding; paralog specificity controls","pmids":["26676744"],"is_preprint":false},{"year":2016,"finding":"WASF3 is present in the HER2 immunocomplex. Suppression of WASF3 suppresses invasion even in the presence of HER2 expression. WASF3's ability to promote invasion is highly dependent on the HER2/HER3 heterodimer. The HER2/HER3 complex facilitates WASF3 phospho-activation and transcriptional upregulation through HER2/HER3 activation of JAK/STAT signaling.","method":"Co-immunoprecipitation, WASF3 suppression/overexpression, invasion assays with HER2/HER3 manipulation, Western blotting (phospho-WASF3)","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP establishing complex, functional epistasis, JAK/STAT pathway linking; single lab","pmids":["26804171"],"is_preprint":false},{"year":2016,"finding":"NCKAP1 is required for WASF3 complex stability and function: silencing NCKAP1 destabilizes the WASF3 complex and suppresses invasive capacity of breast, prostate, and colon cancer cells and metastasis in vivo. Activation of the WASF3 complex requires RAC1 interaction, and inactivation of NCKAP1 prevents RAC1 association with the WASF3 complex. Stapled peptides (WANT3) targeting the NCKAP1-CYFIP1 interface destabilize the WASF3 complex and suppress RAC1 binding and invasion.","method":"shRNA knockdown of NCKAP1, Co-IP (RAC1-WASF3 complex), in vivo spontaneous metastasis model, stapled peptide treatment","journal":"Cancer research","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic KD with in vitro and in vivo validation, RAC1 binding mechanistic data by Co-IP, pharmacologic disruption; replicated across multiple cancer cell types","pmids":["27432794"],"is_preprint":false},{"year":2017,"finding":"WAVE3 interacts with YB1 (Y-box binding protein 1), and this interaction is required for YB1 nuclear translocation in cancer cells and activation of transcription of cancer stem cell-specific genes. WAVE3 is enriched in the CSC subpopulation. CRISPR/Cas9 knockout of WAVE3 attenuates the CSC subpopulation and inhibits transcription of CSC transcription factors.","method":"CRISPR/Cas9 knockout, Co-immunoprecipitation (WAVE3-YB1 interaction), nuclear/cytoplasmic fractionation, CSC marker analysis, transcription factor activity assays","journal":"Oncotarget","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — CRISPR KO with functional readout, Co-IP for interaction, localization assay; single lab","pmids":["29262622"],"is_preprint":false},{"year":2017,"finding":"p63α transcriptionally upregulates HSP70 (Hsp70) expression via E2F1, and HSP70 promotes bladder cancer cell invasion through the Hsp70/WASF3/MMP-9 axis.","method":"Western blotting, invasion assay, transcription analysis (E2F1/p63α-mediated Hsp70 upregulation), pathway inhibition","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — mechanistic pathway placement using molecular biology in a single lab study; consistent with prior HSP70-WASF3 interaction data","pmids":["28794159"],"is_preprint":false},{"year":2017,"finding":"Mutant RAS promotion of invasion and metastasis is dependent on WASF3 activation in a PI3K and AKT-dependent manner. AKT is present in the WASF3 immunocomplex and this association is enhanced by mutant RAS overexpression. Mutant RAS promotes dissociation of p85 from the WASF3 complex, promoting activation of p110. ERK1/2 activation is not affected by loss of WASF3.","method":"Co-immunoprecipitation (AKT in WASF3 complex), proteomics analysis, WASF3 knockdown epistasis, p85/p110 dissociation assay","journal":"Genes, chromosomes & cancer","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP with proteomics, genetic epistasis; single lab, moderate mechanistic depth","pmids":["28233357"],"is_preprint":false},{"year":2019,"finding":"Wasf3 null mice (generated by deletion of exons 4 and 5) are viable with no visible morphological or behavioral abnormalities and no abnormal mammary gland development or brain development. In the MMTV-polyoma middle-T oncogene breast cancer model, Wasf3 is upregulated in metastatic lesions, and Wasf3 null background reduces the number and size of metastatic lung lesions without affecting primary tumor development.","method":"Conditional knockout mouse model (Cre-lox deletion), MMTV-PyMT spontaneous breast cancer model, histological analysis of metastases","journal":"The American journal of pathology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — in vivo genetic KO with spontaneous cancer model, quantitative metastasis assessment; rigorous mammalian genetics","pmids":["31542393"],"is_preprint":false},{"year":2020,"finding":"PCARE (C2orf71) interacts with WASF3/WAVE3 and efficiently recruits it to the primary cilium. Ectopic coexpression of PCARE and WASF3 in ciliated cells causes remarkable expansion of the ciliary tip via actin polymerization. This expansion is disrupted by siRNA knockdown of actin regulators, pharmacological inhibition of actin polymerization, or PCARE with a retinal dystrophy-associated missense mutation. In mouse retina and human retinal organoids, PCARE and WASF3 colocalize with actin at the outer segment base where this process drives disk formation initiation.","method":"Co-immunoprecipitation (PCARE-WASF3), ectopic co-expression in ciliated cells, siRNA knockdown, pharmacological actin inhibition, mouse retinal imaging, human retinal organoids","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal interaction, functional co-expression system, multiple disruption methods, in vivo model validation, disease mutation support","pmids":["32312818"],"is_preprint":false},{"year":2020,"finding":"WAVE3 tyrosine phosphorylation (downstream of PI3K) is also achieved downstream of TGF-β and EGF signaling, and is required for oncogenic activity including migration, tumorsphere growth, and invasion. Loss of WAVE3 phosphorylation also inhibits the activation of PI3K, TGF-β, and EGF signaling downstream effectors, identifying a positive feedback loop between WAVE3 phosphorylation and these pathways.","method":"Phospho-mutant WAVE3 constructs, Western blotting for pathway effectors, migration assay, 3D tumorsphere assay, mouse xenograft model","journal":"Oncogenesis","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — phospho-mutant approach, multiple pathway readouts, in vitro and in vivo; single lab","pmids":["33012785"],"is_preprint":false},{"year":2021,"finding":"Phosphorylation of the proline-rich domain (PRD) of WAVE3 is essential for its interaction with YB1. Loss of PRD phosphorylation inhibits WAVE3-YB1 interaction, prevents YB1-mediated activation of CSC markers, and inhibits WAVE3-mediated EMT activation. PRD phosphorylation is required for migration and invasion in vitro and tumor growth/metastasis in vivo.","method":"Phospho-mutant PRD constructs, co-immunoprecipitation (WAVE3-YB1), invasion/migration assays, CSC marker analysis, in vivo xenograft","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — phospho-mutant with defined domain, Co-IP demonstrating interaction requirement; single lab","pmids":["33594155"],"is_preprint":false},{"year":2021,"finding":"SHOX2 directly activates WASF3 transcription and recruits STAT3 to the WASF3 promoter, where SHOX2 and STAT3 form a functional immunocomplex that cooperatively promotes WASF3 transcriptional activity. WASF3 knockdown abrogates SHOX2-induced metastasis but not SHOX2-dependent tumorigenesis.","method":"ChIP-qPCR and ChIP/re-ChIP (SHOX2 and STAT3 at WASF3 promoter), co-immunoprecipitation (SHOX2-STAT3 complex), shRNA knockdown epistasis, in vivo orthotopic mouse model","journal":"Journal of experimental & clinical cancer research","confidence":"High","confidence_rationale":"Tier 2 / Moderate — ChIP/re-ChIP with Co-IP establishing transcriptional complex, genetic epistasis in vivo; multiple orthogonal methods","pmids":["34465361"],"is_preprint":false},{"year":2023,"finding":"Overexpression of WASF3 disrupts mitochondrial respiratory supercomplex formation and is associated with ER stress. In transgenic mice with increased WASF3 expression, treadmill running capacity was markedly decreased with concomitantly impaired respiratory supercomplex assembly and reduced complex IV levels in skeletal muscle mitochondria. ER stress-induced WASF3 upregulation (by endotoxin) also decreased skeletal muscle complex IV levels. Pharmacologic inhibition of ER stress decreased WASF3 levels and improved mitochondrial function in patient cells.","method":"WASF3 transgenic mouse model, treadmill exercise testing, mitochondrial supercomplex assembly analysis (BN-PAGE or equivalent), complex IV activity measurement, pharmacological ER stress inhibition, patient skeletal muscle biopsy analysis","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — transgenic mouse model with quantitative functional phenotype, biochemical supercomplex analysis, pharmacologic rescue, patient tissue validation; multiple orthogonal methods","pmids":["37579159"],"is_preprint":false},{"year":2023,"finding":"WAVE3/WASF3 phosphorylation is required for β-catenin stabilization; loss of WAVE3 expression or phosphorylation inhibits β-catenin activity and expression. Dual blocking of WAVE3 expression or phosphorylation in combination with chemotherapy suppresses chemoresistant TNBC cell behavior in vitro and in vivo. Re-expression of phospho-active WAVE3 restores oncogenic activity, while phospho-mutant WAVE3 does not.","method":"CRISPR/Cas9 KO, phospho-mutant rescue, Western blotting (β-catenin), 2D/3D invasion assay, xenograft assay","journal":"Breast cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — CRISPR KO and phospho-mutant rescue with β-catenin mechanistic link; single lab","pmids":["36949468"],"is_preprint":false},{"year":2025,"finding":"METTL3 mediates m6A modification of WASF3 mRNA. IGF2BP2 binds to the m6A site in the 3'-UTR of WASF3 mRNA and enhances WASF3 translation. Highly expressed WASF3 activates the MAPK signaling pathway by interacting with phosphorylated p38 (p-p38). Removal of m6A modification of WASF3 mRNA inhibited WASF3 expression and abolished WASF3's ability to bind p-p38 and activate MAPK signaling.","method":"Co-immunoprecipitation (METTL3-WASF3, WASF3-p-p38), m6A modification assay, Western blotting, m6A inhibition experiments, ESCC cell functional assays","journal":"MedComm","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP for WASF3-p-p38 interaction, m6A modification/translation mechanism; single lab, single study","pmids":["41127505"],"is_preprint":false}],"current_model":"WASF3/WAVE3 is a WASP-family actin cytoskeleton remodeling protein that functions as a key node in cancer invasion and metastasis: it localizes to lamellipodia downstream of PI3K (via interaction with the p85 subunit) and is phosphoactivated on four tyrosine residues by c-Abl kinase (facilitated by HER2/HER3 via JAK/STAT signaling); it is stabilized by HSP70 in the cytoplasm and by ATAD3A/GRP78 at the mitochondrial membrane; it is transcriptionally induced by HIF1A, STAT3 (bound directly to its promoter downstream of JAK2/IL-6), and the SHOX2-STAT3 complex; its activity promotes invasion through NFκB-MMP9 signaling, KISS1 suppression, and ZEB1-mediated miR-200 inactivation; it regulates cancer stem cell maintenance through its PRD-phosphorylation-dependent interaction with YB1 nuclear translocation; it forms a pentameric WASF Regulatory Complex with NCKAP1, CYFIP1, ABI, and BRK1 that requires RAC1 interaction for activation; in photoreceptors, WASF3 is recruited by PCARE to the ciliary tip to drive actin-dependent outer segment disk formation; and, in a non-cancer context, WASF3 overexpression disrupts mitochondrial respiratory supercomplex assembly (reducing complex IV levels), potentially mediating exercise intolerance in ME/CFS."},"narrative":{"mechanistic_narrative":"WASF3 (WAVE3) is a WASP-family actin-cytoskeleton regulator that drives cell motility and serves as a central node for cancer invasion and metastasis [PMID:15826941, PMID:17525277]. It localizes to lamellipodia at the leading edge of migrating cells downstream of PI3K signaling, where its N-terminal region binds the p85 regulatory subunit; loss of WASF3 abolishes PDGF-induced lamellipodia and migration [PMID:15826941]. WASF3 functions within a CYFIP1/NCKAP1-containing regulatory complex whose stability and RAC1-dependent activation require both subunits, and disruption of the WASF3-CYFIP1 or NCKAP1-CYFIP1 interfaces with stapled peptides collapses the complex, blocks RAC1 binding, and suppresses invasion in breast, prostate, and colon cancer cells with paralog specificity over WASF1/WASF2 [PMID:26676744, PMID:27432794]. Its activity is gated by tyrosine phosphorylation: c-Abl phosphorylates four tyrosine residues downstream of PDGF, and equivalent phosphoactivation is achieved downstream of TGF-β, EGF, and HER2/HER3 via JAK2/STAT3 signaling, establishing a positive feedback loop with these pathways [PMID:17623672, PMID:23677069, PMID:26804171, PMID:33012785]. Activated WASF3 promotes invasion through an NFκB–MMP9 axis, suppression of the KISS1 metastasis-suppressor and consequent ZEB1-driven inactivation of the miR-200 family during EMT [PMID:21544801, PMID:25329315, PMID:23318438], and through proline-rich-domain-phosphorylation-dependent binding to YB1 that drives YB1 nuclear translocation and cancer stem-cell gene transcription [PMID:29262622, PMID:33594155]. WASF3 abundance is controlled at multiple levels: transcriptionally by HIF1A under hypoxia, by STAT3 and the SHOX2-STAT3 complex at its promoter, and post-transcriptionally by miR-200 and miR-31 targeting its 3'-UTR; its protein is stabilized by HSP70 in the cytoplasm and by ATAD3A/GRP78 at the mitochondrial membrane, where its N-terminus engages the ATAD3A N-terminus [PMID:19801681, PMID:21105030, PMID:22581642, PMID:25823022, PMID:34465361]. Genetic ablation in mice confirms WASF3 is dispensable for development but required for metastatic dissemination in the MMTV-PyMT model [PMID:31542393]. Beyond cancer, WASF3 is recruited by PCARE to the photoreceptor ciliary tip to drive actin-dependent outer-segment disk formation [PMID:32312818], and its overexpression disrupts mitochondrial respiratory supercomplex assembly with reduced complex IV, linking ER-stress-induced WASF3 to exercise intolerance [PMID:37579159].","teleology":[{"year":2005,"claim":"Established WASF3 as an actin-remodeling effector acting downstream of PI3K to build lamellipodia, defining its core cellular role in migration.","evidence":"Microscopy localization, RNAi knockdown of PDGF-induced lamellipodia, yeast two-hybrid and Co-IP mapping the WAVE3 N-terminus to p85 SH2 domain, and LY294002 inhibition","pmids":["15826941"],"confidence":"High","gaps":["Did not establish how p85 binding mechanistically couples to Arp2/3 activation","Direct actin-nucleation activity of WASF3 not measured"]},{"year":2005,"claim":"Connected WASF3 to invasion machinery by showing it sustains p38 MAPK signaling and MMP expression, distinguishing it functionally from WAVE1/WAVE2.","evidence":"siRNA knockdown with Western blots of phospho-kinases, MMP expression, and actin/focal-adhesion staining","pmids":["15907837"],"confidence":"Medium","gaps":["Whether WASF3 directly regulates p38 or acts indirectly was not resolved","MMP regulation mechanism left undefined"]},{"year":2005,"claim":"Placed WASF3 within the same regulatory complexes as WAVE1/WAVE2 and identified LDOC1 as a VH-domain partner whose nuclear export WASF3 drives to suppress apoptosis.","evidence":"Co-IP interaction assays across isoforms; direct binding, fractionation, and apoptosis assays for LDOC1","pmids":["15752430","16272576"],"confidence":"Medium","gaps":["LDOC1 regulation not tested in vivo","Complex composition not yet resolved as a defined pentamer"]},{"year":2007,"claim":"Defined the activation switch by showing c-Abl phosphorylates four WASF3 tyrosines as a requirement for motility, converting WASF3 into a kinase-gated effector.","evidence":"Co-IP, in vitro kinase assay, tyrosine site-directed mutagenesis, STI-571 inhibition, migration assays","pmids":["17623672"],"confidence":"High","gaps":["Structural consequence of tyrosine phosphorylation on complex conformation unknown","Whether all four sites are equivalent not dissected"]},{"year":2007,"claim":"Demonstrated WASF3 is required for metastasis in vivo and operates through a p38-MAPK pathway, validating it as a metastasis driver beyond cell culture.","evidence":"shRNA stable knockdown, Matrigel invasion, lung colonization and orthotopic xenografts, dominant-negative p38 epistasis","pmids":["17525277"],"confidence":"High","gaps":["How WASF3 feeds into p38 activation not mechanistically pinned","Primary tumor versus metastasis-specific effects not fully separated"]},{"year":2009,"claim":"Identified post-transcriptional control of WASF3 by miR-200 (and later miR-31) targeting its 3'-UTR, tying WASF3 levels to EMT/MET state.","evidence":"Luciferase 3'-UTR reporters, Western blots, invasion assays, rescue with miRNA-resistant WASF3 constructs","pmids":["19801681","21105030"],"confidence":"High","gaps":["Whether endogenous miRNA changes fully account for tumor WASF3 levels not quantified","Interplay between the two miRNAs not addressed"]},{"year":2011,"claim":"Mapped the invasion output pathway, placing WASF3 upstream of NFκB to repress the KISS1 metastasis suppressor and drive MMP-9 production.","evidence":"shRNA knockdown, oligonucleotide arrays, KISS1 luciferase reporter, IκBα/NFκB Western blots, KISS1 rescue knockdown, TNF-α treatment","pmids":["21544801"],"confidence":"High","gaps":["Direct molecular link between cytoplasmic WASF3 and NFκB activation not identified","Whether KISS1 repression is transcriptional or indirect not fully defined"]},{"year":2012,"claim":"Extended the invasion axis to invadopodia and survival, and established hypoxic transcriptional induction of WASF3 by HIF1A binding to its promoter.","evidence":"Knockdown/overexpression with invadopodia and ECM degradation assays, TNFα apoptosis assays; ChIP, luciferase reporter and motility assays for HIF1A","pmids":["25329315","22581642"],"confidence":"Medium","gaps":["HIF1A and other inputs not integrated quantitatively","AKT pro-survival linkage mechanism not detailed"]},{"year":2012,"claim":"Resolved chaperone-dependent stability and activation, separating HSP70-dependent WASF3 protein stability from HSP90-dependent ABL stability needed for phosphoactivation.","evidence":"Mass spectrometry, Co-IP, HSP90/70 pharmacological inhibition, shRNA, proteasome inhibition, and HSP70-overexpression negative control in null cells","pmids":["22315230"],"confidence":"High","gaps":["E3 ligase mediating WASF3 proteasomal turnover not identified","Structural basis of HSP70 binding unknown"]},{"year":2013,"claim":"Built the EMT regulatory loop (KISS1→NFκB→ZEB1→miR-200) and established cytokine-driven dual control of WASF3 by JAK2/STAT3 transcription and JAK2 phosphorylation.","evidence":"Gain/loss-of-function Westerns and reporters for ZEB1/miR-200; ChIP, JAK2-WASF3 Co-IP, dominant-negative and pharmacological inhibition with migration/membrane-localization assays","pmids":["23318438","23677069"],"confidence":"High","gaps":["Feedback stability of the ZEB1/miR-200 loop not modeled","Relative contribution of transcriptional versus phospho-activation by JAK2 not weighted"]},{"year":2013,"claim":"Positioned WASF3 as an obligatory effector of TGF-β-induced EMT via Smad2 and β3-integrin, broadening its upstream signaling repertoire.","evidence":"TGF-β treatment with Westerns, stable knockdown, EMT marker analysis, lamellipodia assay, 3D organotypic culture, syngeneic mouse model","pmids":["24197660"],"confidence":"High","gaps":["Direct Smad2 target status of WASF3 promoter not shown","Integrin requirement mechanism not resolved"]},{"year":2015,"claim":"Defined a mitochondrial pool of WASF3 stabilized by ATAD3A and GRP78, distinct from the cytoplasmic HSP70-protected pool, and mapped the N-terminal interaction interface.","evidence":"Mass spectrometry, Co-IP, shRNA, mitochondrial fractionation with proteolysis protection mapping, in vivo xenograft","pmids":["25823022"],"confidence":"High","gaps":["Functional role of mitochondrial WASF3 in invasion versus respiration not delineated at this stage","Whether mitochondrial WASF3 is phosphoactivated unknown"]},{"year":2015,"claim":"Established the WASF3 regulatory complex as a druggable, RAC1-dependent assembly by showing CYFIP1 and NCKAP1 are required for stability and RAC1 binding, with paralog-specific peptide inhibition.","evidence":"shRNA knockdown of CYFIP1/NCKAP1, RAC1-complex Co-IP, WAHM/WANT3 stapled peptides, invasion/motility assays, in vivo metastasis model, WASF1/WASF2 specificity controls","pmids":["26676744","27432794"],"confidence":"High","gaps":["High-resolution structure of the WASF3 complex not determined","Why WASF3 is uniquely required over paralogs not explained molecularly"]},{"year":2016,"claim":"Linked receptor tyrosine kinase oncogenic signaling to WASF3 by showing HER2/HER3 drives both its phosphoactivation and transcription through JAK/STAT, making WASF3 a HER2-pathway effector.","evidence":"Co-IP placing WASF3 in the HER2 complex, suppression/overexpression with HER2/HER3 manipulation, phospho-WASF3 Westerns and invasion epistasis","pmids":["26804171"],"confidence":"Medium","gaps":["Direct versus indirect HER2-WASF3 association not distinguished","Single-lab finding without reciprocal in vivo validation"]},{"year":2017,"claim":"Uncovered a nuclear-transcriptional arm of WASF3 function: PRD-phosphorylation-dependent binding to YB1 drives YB1 nuclear import and cancer stem-cell gene programs.","evidence":"CRISPR/Cas9 knockout, WAVE3-YB1 Co-IP, nuclear/cytoplasmic fractionation, CSC marker and transcription factor activity assays; phospho-mutant PRD constructs","pmids":["29262622","33594155"],"confidence":"Medium","gaps":["Kinase phosphorylating the PRD not identified","How a cytoskeletal protein localizes to drive nuclear YB1 import mechanistically unclear"]},{"year":2017,"claim":"Integrated WASF3 into RAS-PI3K-AKT oncogenic signaling, showing mutant RAS activates WASF3 by dissociating p85 to free p110, with AKT in the WASF3 complex.","evidence":"AKT-WASF3 Co-IP, proteomics, knockdown epistasis, p85/p110 dissociation assays","pmids":["28233357"],"confidence":"Medium","gaps":["Direct versus scaffolded AKT association not resolved","Single-lab finding"]},{"year":2017,"claim":"Connected upstream transcriptional control of the chaperone arm by showing p63α/E2F1-driven HSP70 promotes invasion through the HSP70/WASF3/MMP-9 axis.","evidence":"Westerns, invasion assays, transcriptional analysis of p63α/E2F1-mediated HSP70 induction, pathway inhibition","pmids":["28794159"],"confidence":"Medium","gaps":["Direct effect on WASF3 stability versus indirect not separated here","Cancer-type generality not tested"]},{"year":2019,"claim":"Provided definitive germline genetics: WASF3 is dispensable for normal development but specifically required for metastatic spread in a spontaneous tumor model.","evidence":"Wasf3-null mouse (exon 4/5 deletion), MMTV-PyMT spontaneous breast cancer model, histological metastasis quantification","pmids":["31542393"],"confidence":"High","gaps":["Cell-autonomous versus microenvironmental contribution not dissected","Compensation by WASF1/WASF2 during development not assessed"]},{"year":2020,"claim":"Generalized WASF3 phosphorylation as a feed-forward hub, required for and reinforcing PI3K, TGF-β, and EGF signaling outputs.","evidence":"Phospho-mutant constructs, pathway-effector Westerns, migration, 3D tumorsphere assays, mouse xenografts","pmids":["33012785"],"confidence":"Medium","gaps":["Molecular basis of feedback onto upstream receptors unknown","Identity of all kinases acting on each site unresolved"]},{"year":2020,"claim":"Revealed a non-cancer ciliary role: PCARE recruits WASF3 to the photoreceptor ciliary tip to drive actin-dependent outer-segment disk formation.","evidence":"PCARE-WASF3 Co-IP, ectopic co-expression in ciliated cells, siRNA and pharmacological actin disruption, retinal-dystrophy mutation, mouse retina and human retinal organoids","pmids":["32312818"],"confidence":"High","gaps":["Whether ciliary WASF3 uses the same RAC1-dependent complex as in cancer not tested","Endogenous WASF3 requirement in vivo not genetically confirmed in retina"]},{"year":2021,"claim":"Identified a cooperative transcriptional driver, the SHOX2-STAT3 promoter complex, that selectively governs WASF3-dependent metastasis rather than tumorigenesis.","evidence":"ChIP/re-ChIP of SHOX2 and STAT3 at the WASF3 promoter, SHOX2-STAT3 Co-IP, shRNA epistasis, orthotopic mouse model","pmids":["34465361"],"confidence":"High","gaps":["How SHOX2-STAT3 selectively affects metastatic but not tumorigenic programs unclear","Other promoter co-factors not enumerated"]},{"year":2023,"claim":"Linked WASF3 phosphorylation to β-catenin stabilization and chemoresistance, identifying a co-targeting strategy in triple-negative breast cancer.","evidence":"CRISPR/Cas9 KO, phospho-mutant rescue, β-catenin Westerns, 2D/3D invasion, xenografts","pmids":["36949468"],"confidence":"Medium","gaps":["Direct versus indirect β-catenin regulation not resolved","Single-lab finding"]},{"year":2023,"claim":"Established a distinct pathological role outside cancer: WASF3 overexpression impairs mitochondrial respiratory supercomplex assembly and reduces complex IV, modeling exercise intolerance in ME/CFS.","evidence":"WASF3 transgenic mice, treadmill testing, supercomplex assembly and complex IV analysis, ER-stress induction and pharmacological inhibition, patient muscle biopsies","pmids":["37579159"],"confidence":"High","gaps":["Molecular mechanism by which WASF3 disrupts supercomplex assembly not defined","Connection to its actin/invasion functions unestablished"]},{"year":2025,"claim":"Added m6A-based translational control, with METTL3/IGF2BP2 enhancing WASF3 translation to enable p-p38 binding and MAPK activation.","evidence":"METTL3-WASF3 and WASF3-p-p38 Co-IP, m6A modification assays, m6A inhibition, ESCC functional assays","pmids":["41127505"],"confidence":"Medium","gaps":["Direct WASF3-p-p38 interaction surface not mapped","Single-lab, single cancer-type finding"]},{"year":null,"claim":"How the cytoplasmic actin-regulatory WASF3 complex is mechanistically reconciled with its nuclear (YB1), mitochondrial (ATAD3A/supercomplex), and ciliary (PCARE) functions, and the structural basis of its phospho-activation, remain unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No high-resolution structure of the activated WASF3 complex","Kinases for the PRD and the mechanism of supercomplex disruption unknown","Whether the same RAC1-dependent complex operates across all compartments untested"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[0,16,18,23]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[3,19,25]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[7,11,12]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[0,13]},{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[0,1,23]},{"term_id":"GO:0005739","term_label":"mitochondrion","supporting_discovery_ids":[15,27]},{"term_id":"GO:0005929","term_label":"cilium","supporting_discovery_ids":[23]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[19,25]}],"pathway":[{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[0,4,13,21]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[5,22,28]},{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[10,13,26]},{"term_id":"R-HSA-1266738","term_label":"Developmental Biology","supporting_discovery_ids":[14,23]},{"term_id":"R-HSA-1430728","term_label":"Metabolism","supporting_discovery_ids":[27]}],"complexes":["WASF Regulatory Complex (WASF3-CYFIP1-NCKAP1-ABI-BRK1)"],"partners":["CYFIP1","NCKAP1","RAC1","YB1","ATAD3A","PCARE","JAK2","ABL1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9UPY6","full_name":"Actin-binding protein WASF3","aliases":["Protein WAVE-3","Verprolin homology domain-containing protein 3","Wiskott-Aldrich syndrome protein family member 3","WASP family protein member 3"],"length_aa":502,"mass_kda":55.3,"function":"Downstream effector molecules involved in the transmission of signals from tyrosine kinase receptors and small GTPases to the actin cytoskeleton. Plays a role in the regulation of cell morphology and cytoskeletal organization. Required in the control of cell shape","subcellular_location":"Cytoplasm, cytoskeleton","url":"https://www.uniprot.org/uniprotkb/Q9UPY6/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/WASF3","classification":"Not Classified","n_dependent_lines":3,"n_total_lines":1208,"dependency_fraction":0.0024834437086092716},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"NCKAP1","stoichiometry":10.0},{"gene":"WASF2","stoichiometry":4.0},{"gene":"ACTG1","stoichiometry":0.2},{"gene":"CAPZB","stoichiometry":0.2},{"gene":"WASF1","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/WASF3","total_profiled":1310},"omim":[{"mim_id":"605875","title":"WASP PROTEIN FAMILY, MEMBER 2; WASF2","url":"https://www.omim.org/entry/605875"},{"mim_id":"605068","title":"WASP PROTEIN FAMILY, MEMBER 3; WASF3","url":"https://www.omim.org/entry/605068"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in many","driving_tissues":[{"tissue":"brain","ntpm":58.7},{"tissue":"retina","ntpm":52.2}],"url":"https://www.proteinatlas.org/search/WASF3"},"hgnc":{"alias_symbol":["WAVE3","SCAR3","KIAA0900"],"prev_symbol":[]},"alphafold":{"accession":"Q9UPY6","domains":[{"cath_id":"1.20.5.340","chopping":"9-86","consensus_level":"medium","plddt":93.8173,"start":9,"end":86},{"cath_id":"1.20.5","chopping":"134-188","consensus_level":"medium","plddt":91.4129,"start":134,"end":188}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9UPY6","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9UPY6-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9UPY6-F1-predicted_aligned_error_v6.png","plddt_mean":65.56},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=WASF3","jax_strain_url":"https://www.jax.org/strain/search?query=WASF3"},"sequence":{"accession":"Q9UPY6","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9UPY6.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9UPY6/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9UPY6"}},"corpus_meta":[{"pmid":"17525277","id":"PMC_17525277","title":"Down-regulation 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sciences","url":"https://pubmed.ncbi.nlm.nih.gov/32359534","citation_count":12,"is_preprint":false},{"pmid":"26052252","id":"PMC_26052252","title":"Knockdown of WAVE3 impairs HGF induced migration and invasion of prostate cancer cells.","date":"2015","source":"Cancer cell international","url":"https://pubmed.ncbi.nlm.nih.gov/26052252","citation_count":10,"is_preprint":false},{"pmid":"32793744","id":"PMC_32793744","title":"Elucidating the molecular signaling pathways of WAVE3.","date":"2020","source":"Annals of translational medicine","url":"https://pubmed.ncbi.nlm.nih.gov/32793744","citation_count":9,"is_preprint":false},{"pmid":"35450347","id":"PMC_35450347","title":"In Silico Optimized Stapled Peptides Targeting WASF3 in Breast Cancer.","date":"2022","source":"ACS medicinal chemistry letters","url":"https://pubmed.ncbi.nlm.nih.gov/35450347","citation_count":7,"is_preprint":false},{"pmid":"38125536","id":"PMC_38125536","title":"WASF3 overexpression affects the expression of circular RNA hsa-circ-0100153, which promotes breast cancer progression by sponging hsa-miR-31, hsa-miR-767-3p, and hsa-miR-935.","date":"2023","source":"Heliyon","url":"https://pubmed.ncbi.nlm.nih.gov/38125536","citation_count":6,"is_preprint":false},{"pmid":"25510861","id":"PMC_25510861","title":"MiR-200b modulates the properties of human monocyte-derived dendritic cells by targeting WASF3.","date":"2014","source":"Life sciences","url":"https://pubmed.ncbi.nlm.nih.gov/25510861","citation_count":6,"is_preprint":false},{"pmid":"28233357","id":"PMC_28233357","title":"Promotion of invasion by mutant RAS is dependent on activation of the WASF3 metastasis promoter gene.","date":"2017","source":"Genes, chromosomes & cancer","url":"https://pubmed.ncbi.nlm.nih.gov/28233357","citation_count":5,"is_preprint":false},{"pmid":"33854560","id":"PMC_33854560","title":"Fuzheng Yiliu Formula Regulates Tumor Invasion and Metastasis through Inhibition of WAVE3 Expression.","date":"2021","source":"Evidence-based 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Oncology Societies and of the National Cancer Institute of Mexico","url":"https://pubmed.ncbi.nlm.nih.gov/39080187","citation_count":1,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.02.13.25322223","title":"Integrative Genome-Wide Association Studies of COVID-19 Susceptibility and Hospitalization Reveal Risk Loci for Long COVID","date":"2025-02-14","source":"bioRxiv","url":"https://doi.org/10.1101/2025.02.13.25322223","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":32356,"output_tokens":8500,"usd":0.112284,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":18335,"output_tokens":7653,"usd":0.1415,"stage2_stop_reason":"end_turn"},"total_usd":0.253784,"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\": 2005,\n      \"finding\": \"WAVE3/WASF3 localizes to lamellipodia at the leading edge of migrating cells, and its knockdown prevents PDGF-induced lamellipodia formation and cell migration. WAVE3 interacts physically with the PI3K regulatory subunit p85 (mediated by the N-terminal region of WAVE3 and the C-terminal SH2 domain of p85), placing WAVE3 downstream of PI3K signaling. Treatment with PI3K inhibitor LY294002 also abrogated PDGF-induced lamellipodia formation.\",\n      \"method\": \"Subcellular localization by microscopy, RNA interference knockdown, yeast two-hybrid screen confirmed by co-immunoprecipitation, pharmacological inhibition\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal interaction confirmed by yeast two-hybrid and Co-IP with domain mapping, localization by microscopy, functional KD with defined phenotype\",\n      \"pmids\": [\"15826941\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"WAVE3/WASF3 knockdown decreases phospho-p38 MAPK levels (but not phospho-AKT, phospho-ERK, or phospho-JNK) and inhibits expression of MMP-1, MMP-3, and MMP-9 (but not MMP-2), leading to inhibition of cell motility and invasion with increased actin stress fiber formation and reorganization of focal adhesion complexes. WAVE1 and WAVE2 expression levels were not affected by loss of WAVE3.\",\n      \"method\": \"RNA interference knockdown, Western blotting, invasion assay, actin/focal adhesion staining\",\n      \"journal\": \"Experimental cell research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — siRNA KD with multiple downstream readouts (kinase phosphorylation, MMP expression, cell morphology), single lab\",\n      \"pmids\": [\"15907837\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"WAVE3/WASF3 includes the Scar/WAVE family proteins in the same kinds of protein complexes as WAVE1 and WAVE2, all three isoforms interacting with previously described binding partners, suggesting participation in the same regulatory complexes for actin assembly.\",\n      \"method\": \"Co-immunoprecipitation, interaction assays with multiple binding partners across isoforms\",\n      \"journal\": \"BMC cell biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Co-IP-based interaction data replicated across three isoforms in single study\",\n      \"pmids\": [\"15752430\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"WAVE3 binds directly to LDOC1 through the verprolin homology (VH) domain of WAVE3. WAVE3 expression induces translocation of LDOC1 from the nucleus to the cytoplasm, inhibiting LDOC1-induced apoptosis (which requires nuclear LDOC1 and p53 stabilization). Thus WAVE3 negatively regulates LDOC1 function.\",\n      \"method\": \"Direct binding assay, ectopic expression, subcellular localization (nuclear/cytoplasmic fractionation and microscopy), apoptosis assay\",\n      \"journal\": \"Journal of biochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct binding established with domain mapping (VH domain), functional consequence (apoptosis suppression) demonstrated by localization change; single lab\",\n      \"pmids\": [\"16272576\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"c-Abl tyrosine kinase interacts with WAVE3/WASF3 upon PDGF stimulation, and phosphorylates four tyrosine residues on WAVE3. Abl-mediated phosphorylation of WAVE3 is required for stimulation of lamellipodia formation and cell migration. The Abl inhibitor STI-571 abrogates Abl-mediated phosphorylation of WAVE3.\",\n      \"method\": \"Co-immunoprecipitation, in vitro kinase assay, site-directed mutagenesis of tyrosine residues, pharmacological inhibition (STI-571), lamellipodia/migration assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro kinase assay with mutagenesis of phosphorylation sites, co-IP, pharmacological inhibition, and functional migration readout in one study\",\n      \"pmids\": [\"17623672\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Stable shRNA-mediated knockdown of WAVE3 in MDA-MB-231 breast cancer cells reduces Matrigel invasion, lung colony formation after tail-vein injection, and primary tumor growth in orthotopic xenograft models. Suppression of p38 MAPK activity by dominant-negative p38 produces comparable phenotypes, establishing the WAVE3-p38 pathway in metastasis.\",\n      \"method\": \"shRNA stable knockdown, Matrigel invasion assay, xenograft/orthotopic mouse models, dominant-negative p38 construct\",\n      \"journal\": \"The American journal of pathology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic KD with defined phenotype confirmed in vitro and in vivo, pathway placement by dominant-negative epistasis\",\n      \"pmids\": [\"17525277\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"miR-200 family microRNAs directly target the 3'-UTR of WAVE3 mRNA and inhibit its expression. miR-200-mediated downregulation of WAVE3 reduces cancer cell invasiveness and causes morphological changes resembling mesenchymal-to-epithelial transition. Re-expression of a miR-200-resistant WAVE3 reverses these effects, confirming specificity.\",\n      \"method\": \"Luciferase 3'-UTR reporter assay, Western blotting, invasion assay, rescue experiment with miR-200-resistant WAVE3\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct 3'-UTR targeting confirmed by reporter assay, functional rescue with resistant construct, multiple orthogonal methods\",\n      \"pmids\": [\"19801681\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"WAVE3/WASF3 knockdown leads to upregulation of the KISS1 metastasis suppressor gene, elevated IκBα levels in the cytoplasm, and reduced nuclear NF-κB (p65/50). Knockdown of KISS1 in WASF3-silenced cells recovers the invasion phenotype. TNF-α treatment has no effect on invasion or NF-κB nuclear translocation in WASF3 knockdown cells, placing WASF3 upstream of NF-κB/KISS1 in the regulation of MMP-9 production.\",\n      \"method\": \"shRNA knockdown, oligonucleotide arrays, luciferase reporter (KISS1 transcription), Western blotting (IκBα/NF-κB), rescue knockdown of KISS1, TNF-α treatment\",\n      \"journal\": \"International journal of cancer\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal methods (transcriptomics, reporter assay, rescue KD, pathway protein analysis), epistasis established by double-KD rescue\",\n      \"pmids\": [\"21544801\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"miR-31 directly targets the 3'-UTR of WAVE3 mRNA and inhibits its expression. Loss of miR-31 correlates with increased WAVE3 in invasive breast cancer. Re-expression of miR-31-resistant WAVE3 reverses miR-31-mediated inhibition of cancer cell invasion.\",\n      \"method\": \"3'-UTR targeting validation, Western blotting, invasion assay, rescue with miR-31-resistant WAVE3\",\n      \"journal\": \"International journal of cancer\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct 3'-UTR targeting, functional rescue with resistant construct, correlation in cell lines and human tumors\",\n      \"pmids\": [\"21105030\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"HSP90 is present in the WASF3/WAVE3 immunocomplex from prostate cancer cells. Inactivation of HSP90 does not affect WASF3 stability but prevents its phosphoactivation by destabilizing ABL kinase. HSP70 is also present in the WASF3 immunocomplex; inactivation of HSP70 leads to WASF3 destabilization through proteasome degradation. Overexpression of HSP70 in WASF3-null cells does not enhance invasion.\",\n      \"method\": \"Mass spectrometry, co-immunoprecipitation, pharmacological inhibition of HSP90/70, shRNA knockdown, proteasome inhibition assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Moderate — mass spectrometry identification, Co-IP, multiple functional validations (stability vs phospho-activation distinction), negative control (HSP70 OE in null cells)\",\n      \"pmids\": [\"22315230\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"HIF1A binds to hypoxia response elements (HRE) in the WASF3 promoter under hypoxic conditions, as shown by ChIP assay, and induces WASF3 transcription. Hypoxia also increases WASF3 phosphoactivation. WASF3 knockdown cells show no motility response to hypoxia.\",\n      \"method\": \"Chromatin immunoprecipitation (ChIP), luciferase reporter assay, scratch wound motility assay, Western blotting for phospho-WASF3\",\n      \"journal\": \"International journal of cancer\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP demonstrating direct HIF1A binding, luciferase reporter, functional validation with KD; multiple orthogonal methods in single study\",\n      \"pmids\": [\"22581642\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Loss of WAVE3 in cancer cells leads to inhibition of NFκB signaling through decreased nuclear translocation of NFκB, resulting in loss of activation of NFκB target genes (including MMP9), inhibition of invadopodia formation and ECM degradation, and sensitization of cancer cells to TNFα-induced apoptosis through inhibition of the AKT pro-survival pathway. Conversely, overexpression of WAVE3 enhances NFκB activity.\",\n      \"method\": \"WAVE3 knockdown/overexpression, NFκB nuclear translocation assay, invadopodia formation assay, ECM degradation assay, MMP9 expression analysis, apoptosis assay\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple functional readouts with both loss- and gain-of-function approaches; single lab\",\n      \"pmids\": [\"25329315\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"WASF3/WAVE3 overexpression increases ZEB1/2 levels, which suppress the miR-200a/200b/429 cluster. This occurs through WASF3-mediated downregulation of KISS1, which releases IκBα inhibition of NFκB, and ZEB1 expression is regulated by NFκB. Knockdown of WASF3 leads to reduced ZEB1 levels, increased miR-200 and E-cadherin levels, and loss of invasion.\",\n      \"method\": \"WASF3 overexpression/knockdown, Western blotting, luciferase reporter, NFκB pathway analysis\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — pathway dissection by gain/loss-of-function with multiple molecular readouts; single lab\",\n      \"pmids\": [\"23318438\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"IL-6 induces WASF3 expression and phosphoactivation through the JAK2/STAT3 pathway in two ways: (1) STAT3 directly binds the WASF3 promoter and increases transcription; (2) JAK2 interacts with WASF3 and directly activates (phosphorylates) it. Inhibition of STAT3 (shRNA, dominant negative, or S3I-201) reduces WASF3 levels and migration. Inhibition of JAK2 (shRNA or AG490) reduces WASF3 activation and prevents its membrane localization.\",\n      \"method\": \"ChIP demonstrating STAT3 binding to WASF3 promoter, Co-IP (JAK2-WASF3), shRNA, dominant-negative constructs, pharmacological inhibition, migration assay, membrane localization assay\",\n      \"journal\": \"Carcinogenesis\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP, Co-IP, pharmacological and genetic inhibition with functional readouts; multiple orthogonal methods\",\n      \"pmids\": [\"23677069\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"TGF-β selectively and robustly induces WAVE3 expression in metastatic breast cancer cells through a Smad2- and β3 integrin-dependent mechanism. WAVE3 is required for TGF-β-induced EMT: stable depletion of WAVE3 prevents TGF-β from inducing EMT programs, lamellipodia formation, and cell migration.\",\n      \"method\": \"TGF-β treatment with downstream Western blotting, stable WAVE3 knockdown, EMT marker analysis, lamellipodia assay, 3D organotypic culture, in vivo syngeneic mouse model\",\n      \"journal\": \"Breast cancer research and treatment\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — pathway placement by genetic KD with multiple readouts (EMT markers, lamellipodia, migration), in vitro and in vivo validation\",\n      \"pmids\": [\"24197660\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"ATAD3A (mitochondrial membrane protein) interacts with WASF3/WAVE3 as demonstrated by mass spectrometry. ATAD3A knockdown decreases WASF3 protein levels. HSP70 stabilizes WASF3 in the cytoplasm, but inactivation of HSP70 does not destabilize WASF3 at the mitochondrial membrane where ATAD3A protects it. GRP78 upregulation (during ER stress) increases WASF3 levels, and ATAD3A is present in a WASF3-GRP78 complex. Suppression of GRP78 leads to ATAD3A-dependent destabilization of WASF3 at the mitochondrial membrane. The N-terminal end of WASF3 is within the mitochondria and is the interaction site with the N-terminal end of ATAD3A.\",\n      \"method\": \"Mass spectrometry, co-immunoprecipitation, shRNA knockdown, mitochondrial fractionation with proteolysis protection assay, in vivo xenograft\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Moderate — mass spectrometry interaction, mitochondrial proteolysis domain mapping, multiple KD validations, Co-IP\",\n      \"pmids\": [\"25823022\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Genetic knockdown of CYFIP1 in cancer cells destabilizes the WASF3 complex, causes loss of WASF3 function, and suppresses invasion. Stapled peptides (WAHM) targeting the α-helical interface between WASF3 and CYFIP1 suppress motility and invasion in breast and prostate cancer cells, suppress Rac interaction with the WASF3 complex, and dysregulate downstream targets MMP-9 and KISS1. Depletion of WASF1 and WASF2, which also bind CYFIP1, did not affect invasion, demonstrating specificity for WASF3.\",\n      \"method\": \"shRNA knockdown of CYFIP1, stapled peptide treatment, invasion/motility assay, Rac interaction assay (Co-IP), MMP-9 and KISS1 expression analysis\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic and pharmacologic disruption of defined protein-protein interface with multiple functional readouts including RAC binding; paralog specificity controls\",\n      \"pmids\": [\"26676744\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"WASF3 is present in the HER2 immunocomplex. Suppression of WASF3 suppresses invasion even in the presence of HER2 expression. WASF3's ability to promote invasion is highly dependent on the HER2/HER3 heterodimer. The HER2/HER3 complex facilitates WASF3 phospho-activation and transcriptional upregulation through HER2/HER3 activation of JAK/STAT signaling.\",\n      \"method\": \"Co-immunoprecipitation, WASF3 suppression/overexpression, invasion assays with HER2/HER3 manipulation, Western blotting (phospho-WASF3)\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP establishing complex, functional epistasis, JAK/STAT pathway linking; single lab\",\n      \"pmids\": [\"26804171\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"NCKAP1 is required for WASF3 complex stability and function: silencing NCKAP1 destabilizes the WASF3 complex and suppresses invasive capacity of breast, prostate, and colon cancer cells and metastasis in vivo. Activation of the WASF3 complex requires RAC1 interaction, and inactivation of NCKAP1 prevents RAC1 association with the WASF3 complex. Stapled peptides (WANT3) targeting the NCKAP1-CYFIP1 interface destabilize the WASF3 complex and suppress RAC1 binding and invasion.\",\n      \"method\": \"shRNA knockdown of NCKAP1, Co-IP (RAC1-WASF3 complex), in vivo spontaneous metastasis model, stapled peptide treatment\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic KD with in vitro and in vivo validation, RAC1 binding mechanistic data by Co-IP, pharmacologic disruption; replicated across multiple cancer cell types\",\n      \"pmids\": [\"27432794\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"WAVE3 interacts with YB1 (Y-box binding protein 1), and this interaction is required for YB1 nuclear translocation in cancer cells and activation of transcription of cancer stem cell-specific genes. WAVE3 is enriched in the CSC subpopulation. CRISPR/Cas9 knockout of WAVE3 attenuates the CSC subpopulation and inhibits transcription of CSC transcription factors.\",\n      \"method\": \"CRISPR/Cas9 knockout, Co-immunoprecipitation (WAVE3-YB1 interaction), nuclear/cytoplasmic fractionation, CSC marker analysis, transcription factor activity assays\",\n      \"journal\": \"Oncotarget\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — CRISPR KO with functional readout, Co-IP for interaction, localization assay; single lab\",\n      \"pmids\": [\"29262622\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"p63α transcriptionally upregulates HSP70 (Hsp70) expression via E2F1, and HSP70 promotes bladder cancer cell invasion through the Hsp70/WASF3/MMP-9 axis.\",\n      \"method\": \"Western blotting, invasion assay, transcription analysis (E2F1/p63α-mediated Hsp70 upregulation), pathway inhibition\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — mechanistic pathway placement using molecular biology in a single lab study; consistent with prior HSP70-WASF3 interaction data\",\n      \"pmids\": [\"28794159\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Mutant RAS promotion of invasion and metastasis is dependent on WASF3 activation in a PI3K and AKT-dependent manner. AKT is present in the WASF3 immunocomplex and this association is enhanced by mutant RAS overexpression. Mutant RAS promotes dissociation of p85 from the WASF3 complex, promoting activation of p110. ERK1/2 activation is not affected by loss of WASF3.\",\n      \"method\": \"Co-immunoprecipitation (AKT in WASF3 complex), proteomics analysis, WASF3 knockdown epistasis, p85/p110 dissociation assay\",\n      \"journal\": \"Genes, chromosomes & cancer\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP with proteomics, genetic epistasis; single lab, moderate mechanistic depth\",\n      \"pmids\": [\"28233357\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Wasf3 null mice (generated by deletion of exons 4 and 5) are viable with no visible morphological or behavioral abnormalities and no abnormal mammary gland development or brain development. In the MMTV-polyoma middle-T oncogene breast cancer model, Wasf3 is upregulated in metastatic lesions, and Wasf3 null background reduces the number and size of metastatic lung lesions without affecting primary tumor development.\",\n      \"method\": \"Conditional knockout mouse model (Cre-lox deletion), MMTV-PyMT spontaneous breast cancer model, histological analysis of metastases\",\n      \"journal\": \"The American journal of pathology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo genetic KO with spontaneous cancer model, quantitative metastasis assessment; rigorous mammalian genetics\",\n      \"pmids\": [\"31542393\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"PCARE (C2orf71) interacts with WASF3/WAVE3 and efficiently recruits it to the primary cilium. Ectopic coexpression of PCARE and WASF3 in ciliated cells causes remarkable expansion of the ciliary tip via actin polymerization. This expansion is disrupted by siRNA knockdown of actin regulators, pharmacological inhibition of actin polymerization, or PCARE with a retinal dystrophy-associated missense mutation. In mouse retina and human retinal organoids, PCARE and WASF3 colocalize with actin at the outer segment base where this process drives disk formation initiation.\",\n      \"method\": \"Co-immunoprecipitation (PCARE-WASF3), ectopic co-expression in ciliated cells, siRNA knockdown, pharmacological actin inhibition, mouse retinal imaging, human retinal organoids\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal interaction, functional co-expression system, multiple disruption methods, in vivo model validation, disease mutation support\",\n      \"pmids\": [\"32312818\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"WAVE3 tyrosine phosphorylation (downstream of PI3K) is also achieved downstream of TGF-β and EGF signaling, and is required for oncogenic activity including migration, tumorsphere growth, and invasion. Loss of WAVE3 phosphorylation also inhibits the activation of PI3K, TGF-β, and EGF signaling downstream effectors, identifying a positive feedback loop between WAVE3 phosphorylation and these pathways.\",\n      \"method\": \"Phospho-mutant WAVE3 constructs, Western blotting for pathway effectors, migration assay, 3D tumorsphere assay, mouse xenograft model\",\n      \"journal\": \"Oncogenesis\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — phospho-mutant approach, multiple pathway readouts, in vitro and in vivo; single lab\",\n      \"pmids\": [\"33012785\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Phosphorylation of the proline-rich domain (PRD) of WAVE3 is essential for its interaction with YB1. Loss of PRD phosphorylation inhibits WAVE3-YB1 interaction, prevents YB1-mediated activation of CSC markers, and inhibits WAVE3-mediated EMT activation. PRD phosphorylation is required for migration and invasion in vitro and tumor growth/metastasis in vivo.\",\n      \"method\": \"Phospho-mutant PRD constructs, co-immunoprecipitation (WAVE3-YB1), invasion/migration assays, CSC marker analysis, in vivo xenograft\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — phospho-mutant with defined domain, Co-IP demonstrating interaction requirement; single lab\",\n      \"pmids\": [\"33594155\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"SHOX2 directly activates WASF3 transcription and recruits STAT3 to the WASF3 promoter, where SHOX2 and STAT3 form a functional immunocomplex that cooperatively promotes WASF3 transcriptional activity. WASF3 knockdown abrogates SHOX2-induced metastasis but not SHOX2-dependent tumorigenesis.\",\n      \"method\": \"ChIP-qPCR and ChIP/re-ChIP (SHOX2 and STAT3 at WASF3 promoter), co-immunoprecipitation (SHOX2-STAT3 complex), shRNA knockdown epistasis, in vivo orthotopic mouse model\",\n      \"journal\": \"Journal of experimental & clinical cancer research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP/re-ChIP with Co-IP establishing transcriptional complex, genetic epistasis in vivo; multiple orthogonal methods\",\n      \"pmids\": [\"34465361\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Overexpression of WASF3 disrupts mitochondrial respiratory supercomplex formation and is associated with ER stress. In transgenic mice with increased WASF3 expression, treadmill running capacity was markedly decreased with concomitantly impaired respiratory supercomplex assembly and reduced complex IV levels in skeletal muscle mitochondria. ER stress-induced WASF3 upregulation (by endotoxin) also decreased skeletal muscle complex IV levels. Pharmacologic inhibition of ER stress decreased WASF3 levels and improved mitochondrial function in patient cells.\",\n      \"method\": \"WASF3 transgenic mouse model, treadmill exercise testing, mitochondrial supercomplex assembly analysis (BN-PAGE or equivalent), complex IV activity measurement, pharmacological ER stress inhibition, patient skeletal muscle biopsy analysis\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — transgenic mouse model with quantitative functional phenotype, biochemical supercomplex analysis, pharmacologic rescue, patient tissue validation; multiple orthogonal methods\",\n      \"pmids\": [\"37579159\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"WAVE3/WASF3 phosphorylation is required for β-catenin stabilization; loss of WAVE3 expression or phosphorylation inhibits β-catenin activity and expression. Dual blocking of WAVE3 expression or phosphorylation in combination with chemotherapy suppresses chemoresistant TNBC cell behavior in vitro and in vivo. Re-expression of phospho-active WAVE3 restores oncogenic activity, while phospho-mutant WAVE3 does not.\",\n      \"method\": \"CRISPR/Cas9 KO, phospho-mutant rescue, Western blotting (β-catenin), 2D/3D invasion assay, xenograft assay\",\n      \"journal\": \"Breast cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — CRISPR KO and phospho-mutant rescue with β-catenin mechanistic link; single lab\",\n      \"pmids\": [\"36949468\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"METTL3 mediates m6A modification of WASF3 mRNA. IGF2BP2 binds to the m6A site in the 3'-UTR of WASF3 mRNA and enhances WASF3 translation. Highly expressed WASF3 activates the MAPK signaling pathway by interacting with phosphorylated p38 (p-p38). Removal of m6A modification of WASF3 mRNA inhibited WASF3 expression and abolished WASF3's ability to bind p-p38 and activate MAPK signaling.\",\n      \"method\": \"Co-immunoprecipitation (METTL3-WASF3, WASF3-p-p38), m6A modification assay, Western blotting, m6A inhibition experiments, ESCC cell functional assays\",\n      \"journal\": \"MedComm\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP for WASF3-p-p38 interaction, m6A modification/translation mechanism; single lab, single study\",\n      \"pmids\": [\"41127505\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"WASF3/WAVE3 is a WASP-family actin cytoskeleton remodeling protein that functions as a key node in cancer invasion and metastasis: it localizes to lamellipodia downstream of PI3K (via interaction with the p85 subunit) and is phosphoactivated on four tyrosine residues by c-Abl kinase (facilitated by HER2/HER3 via JAK/STAT signaling); it is stabilized by HSP70 in the cytoplasm and by ATAD3A/GRP78 at the mitochondrial membrane; it is transcriptionally induced by HIF1A, STAT3 (bound directly to its promoter downstream of JAK2/IL-6), and the SHOX2-STAT3 complex; its activity promotes invasion through NFκB-MMP9 signaling, KISS1 suppression, and ZEB1-mediated miR-200 inactivation; it regulates cancer stem cell maintenance through its PRD-phosphorylation-dependent interaction with YB1 nuclear translocation; it forms a pentameric WASF Regulatory Complex with NCKAP1, CYFIP1, ABI, and BRK1 that requires RAC1 interaction for activation; in photoreceptors, WASF3 is recruited by PCARE to the ciliary tip to drive actin-dependent outer segment disk formation; and, in a non-cancer context, WASF3 overexpression disrupts mitochondrial respiratory supercomplex assembly (reducing complex IV levels), potentially mediating exercise intolerance in ME/CFS.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"WASF3 (WAVE3) is a WASP-family actin-cytoskeleton regulator that drives cell motility and serves as a central node for cancer invasion and metastasis [#0, #5]. It localizes to lamellipodia at the leading edge of migrating cells downstream of PI3K signaling, where its N-terminal region binds the p85 regulatory subunit; loss of WASF3 abolishes PDGF-induced lamellipodia and migration [#0]. WASF3 functions within a CYFIP1/NCKAP1-containing regulatory complex whose stability and RAC1-dependent activation require both subunits, and disruption of the WASF3-CYFIP1 or NCKAP1-CYFIP1 interfaces with stapled peptides collapses the complex, blocks RAC1 binding, and suppresses invasion in breast, prostate, and colon cancer cells with paralog specificity over WASF1/WASF2 [#16, #18]. Its activity is gated by tyrosine phosphorylation: c-Abl phosphorylates four tyrosine residues downstream of PDGF, and equivalent phosphoactivation is achieved downstream of TGF-\\u03b2, EGF, and HER2/HER3 via JAK2/STAT3 signaling, establishing a positive feedback loop with these pathways [#4, #13, #17, #24]. Activated WASF3 promotes invasion through an NF\\u03baB\\u2013MMP9 axis, suppression of the KISS1 metastasis-suppressor and consequent ZEB1-driven inactivation of the miR-200 family during EMT [#7, #11, #12], and through proline-rich-domain-phosphorylation-dependent binding to YB1 that drives YB1 nuclear translocation and cancer stem-cell gene transcription [#19, #25]. WASF3 abundance is controlled at multiple levels: transcriptionally by HIF1A under hypoxia, by STAT3 and the SHOX2-STAT3 complex at its promoter, and post-transcriptionally by miR-200 and miR-31 targeting its 3'-UTR; its protein is stabilized by HSP70 in the cytoplasm and by ATAD3A/GRP78 at the mitochondrial membrane, where its N-terminus engages the ATAD3A N-terminus [#6, #8, #10, #15, #26]. Genetic ablation in mice confirms WASF3 is dispensable for development but required for metastatic dissemination in the MMTV-PyMT model [#22]. Beyond cancer, WASF3 is recruited by PCARE to the photoreceptor ciliary tip to drive actin-dependent outer-segment disk formation [#23], and its overexpression disrupts mitochondrial respiratory supercomplex assembly with reduced complex IV, linking ER-stress-induced WASF3 to exercise intolerance [#27].\",\n  \"teleology\": [\n    {\n      \"year\": 2005,\n      \"claim\": \"Established WASF3 as an actin-remodeling effector acting downstream of PI3K to build lamellipodia, defining its core cellular role in migration.\",\n      \"evidence\": \"Microscopy localization, RNAi knockdown of PDGF-induced lamellipodia, yeast two-hybrid and Co-IP mapping the WAVE3 N-terminus to p85 SH2 domain, and LY294002 inhibition\",\n      \"pmids\": [\"15826941\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not establish how p85 binding mechanistically couples to Arp2/3 activation\", \"Direct actin-nucleation activity of WASF3 not measured\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Connected WASF3 to invasion machinery by showing it sustains p38 MAPK signaling and MMP expression, distinguishing it functionally from WAVE1/WAVE2.\",\n      \"evidence\": \"siRNA knockdown with Western blots of phospho-kinases, MMP expression, and actin/focal-adhesion staining\",\n      \"pmids\": [\"15907837\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether WASF3 directly regulates p38 or acts indirectly was not resolved\", \"MMP regulation mechanism left undefined\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Placed WASF3 within the same regulatory complexes as WAVE1/WAVE2 and identified LDOC1 as a VH-domain partner whose nuclear export WASF3 drives to suppress apoptosis.\",\n      \"evidence\": \"Co-IP interaction assays across isoforms; direct binding, fractionation, and apoptosis assays for LDOC1\",\n      \"pmids\": [\"15752430\", \"16272576\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"LDOC1 regulation not tested in vivo\", \"Complex composition not yet resolved as a defined pentamer\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Defined the activation switch by showing c-Abl phosphorylates four WASF3 tyrosines as a requirement for motility, converting WASF3 into a kinase-gated effector.\",\n      \"evidence\": \"Co-IP, in vitro kinase assay, tyrosine site-directed mutagenesis, STI-571 inhibition, migration assays\",\n      \"pmids\": [\"17623672\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural consequence of tyrosine phosphorylation on complex conformation unknown\", \"Whether all four sites are equivalent not dissected\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Demonstrated WASF3 is required for metastasis in vivo and operates through a p38-MAPK pathway, validating it as a metastasis driver beyond cell culture.\",\n      \"evidence\": \"shRNA stable knockdown, Matrigel invasion, lung colonization and orthotopic xenografts, dominant-negative p38 epistasis\",\n      \"pmids\": [\"17525277\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How WASF3 feeds into p38 activation not mechanistically pinned\", \"Primary tumor versus metastasis-specific effects not fully separated\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Identified post-transcriptional control of WASF3 by miR-200 (and later miR-31) targeting its 3'-UTR, tying WASF3 levels to EMT/MET state.\",\n      \"evidence\": \"Luciferase 3'-UTR reporters, Western blots, invasion assays, rescue with miRNA-resistant WASF3 constructs\",\n      \"pmids\": [\"19801681\", \"21105030\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether endogenous miRNA changes fully account for tumor WASF3 levels not quantified\", \"Interplay between the two miRNAs not addressed\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Mapped the invasion output pathway, placing WASF3 upstream of NF\\u03baB to repress the KISS1 metastasis suppressor and drive MMP-9 production.\",\n      \"evidence\": \"shRNA knockdown, oligonucleotide arrays, KISS1 luciferase reporter, I\\u03baB\\u03b1/NF\\u03baB Western blots, KISS1 rescue knockdown, TNF-\\u03b1 treatment\",\n      \"pmids\": [\"21544801\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct molecular link between cytoplasmic WASF3 and NF\\u03baB activation not identified\", \"Whether KISS1 repression is transcriptional or indirect not fully defined\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Extended the invasion axis to invadopodia and survival, and established hypoxic transcriptional induction of WASF3 by HIF1A binding to its promoter.\",\n      \"evidence\": \"Knockdown/overexpression with invadopodia and ECM degradation assays, TNF\\u03b1 apoptosis assays; ChIP, luciferase reporter and motility assays for HIF1A\",\n      \"pmids\": [\"25329315\", \"22581642\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"HIF1A and other inputs not integrated quantitatively\", \"AKT pro-survival linkage mechanism not detailed\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Resolved chaperone-dependent stability and activation, separating HSP70-dependent WASF3 protein stability from HSP90-dependent ABL stability needed for phosphoactivation.\",\n      \"evidence\": \"Mass spectrometry, Co-IP, HSP90/70 pharmacological inhibition, shRNA, proteasome inhibition, and HSP70-overexpression negative control in null cells\",\n      \"pmids\": [\"22315230\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"E3 ligase mediating WASF3 proteasomal turnover not identified\", \"Structural basis of HSP70 binding unknown\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Built the EMT regulatory loop (KISS1\\u2192NF\\u03baB\\u2192ZEB1\\u2192miR-200) and established cytokine-driven dual control of WASF3 by JAK2/STAT3 transcription and JAK2 phosphorylation.\",\n      \"evidence\": \"Gain/loss-of-function Westerns and reporters for ZEB1/miR-200; ChIP, JAK2-WASF3 Co-IP, dominant-negative and pharmacological inhibition with migration/membrane-localization assays\",\n      \"pmids\": [\"23318438\", \"23677069\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Feedback stability of the ZEB1/miR-200 loop not modeled\", \"Relative contribution of transcriptional versus phospho-activation by JAK2 not weighted\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Positioned WASF3 as an obligatory effector of TGF-\\u03b2-induced EMT via Smad2 and \\u03b23-integrin, broadening its upstream signaling repertoire.\",\n      \"evidence\": \"TGF-\\u03b2 treatment with Westerns, stable knockdown, EMT marker analysis, lamellipodia assay, 3D organotypic culture, syngeneic mouse model\",\n      \"pmids\": [\"24197660\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct Smad2 target status of WASF3 promoter not shown\", \"Integrin requirement mechanism not resolved\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Defined a mitochondrial pool of WASF3 stabilized by ATAD3A and GRP78, distinct from the cytoplasmic HSP70-protected pool, and mapped the N-terminal interaction interface.\",\n      \"evidence\": \"Mass spectrometry, Co-IP, shRNA, mitochondrial fractionation with proteolysis protection mapping, in vivo xenograft\",\n      \"pmids\": [\"25823022\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Functional role of mitochondrial WASF3 in invasion versus respiration not delineated at this stage\", \"Whether mitochondrial WASF3 is phosphoactivated unknown\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Established the WASF3 regulatory complex as a druggable, RAC1-dependent assembly by showing CYFIP1 and NCKAP1 are required for stability and RAC1 binding, with paralog-specific peptide inhibition.\",\n      \"evidence\": \"shRNA knockdown of CYFIP1/NCKAP1, RAC1-complex Co-IP, WAHM/WANT3 stapled peptides, invasion/motility assays, in vivo metastasis model, WASF1/WASF2 specificity controls\",\n      \"pmids\": [\"26676744\", \"27432794\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"High-resolution structure of the WASF3 complex not determined\", \"Why WASF3 is uniquely required over paralogs not explained molecularly\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Linked receptor tyrosine kinase oncogenic signaling to WASF3 by showing HER2/HER3 drives both its phosphoactivation and transcription through JAK/STAT, making WASF3 a HER2-pathway effector.\",\n      \"evidence\": \"Co-IP placing WASF3 in the HER2 complex, suppression/overexpression with HER2/HER3 manipulation, phospho-WASF3 Westerns and invasion epistasis\",\n      \"pmids\": [\"26804171\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct versus indirect HER2-WASF3 association not distinguished\", \"Single-lab finding without reciprocal in vivo validation\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Uncovered a nuclear-transcriptional arm of WASF3 function: PRD-phosphorylation-dependent binding to YB1 drives YB1 nuclear import and cancer stem-cell gene programs.\",\n      \"evidence\": \"CRISPR/Cas9 knockout, WAVE3-YB1 Co-IP, nuclear/cytoplasmic fractionation, CSC marker and transcription factor activity assays; phospho-mutant PRD constructs\",\n      \"pmids\": [\"29262622\", \"33594155\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Kinase phosphorylating the PRD not identified\", \"How a cytoskeletal protein localizes to drive nuclear YB1 import mechanistically unclear\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Integrated WASF3 into RAS-PI3K-AKT oncogenic signaling, showing mutant RAS activates WASF3 by dissociating p85 to free p110, with AKT in the WASF3 complex.\",\n      \"evidence\": \"AKT-WASF3 Co-IP, proteomics, knockdown epistasis, p85/p110 dissociation assays\",\n      \"pmids\": [\"28233357\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct versus scaffolded AKT association not resolved\", \"Single-lab finding\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Connected upstream transcriptional control of the chaperone arm by showing p63\\u03b1/E2F1-driven HSP70 promotes invasion through the HSP70/WASF3/MMP-9 axis.\",\n      \"evidence\": \"Westerns, invasion assays, transcriptional analysis of p63\\u03b1/E2F1-mediated HSP70 induction, pathway inhibition\",\n      \"pmids\": [\"28794159\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct effect on WASF3 stability versus indirect not separated here\", \"Cancer-type generality not tested\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Provided definitive germline genetics: WASF3 is dispensable for normal development but specifically required for metastatic spread in a spontaneous tumor model.\",\n      \"evidence\": \"Wasf3-null mouse (exon 4/5 deletion), MMTV-PyMT spontaneous breast cancer model, histological metastasis quantification\",\n      \"pmids\": [\"31542393\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Cell-autonomous versus microenvironmental contribution not dissected\", \"Compensation by WASF1/WASF2 during development not assessed\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Generalized WASF3 phosphorylation as a feed-forward hub, required for and reinforcing PI3K, TGF-\\u03b2, and EGF signaling outputs.\",\n      \"evidence\": \"Phospho-mutant constructs, pathway-effector Westerns, migration, 3D tumorsphere assays, mouse xenografts\",\n      \"pmids\": [\"33012785\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular basis of feedback onto upstream receptors unknown\", \"Identity of all kinases acting on each site unresolved\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Revealed a non-cancer ciliary role: PCARE recruits WASF3 to the photoreceptor ciliary tip to drive actin-dependent outer-segment disk formation.\",\n      \"evidence\": \"PCARE-WASF3 Co-IP, ectopic co-expression in ciliated cells, siRNA and pharmacological actin disruption, retinal-dystrophy mutation, mouse retina and human retinal organoids\",\n      \"pmids\": [\"32312818\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether ciliary WASF3 uses the same RAC1-dependent complex as in cancer not tested\", \"Endogenous WASF3 requirement in vivo not genetically confirmed in retina\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Identified a cooperative transcriptional driver, the SHOX2-STAT3 promoter complex, that selectively governs WASF3-dependent metastasis rather than tumorigenesis.\",\n      \"evidence\": \"ChIP/re-ChIP of SHOX2 and STAT3 at the WASF3 promoter, SHOX2-STAT3 Co-IP, shRNA epistasis, orthotopic mouse model\",\n      \"pmids\": [\"34465361\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How SHOX2-STAT3 selectively affects metastatic but not tumorigenic programs unclear\", \"Other promoter co-factors not enumerated\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Linked WASF3 phosphorylation to \\u03b2-catenin stabilization and chemoresistance, identifying a co-targeting strategy in triple-negative breast cancer.\",\n      \"evidence\": \"CRISPR/Cas9 KO, phospho-mutant rescue, \\u03b2-catenin Westerns, 2D/3D invasion, xenografts\",\n      \"pmids\": [\"36949468\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct versus indirect \\u03b2-catenin regulation not resolved\", \"Single-lab finding\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Established a distinct pathological role outside cancer: WASF3 overexpression impairs mitochondrial respiratory supercomplex assembly and reduces complex IV, modeling exercise intolerance in ME/CFS.\",\n      \"evidence\": \"WASF3 transgenic mice, treadmill testing, supercomplex assembly and complex IV analysis, ER-stress induction and pharmacological inhibition, patient muscle biopsies\",\n      \"pmids\": [\"37579159\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular mechanism by which WASF3 disrupts supercomplex assembly not defined\", \"Connection to its actin/invasion functions unestablished\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Added m6A-based translational control, with METTL3/IGF2BP2 enhancing WASF3 translation to enable p-p38 binding and MAPK activation.\",\n      \"evidence\": \"METTL3-WASF3 and WASF3-p-p38 Co-IP, m6A modification assays, m6A inhibition, ESCC functional assays\",\n      \"pmids\": [\"41127505\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct WASF3-p-p38 interaction surface not mapped\", \"Single-lab, single cancer-type finding\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How the cytoplasmic actin-regulatory WASF3 complex is mechanistically reconciled with its nuclear (YB1), mitochondrial (ATAD3A/supercomplex), and ciliary (PCARE) functions, and the structural basis of its phospho-activation, remain unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No high-resolution structure of the activated WASF3 complex\", \"Kinases for the PRD and the mechanism of supercomplex disruption unknown\", \"Whether the same RAC1-dependent complex operates across all compartments untested\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [0, 16, 18, 23]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [3, 19, 25]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [7, 11, 12]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [0, 13]},\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [0, 1, 23]},\n      {\"term_id\": \"GO:0005739\", \"supporting_discovery_ids\": [15, 27]},\n      {\"term_id\": \"GO:0005929\", \"supporting_discovery_ids\": [23]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [19, 25]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [0, 4, 13, 21]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [5, 22, 28]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [10, 13, 26]},\n      {\"term_id\": \"R-HSA-1266738\", \"supporting_discovery_ids\": [14, 23]},\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [27]}\n    ],\n    \"complexes\": [\"WASF Regulatory Complex (WASF3-CYFIP1-NCKAP1-ABI-BRK1)\"],\n    \"partners\": [\"CYFIP1\", \"NCKAP1\", \"RAC1\", \"YB1\", \"ATAD3A\", \"PCARE\", \"JAK2\", \"ABL1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":8,"faith_total":8,"faith_pct":100.0}}