{"gene":"ASAP1","run_date":"2026-06-09T22:02:44","timeline":{"discoveries":[{"year":1998,"finding":"ASAP1 was purified and cloned as a PIP2-dependent Arf1 GTPase-activating protein (GAP). The PH, zinc finger, and ANK repeat regions together possess PIP2-dependent GAP activity on Arf1 and Arf5 in vitro. ASAP1 associates with the SH3 domains of Src family members and Crk adapter protein in vitro, coprecipitates with Src from cell lysates, and is phosphorylated on tyrosine residues in cells expressing activated Src through the same proline-rich class II Src SH3 binding site.","method":"Protein purification, in vitro GAP assay, GST pulldown, co-immunoprecipitation, in-cell phosphorylation","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — direct biochemical reconstitution of GAP activity, multiple orthogonal binding assays, replicated across multiple Arf substrates","pmids":["9819391"],"is_preprint":false},{"year":2000,"finding":"ASAP1 localizes to focal adhesions and cycles with focal adhesion proteins during cell migration. Overexpression of ASAP1 altered focal adhesion morphology and blocked cell spreading and PDGF-induced dorsal ruffle formation; a GAP-inactive mutant had reduced effect on spreading and increased dorsal ruffle formation, demonstrating that GAP activity is required for these cytoskeletal functions.","method":"Fluorescence microscopy, cell spreading assay, PDGF-stimulation assay, overexpression of wild-type vs. catalytically inactive mutant","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct localization with functional consequence, loss-of-function and gain-of-function with defined cellular phenotype, replicated findings","pmids":["10725410"],"is_preprint":false},{"year":2000,"finding":"PIP2 activates ASAP1 GAP activity by binding the PH domain, which acts as an allosteric site rather than merely a membrane recruitment signal. The PH domain is necessary for GAP activity even in the absence of phospholipids; PIP2 binding causes a conformational change in the Arf GAP domain. Activation and membrane recruitment can be uncoupled.","method":"In vitro GAP activity assays, limited proteolysis, domain deletion/mutation analysis","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — reconstitution in vitro with mutagenesis and multiple orthogonal biochemical methods","pmids":["10734117"],"is_preprint":false},{"year":2001,"finding":"ASAP1 (DEF-1) functions as an ARF GAP for ARF1 but not ARF6 in vivo (cell-based ARF GAP assay), unlike ACAP1, ACAP2, and ARFGAP1 which act on both. Enhancement of cell motility by ASAP1 is dependent on GAP activity, whereas inhibition of cell spreading by ASAP1 is GAP-activity independent, indicating two distinct downstream pathways.","method":"Cell-based ARF GAP assay, stable overexpression, cell motility assay, GAP-domain deletion mutant","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — cell-based GAP assay with direct comparison of substrates and domain mutants showing mechanistic separation of phenotypes","pmids":["11773070"],"is_preprint":false},{"year":2002,"finding":"ASAP1 binds to focal adhesion kinase (FAK) via an interaction between the C-terminal SH3 domain of ASAP1 and the second proline-rich motif in the C-terminal region of FAK. This interaction contributes to focal adhesion assembly; ASAP1 variants that could not bind FAK or lacked GAP activity showed reduced inhibition of cell spreading and failed to prevent paxillin/FAK organization in focal adhesions.","method":"Affinity chromatography, yeast two-hybrid, GST pulldown, co-immunoprecipitation, cell spreading assay, overexpression of mutants","journal":"Molecular biology of the cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP and pulldown confirming direct interaction, mutagenesis linking binding to cellular function","pmids":["12058076"],"is_preprint":false},{"year":2003,"finding":"CrkL binds ASAP1 via its N-terminal SH3 domain and directs ASAP1 to peripheral focal adhesions. CrkL co-expression recruited endogenous and exogenous ASAP1 to CrkL-induced focal adhesions; an SH2-mutated CrkL that cannot localize to focal adhesions failed to recruit ASAP1.","method":"Pulldown/mass spectrometry, co-immunoprecipitation, fluorescence microscopy, overexpression of wild-type vs. SH2-mutant CrkL","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct identification in platelets, confirmed in cells with mutagenesis; single lab","pmids":["12522101"],"is_preprint":false},{"year":2003,"finding":"Pyk2 interacts with ASAP1 through the proline-rich regions of Pyk2 and the SH3 domain of ASAP1. Pyk2 directly phosphorylates ASAP1 on tyrosine residues (Y308 and Y782) in vitro and in cells; this phosphorylation inhibits ASAP1 GAP activity toward Arf1 as measured by fluorimetric GTPase assay.","method":"Yeast two-hybrid, pulldown, co-immunoprecipitation, in vitro kinase assay, fluorimetric Arf-GTPase assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — in vitro kinase assay with mapped phosphorylation sites and direct measurement of GAP activity inhibition; single lab with multiple orthogonal methods","pmids":["12771146"],"is_preprint":false},{"year":2004,"finding":"ASAP1 interacts with the N-terminus (amino acids 2–17) of Arf1 at an interface distinct from other Arf GAPs (AGAP1, ArfGAP1). Specific mutations in Arf1 alpha-helix 3 and switch regions (notably I46D reducing ASAP1-catalyzed hydrolysis ~10,000-fold with isolated effect on kcat) distinguish ASAP1's catalytic interface from other Arf GAPs.","method":"In vitro GAP activity assay, Arf1 mutagenesis, antibody epitope sequestration, in vivo localization studies","journal":"Cellular signalling","confidence":"High","confidence_rationale":"Tier 1 / Moderate — quantitative kinetic analysis with multiple point mutants and in vivo validation","pmids":["15212764"],"is_preprint":false},{"year":2005,"finding":"ASAP1 regulation by phospholipids requires a direct interaction between its PH and Arf GAP domains; the two domains form a composite substrate-binding site. Saturation kinetics, limited proteolysis, FRET, and fluorescence spectrometry support a two-step model: conformational change upon membrane recruitment followed by a second change upon PIP2 binding.","method":"Saturation kinetics, limited proteolysis, FRET, fluorescence spectrometry, analytical ultracentrifugation","journal":"Cellular signalling","confidence":"High","confidence_rationale":"Tier 1 / Moderate — multiple orthogonal biophysical methods in vitro characterizing domain interaction and conformational mechanism","pmids":["16038802"],"is_preprint":false},{"year":2005,"finding":"CD2AP (CD2-associated protein) binds ASAP1 through its N-terminal SH3 domains. Sequestration of endogenous ASAP1 to mitochondria via a CD2AP SH3-mito fusion protein inhibited cell spreading and migration in response to fibronectin and caused increased GTP loading on Arf1 and loss of paxillin from adhesions. siRNA knockdown of ASAP1 produced the same phenotypes.","method":"Affinity chromatography/mass spectrometry, co-immunoprecipitation, GST pulldown, mitochondria mislocalization strategy, siRNA knockdown, GTP-loading assay, fluorescence microscopy","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct identification of binding partner, two independent loss-of-function strategies showing same cellular phenotype and downstream Arf1 GTP loading","pmids":["15632162"],"is_preprint":false},{"year":2005,"finding":"Arf1 mutant I46D selectively abolishes ASAP1-catalyzed GTP hydrolysis (~10,000-fold reduction in kcat) while having minimal effect on AGAP1 (~3-fold). In vivo, [I46D]Arf1 acts as a constitutively active mutant at the cell periphery, disrupting ASAP1 and paxillin localization.","method":"In vitro GAP assay with kinetic analysis, in vivo localization by fluorescence microscopy, mutagenesis","journal":"Current biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — quantitative kinetics with mutagenesis and in vivo functional validation; single lab","pmids":["16332543"],"is_preprint":false},{"year":2006,"finding":"ASAP1 contains an N-terminal BAR domain that together with the PH domain dimerizes into an extended structure that binds acidic phospholipid-containing vesicles and bends membranes to form tubular structures. This bending activity is regulated by Arf1·GTP binding to the Arf GAP domain (acting as an Arf effector). ASAP1 colocalizes with EGFR in tubular recycling structures; the BAR domain is necessary for ASAP1 function in EGFR trafficking and cell spreading.","method":"In vitro membrane tubulation assay with electron microscopy, vesicle sedimentation, dimerization analysis, live-cell imaging, EGFR trafficking assay, BAR domain deletion mutant","journal":"Current biology","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — in vitro reconstitution of membrane bending, EM, and cell-based trafficking/spreading assays with domain deletion mutant","pmids":["16431365"],"is_preprint":false},{"year":2007,"finding":"ASAP1 uses Arf1-GTP as substrate with kcat ~57 s−1 and Km ~2.2 μM (steady-state). AlF4− stabilizes an Arf1-GDP·ASAP1 transition-state complex. Arg-497 mutation severely affects kcat with minimal effect on Km. Mutations of residues predicted to affect Arf1 affinity (W479, I490, R505, L511, D512) instead primarily affected kcat, supporting a conformational change in the Arf1-GTP·ASAP1 complex during catalysis.","method":"Steady-state and single-turnover kinetics, AlF4− trapping, mutagenesis, in vivo dorsal ruffle assay","journal":"The Biochemical journal","confidence":"High","confidence_rationale":"Tier 1 / Moderate — rigorous quantitative kinetics with mutagenesis and transition-state trapping; single lab with multiple methods","pmids":["17112341"],"is_preprint":false},{"year":2007,"finding":"ASAP1 is required for formation of invadopodia and podosomes. The BAR domain, SH3 domain, and Src phosphorylation site of ASAP1 are each required for podosome formation. The Src binding site and GAP activity are dispensable for podosome formation, suggesting ASAP1 functions as a coincidence detector integrating SH3-domain protein interactions, BAR domain scaffolding, and Src phosphorylation.","method":"siRNA knockdown, rescue with ASAP1 mutants (BAR-deleted, SH3-deleted, phosphorylation site mutant, GAP-inactive), fluorescence microscopy","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — systematic domain mutagenesis with siRNA rescue defining necessary structural elements for a specific cellular phenotype","pmids":["17893324"],"is_preprint":false},{"year":2008,"finding":"ASAP1 directly binds FIP3 (a Rab11 and Arf6 effector) through its BAR domain as identified by yeast two-hybrid and confirmed by co-immunoprecipitation and in vitro pulldown. FIP3 binding to the BAR domain stimulates ASAP1 GAP activity against Arf1 but not Arf6. ASAP1 forms a ternary complex with Rab11 via FIP3. ASAP1 colocalizes with FIP3 in pericentrosomal recycling endosomes; ASAP1 or FIP3 depletion alters transferrin receptor localization and transferrin trafficking.","method":"Yeast two-hybrid, co-immunoprecipitation, in vitro pulldown, GAP activity assay, siRNA knockdown, fluorescence microscopy, transferrin trafficking assay","journal":"Molecular biology of the cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal binding assays, functional GAP stimulation assay, and localization with trafficking phenotype","pmids":["18685082"],"is_preprint":false},{"year":2008,"finding":"The BAR domain of ASAP1 autoinhibits GAP activity by intramolecular interaction with the PH and/or Arf GAP domains. The catalytic power of PZA (PH+GAP+Ank) is greater than BAR-PZA; the BAR domain increases Km and decreases kcat. The effect requires the N-terminal loop of the BAR domain and is not due to differential membrane association or membrane curvature changes.","method":"Sedimentation velocity analytical ultracentrifugation, in vitro GAP assay on large unilamellar vesicles, steady-state and single-turnover kinetics, BAR loop mutagenesis","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — quantitative kinetic analysis with domain deletion and mutagenesis demonstrating autoinhibitory mechanism in vitro","pmids":["19017632"],"is_preprint":false},{"year":2008,"finding":"NMR spectroscopy reveals a dynamic interaction between the PH and Arf GAP domains of ASAP1: the domains interact transiently, with the interaction partially occluding the PIP2 binding site in solution. PIP2 binding alters PH domain conformation but has little effect on Arf GAP domain structure. PH domain loop mutations at the GAP-domain interface affect PIP2 binding and both Km and kcat for Arf1 GTP hydrolysis.","method":"NMR spectroscopy, in vitro GAP kinetics, mutagenesis, analytical ultracentrifugation, lipid binding assays","journal":"Cellular signalling","confidence":"High","confidence_rationale":"Tier 1 / Moderate — NMR structural data combined with quantitative kinetics and mutagenesis; single lab with multiple orthogonal methods","pmids":["18675341"],"is_preprint":false},{"year":2011,"finding":"GEFH1 (a RhoA guanine nucleotide exchange factor) binds the BAR domain of ASAP1, colocalizes with ASAP1 in podosomes, inhibits ASAP1 GAP activity, and negatively regulates podosome assembly. GEFH1 overexpression inhibited podosome assembly and a GEFH1 mutant lacking the BAR-binding domain was less effective; GEFH1 siRNA knockdown increased the rate of podosome assembly.","method":"Yeast two-hybrid, co-immunoprecipitation, fluorescence colocalization, GAP activity assay, overexpression, siRNA knockdown","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP confirming endogenous interaction, functional assays showing GAP inhibition and podosome regulation; single lab","pmids":["21352810"],"is_preprint":false},{"year":2012,"finding":"ASAP1 is a scaffold for ciliary receptor targeting that brings together Arf4, Rab11, Rab8-GEF Rabin8, and rhodopsin. Ablation of ASAP1 abolishes ciliary targeting of rhodopsin and causes actin-rich periciliary membrane projections with mislocalized rhodopsin. ASAP1 recognizes the FR ciliary targeting signal of rhodopsin; rhodopsin FR-AA mutant fails to interact with Rab8 and cannot cross the periciliary diffusion barrier.","method":"siRNA knockdown, fluorescence microscopy, co-immunoprecipitation, transport assay in photoreceptors","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — loss-of-function with defined trafficking phenotype, identification of binding signal on cargo, multiple interaction partners confirmed","pmids":["22983554"],"is_preprint":false},{"year":2015,"finding":"Crystal structure of the ASAP1 PH domain was solved in unliganded and dibutyryl-PIP2-bound forms. PIP2 contacts both a canonical site (C site) and an atypical site (A site); PIP2 dependence of vesicle binding and GAP activity is sigmoidal (cooperative), distinct from the hyperbolic binding seen for PLC-δ1 PH domain. Mutations in either the C or A site reduced PIP2-dependent vesicle binding and GAP activity, supporting cooperative binding mechanism for rapid switching.","method":"X-ray crystallography, vesicle binding assay, in vitro GAP activity assay, site-directed mutagenesis","journal":"Structure","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure with functional validation by mutagenesis and cooperative binding analysis","pmids":["26365802"],"is_preprint":false},{"year":2015,"finding":"ASAP1-depleted dendritic cells show impaired matrix degradation and migration. ASAP1 is involved in actin and membrane remodeling associated with podosomes in dendritic cells; genetic variants reducing ASAP1 expression in M. tuberculosis-infected dendritic cells may impair their migration.","method":"siRNA knockdown of ASAP1 in primary human dendritic cells, matrix degradation assay, migration assay","journal":"Nature genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct loss-of-function in primary cells with defined functional phenotype; single study in DCs","pmids":["25774636"],"is_preprint":false},{"year":2015,"finding":"FIP3 competes with rhodopsin for binding to ASAP1 and displaces it from the ternary complex with Arf4-GTP and ASAP1. FIP3 promotes Rab11a activity and coordinates ASAP1 and Rab11a interactions with Rabin8, facilitating orderly Rab11-Rabin8-Rab8 cascade assembly for ciliary receptor trafficking. Ablation of FIP3 abolishes ciliary targeting similarly to ASAP1 ablation.","method":"Co-immunoprecipitation, competition binding assay, siRNA knockdown, fluorescence microscopy, trafficking assay","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP competition and loss-of-function in single lab; extends previous ciliary trafficking mechanism","pmids":["25673879"],"is_preprint":false},{"year":2016,"finding":"The ASAP1 BAR domain together with the PH domain directly binds nonmuscle myosin 2A (NM2A) in vitro. ASAP1 and NM2A co-immunoprecipitate and colocalize in cells. ASAP1 knockdown reduced colocalization of NM2A and F-actin. Knockdown of either ASAP1 or NM2A produced similar defects in focal adhesions, cell migration, spreading, and circular dorsal ruffles. Exogenous NM2A rescued ASAP1-knockdown CDR defects, positioning ASAP1 as a positive regulator of NM2A.","method":"In vitro binding assay, co-immunoprecipitation, siRNA knockdown, rescue experiments, fluorescence microscopy","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Moderate — direct in vitro binding, reciprocal Co-IP, and epistasis by rescue experiments; single lab, multiple orthogonal methods","pmids":["26893376"],"is_preprint":false},{"year":2019,"finding":"PIP2 controls binding of the N-terminal extension (residues 2–17) of ARF1 to the PH domain of ASAP1 and thereby regulates GAP activity. Deletion of the ARF1 N-terminus ([Δ17]ARF1) makes GAP activity largely PIP2-independent. A peptide of residues 2–17 inhibits GAP activity and binds PIP2-dependently to the PH domain including a 17-amino acid interdomain linker N-terminal to the first β-strand. Mutations in the linker or C-terminal α-helix of the PH domain decrease both ARF1 N-terminal binding and GAP activity, and reduce cellular actin remodeling.","method":"In vitro GAP assay with truncation and point mutants, peptide inhibition assay, NMR binding assay, cell-based actin remodeling assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — quantitative in vitro kinetics with mutagenesis, NMR binding validation, and cell-based functional assay; single lab, multiple orthogonal methods","pmids":["31591270"],"is_preprint":false},{"year":2019,"finding":"ASAP1 depletion causes defects in actin stress fiber organization. The BAR-PH fragment is sufficient to affect actin; the N-BAR domain of ASAP1 directly binds and bundles actin filaments in vitro, whereas the Arf GAP and C-terminal SH3 domain reduce BAR-PH binding and bundling. Overexpression of ASAP1 enhanced actin remodeling; replacing the ASAP1 BAR domain with the ACAP1 BAR domain abolished actin effects.","method":"siRNA knockdown, overexpression, actin co-sedimentation, domain swap mutants, fluorescence microscopy","journal":"iScience","confidence":"High","confidence_rationale":"Tier 2 / Moderate — direct in vitro actin binding and bundling assays combined with domain mutants and cell-based phenotype; single lab","pmids":["31785555"],"is_preprint":false},{"year":2019,"finding":"NMR methyl-geoHARD analysis of the ASAP1 ZA (PH-ArfGAP) domain reveals wide-range conformational dynamics (kex 10²–10⁵ s⁻¹) in the hydrophobic interior, including collective and local motions that may correlate with catalytic function and substrate recognition.","method":"Methyl-TROSY NMR, adiabatic relaxation dispersion, CPMG relaxation dispersion","journal":"Journal of the American Chemical Society","confidence":"Low","confidence_rationale":"Tier 1 / Weak — rigorous NMR technique but functional correlation remains speculative; single lab, dynamics-only study","pmids":["31293161"],"is_preprint":false},{"year":2019,"finding":"Loss of ASAP1 in mice (gene-trap) impairs adipogenic and osteogenic differentiation of mesenchymal progenitor cells, causing growth retardation and delayed ossification. Mechanistically, FAK/Src and PI3K/AKT signaling is compromised in ASAP1-null MEFs, leading to impaired adipogenic and osteogenic differentiation.","method":"Gene-trap mouse model, in vitro differentiation assays, Western blotting for FAK/Src and PI3K/AKT pathway components","journal":"PLoS genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic loss-of-function with defined differentiation phenotype and pathway readout; single lab","pmids":["31246957"],"is_preprint":false},{"year":2020,"finding":"Binding of multiple PI(4,5)P2 molecules to the ASAP1 PH domain triggers a functionally relevant allosteric conformational switch and maintains the PH domain in a defined orientation that allows critical contacts with Arf1 at the membrane, as determined by combining NMR, neutron reflectometry, and molecular dynamics simulation.","method":"NMR, neutron reflectometry, molecular dynamics simulation","journal":"Science advances","confidence":"High","confidence_rationale":"Tier 1 / Moderate — three complementary structural/biophysical methods providing atomic-level mechanism; single integrated study","pmids":["32998886"],"is_preprint":false},{"year":2020,"finding":"The ASAP1 N-BAR domain directly binds F-actin, homodimerization of ASAP1 aligns F-actin in predominantly unipolar bundles, and ASAP1 stabilizes actin filaments against depolymerization. The N-BAR domain moderately reduces spontaneous G-actin polymerization. Overexpression of ASAP1 BAR-PH tandem induced actin-filled cellular projections; an ASAP1 construct lacking the N-BAR domain failed to induce projections.","method":"Actin cosedimentation, polymerization and depolymerization assays, TIRF microscopy, confocal microscopy, electron microscopy, overexpression in fibroblasts","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — direct in vitro binding reconstitution with multiple structural and cell-based assays; multiple orthogonal methods","pmids":["32444496"],"is_preprint":false},{"year":2021,"finding":"Knockdown of ASAP1 in THP1-derived macrophages increased efficiency of Mycobacterium tuberculosis H37Ra entry and enhanced F-actin aggregation and vinculin/paxillin-rich puncta formation, identifying ASAP1 as a regulator of Mtb uptake through actin cytoskeleton remodeling.","method":"siRNA knockdown, fluorescence confocal microscopy, colony forming unit assay, F-actin staining","journal":"Tuberculosis","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function with defined entry and actin phenotype; single lab, single method type","pmids":["34058694"],"is_preprint":false},{"year":2022,"finding":"A lysine-rich cluster (K75, K76, K79) in the N-BAR domain of ASAP1 is required for binding and bundling actin filaments. Charge-neutralizing or charge-reversing mutations at these positions reduced BAR-PH binding to F-actin and abrogated actin bundle formation in vitro and cellular actin remodeling in U2OS cells; [K75E, K76E, K79E] full-length ASAP1 did not rescue endogenous ASAP1 knockdown-induced reduction of stress fibers.","method":"Structural modeling, mutagenesis, actin co-sedimentation, in vitro bundling assay, cell-based actin remodeling assay, siRNA rescue","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — mutagenesis with in vitro reconstitution and cell-based rescue assay; single lab, multiple orthogonal methods","pmids":["35143843"],"is_preprint":false},{"year":2023,"finding":"Membrane-bound active Arf1 (Myr-Arf1) explores large conformational dynamics with its G domain oscillating between membrane-associated and membrane-distal conformations. Interaction with the ASAP1 PH domain restricts Arf1 G domain motions and locks it in a conformation exposing functionally relevant regions for catalysis.","method":"NMR, neutron reflectometry, molecular dynamics simulations","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1 / Moderate — three complementary structural/biophysical methods; mechanistic insight into ASAP1–Arf1 interaction at membrane","pmids":["37989735"],"is_preprint":false},{"year":2023,"finding":"Crystal structure of the ASAP1 SH3 domain in complex with the MICAL1 proline-rich motif (PRM) revealed a unique binding mode: ASAP1 SH3 contains two negatively charged patches that recognize the 'xPx+Px+' sequence in MICAL1 PRM, yielding sub-μM affinity. This binding pocket (termed SH3AGS) is also found in GRAF and SKAP1 SH3 domains.","method":"X-ray crystallography, ITC/binding affinity measurement, mutagenesis","journal":"International journal of molecular sciences","confidence":"High","confidence_rationale":"Tier 1 / Moderate — crystal structure with biochemical binding validation; single lab","pmids":["36674928"],"is_preprint":false},{"year":2023,"finding":"ASAP1 acts as an effector for ARF6 and mediates HGF/IGF-1 signaling to promote nuclear localization and transcriptional activity of NFAT1 in uveal melanoma. HGF and IGF-1 hyperactivate ARF6, which then interacts with ASAP1 to induce NFAT1 nuclear translocation; inhibition of ASAP1 or NFAT impairs cellular invasiveness and reduces metastasis in a xenograft model.","method":"Co-immunoprecipitation (ARF6-ASAP1), NFAT nuclear localization assay, siRNA knockdown, xenograft mouse model","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP for ARF6-ASAP1 interaction, functional loss-of-function with defined signaling phenotype and in vivo validation; single lab","pmids":["37500798"],"is_preprint":false},{"year":2023,"finding":"ASAP1 activates the IQGAP1/CDC42 pathway by inhibiting ubiquitin-mediated degradation of IQGAP1, thereby enhancing CDC42 activity. Activated CDC42 upregulates the EGFR-MAPK pathway to promote chemotherapy resistance in gastric cancer.","method":"siRNA/overexpression, Co-IP, ubiquitination assay, Western blotting for CDC42/EGFR/MAPK pathway, in vitro and in vivo tumor assays","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP and ubiquitination assay identifying mechanism; single lab with multiple assays","pmids":["36792578"],"is_preprint":false},{"year":2024,"finding":"CUL1 promotes ubiquitination and degradation of ASAP1 via the SCF-FBXW7 complex, suppressing osteoblast proliferation and osteogenesis. CUL1 silencing moderated Dex-induced inhibition of proliferation and osteogenesis by restoring ASAP1 levels.","method":"Co-immunoprecipitation, ubiquitination assay, CUL1 siRNA knockdown, osteogenesis assay, mouse osteoporosis model","journal":"Hormones","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP and ubiquitination assay identifying the E3 complex; single lab with in vivo validation","pmids":["39287759"],"is_preprint":false},{"year":2024,"finding":"The SH3 domain of ASAP1 binds a 12-residue positively charged peptide from the neuronal scaffold protein 440 kDa ankyrin-B via a noncanonical SH3-ligand binding mode. The crystal structure of ASAP1-SH3 in complex with this gAnkB peptide defined a consensus ASAP1-SH3 binding motif, enabling identification of novel binding partners including Clasp1/2, ALS2, β-Pix, DAPK3, PHIP, and Limk1.","method":"Crystal structure determination, ITC/binding affinity measurement, mutagenesis, in silico database search","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — crystal structure with quantitative binding and mutagenesis; single lab with multiple orthogonal methods","pmids":["39265663"],"is_preprint":false},{"year":2024,"finding":"MSUT2 regulates tau seed internalization into neurons via adenosine receptor 1 (A1AR)-mediated modulation of ASAP1 activity. Down-regulation or inhibition of A1AR modulates ASAP1 activity, reducing internalization of pathogenic tau seeds and tau pathology in neuron cultures and mouse models.","method":"siRNA knockdown, tau seeding assay, A1AR inhibitor treatment, neuron culture and mouse model of tau pathology","journal":"Acta neuropathologica","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function genetics with defined tau seeding phenotype and pathway placement; single lab","pmids":["38472475"],"is_preprint":false},{"year":2025,"finding":"ASAP1 and ARF1 are necessary for myogenic differentiation in FN-RMS. Loss of ASAP1 or ARF1/ARF5 (GAP substrates) blocks differentiation and prevents MEK-inhibition-induced inactivation of TAZ (WWTR1), a pro-proliferative transcriptional co-activator. Dual knockdown of ASAP1 and WWTR1 rescued MEKi-induced differentiation, placing ASAP1 upstream of TAZ inactivation.","method":"siRNA knockdown, MEK inhibitor treatment, Western blotting for TAZ phosphorylation/activation, myogenic transcription factor expression assay, epistasis rescue experiment","journal":"Molecular cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — epistasis experiments placing ASAP1 upstream of TAZ in differentiation pathway; single lab with multiple knockdown conditions","pmids":["39495123"],"is_preprint":false},{"year":2025,"finding":"MYO1F interacts with ASAP1 through an SH3-domain-dependent interaction (proximity labeling proteomics, structural modeling, mutagenesis), and ASAP1 colocalizes with MYO1F at actin-rich podosomes and phagocytic cups in macrophages and microglia.","method":"Proximity labeling/proteomics, structural modeling, mutagenesis, immunofluorescence colocalization","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — proximity proteomics with structural and mutagenesis validation; functional consequence not directly tested for ASAP1 specifically","pmids":["41208482"],"is_preprint":false},{"year":2025,"finding":"ASAP1 interacts with the SMAD2/3 complex and forms a positive feedback loop with TGFβ signaling, promoting EMT and cell invasiveness in papillary thyroid cancer cells. ASAP1 knockdown reduced p-SMAD2 levels; co-immunoprecipitation confirmed ASAP1-SMAD2/3 interaction.","method":"Co-immunoprecipitation, immunofluorescence, Western blotting for p-SMAD2, luciferase reporter assay, lentiviral knockdown/overexpression","journal":"Cancer medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP confirming physical interaction and TGFβ pathway assays; single lab","pmids":["40742091"],"is_preprint":false},{"year":2025,"finding":"Both the Arf GAP domain activity and the BAR domain actin/NM2A-binding activity of ASAP1 are required coordinately to maintain focal adhesions and actin stress fibers; neither domain alone is sufficient. Arf5 (a GAP substrate) loss-of-function phenocopies ASAP1 knockdown on SFs and FAs.","method":"siRNA knockdown, rescue with domain mutants (GAP-inactive, BAR-deleted), dominant negative and GTPase-deficient Arf5 mutants, fluorescence microscopy in four cell lines","journal":"Biology of the cell","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — systematic domain rescue in multiple cell lines with epistatic Arf5 experiments; single lab","pmids":["40194952"],"is_preprint":false},{"year":2024,"finding":"The PH domain of ASAP1 enhances GAP activity by >7 orders of magnitude by acting as an active catalytic component, not merely a membrane recruitment signal. NMR and MD simulations show the PH domain directly contacts Arf·GTP at the membrane and allosterically drives conformational rearrangements of the GTP binding site to facilitate charge stabilization and accelerate GTP hydrolysis; mathematical modeling indicates this allosteric contribution equals membrane recruitment in importance.","method":"NMR, molecular dynamics simulation, kinetic assays, mutagenesis, mathematical modeling","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — rigorous structural and kinetic study with mutagenesis; preprint, not yet peer-reviewed","pmids":["39763923"],"is_preprint":true}],"current_model":"ASAP1 is a multi-domain scaffold protein that acts as a PIP2-dependent Arf GTPase-activating protein (primarily for Arf1/Arf5), in which the PH domain allosterically activates catalysis by directly engaging Arf·GTP at the membrane surface, the BAR domain autoinhibits GAP activity intramolecularly and independently bundles actin filaments via a lysine-rich cluster, and the SH3 domain mediates interactions with Src, FAK, Pyk2, CrkL, and other proline-rich partners; Src and Pyk2 phosphorylate ASAP1 to modulate its GAP activity and localization, while ASAP1 coordinates focal adhesion dynamics, invadopodium/podosome formation, circular dorsal ruffle generation, endocytic recycling, and ciliary receptor trafficking by integrating signals from phospholipids, Arf GTPases, Rab proteins, and the actomyosin cytoskeleton."},"narrative":{"mechanistic_narrative":"ASAP1 is a multi-domain scaffold and PIP2-dependent Arf GTPase-activating protein that integrates phospholipid signals, Arf GTPases, and the actomyosin cytoskeleton to control focal adhesion dynamics, invadopodium/podosome formation, dorsal ruffle generation, endocytic recycling, and ciliary receptor trafficking [PMID:9819391, PMID:10725410, PMID:18685082, PMID:22983554]. It was originally purified as a PIP2-dependent GAP acting on Arf1 and Arf5 but not Arf6, with catalysis requiring the PH domain in concert with the Arf GAP domain [PMID:9819391, PMID:11773070, PMID:15212764]. PIP2 binding to the PH domain is not merely a recruitment cue but an allosteric activator: cooperative binding at canonical and atypical sites drives a conformational switch that orients the PH domain to engage Arf·GTP at the membrane and accelerate hydrolysis, restricting the conformational motions of membrane-bound Arf1 to expose its catalytic regions [PMID:10734117, PMID:26365802, PMID:32998886, PMID:37989735]. The N-terminal BAR domain serves dual roles—it autoinhibits GAP activity through intramolecular contacts with the PH/GAP domains, and it independently dimerizes to bend membranes, bind and bundle actin filaments via a lysine-rich cluster (K75/K76/K79), and bind nonmuscle myosin 2A, thereby coupling membrane remodeling to actin organization [PMID:16431365, PMID:19017632, PMID:26893376, PMID:31785555, PMID:32444496, PMID:35143843]. GAP activity and BAR-domain actin/NM2A binding act coordinately to maintain focal adhesions and stress fibers, with Arf5 loss phenocopying ASAP1 depletion [PMID:40194952]. The C-terminal SH3 domain, through a noncanonical acidic binding pocket, mediates interactions with proline-rich partners including FAK, Pyk2, and MICAL1, and Pyk2 phosphorylates ASAP1 to inhibit its GAP activity [PMID:12058076, PMID:12771146, PMID:36674928, PMID:39265663]. ASAP1 is recruited to focal adhesions by CrkL and CD2AP and nucleates a ciliary trafficking complex with Arf4, Rab11, FIP3, and Rabin8 that targets rhodopsin to the cilium [PMID:12522101, PMID:15632162, PMID:18685082, PMID:22983554, PMID:25673879]. In disease contexts, ASAP1 functions as an Arf6 effector promoting NFAT1-driven invasion in uveal melanoma and participates in TGFβ/SMAD and IQGAP1/CDC42 signaling in cancers [PMID:37500798, PMID:36792578, PMID:40742091].","teleology":[{"year":1998,"claim":"Established ASAP1's defining biochemical activity—answering whether it was an enzyme by showing it is a PIP2-dependent Arf GAP physically coupled to Src-family signaling.","evidence":"Protein purification, in vitro GAP assay, GST pulldown, and in-cell phosphorylation","pmids":["9819391"],"confidence":"High","gaps":["Did not resolve how PIP2 stimulates catalysis","Substrate specificity among Arf isoforms not yet defined"]},{"year":2000,"claim":"Linked the enzymatic activity to cellular function by localizing ASAP1 to focal adhesions and showing GAP activity is required for cell spreading and dorsal ruffle control.","evidence":"Fluorescence microscopy and cell spreading/PDGF assays with wild-type vs catalytically inactive mutant","pmids":["10725410"],"confidence":"High","gaps":["GAP-independent functions hinted but not mechanistically separated"]},{"year":2000,"claim":"Resolved the role of PIP2 by showing the PH domain is an allosteric activator distinct from a membrane recruitment signal.","evidence":"In vitro GAP assays, limited proteolysis, and domain deletion analysis","pmids":["10734117"],"confidence":"High","gaps":["Atomic structure of the conformational change unresolved"]},{"year":2001,"claim":"Defined in vivo substrate specificity (Arf1 not Arf6) and dissected two independent downstream pathways—GAP-dependent motility versus GAP-independent spreading inhibition.","evidence":"Cell-based ARF GAP assay, stable overexpression, motility assay, GAP-domain deletion mutant","pmids":["11773070"],"confidence":"High","gaps":["Effectors of the GAP-independent spreading pathway not identified"]},{"year":2002,"claim":"Identified FAK as a direct SH3-domain partner linking ASAP1 to focal adhesion assembly.","evidence":"Yeast two-hybrid, reciprocal Co-IP/pulldown, cell spreading assay with binding/GAP mutants","pmids":["12058076"],"confidence":"High","gaps":["Whether FAK binding regulates GAP activity not tested"]},{"year":2003,"claim":"Established phospho-regulation and adapter recruitment—Pyk2 phosphorylates ASAP1 to inhibit GAP activity, and CrkL directs it to peripheral adhesions.","evidence":"Yeast two-hybrid, Co-IP, in vitro kinase assay with mapped sites, fluorimetric GTPase assay; CrkL recruitment with SH2-mutant","pmids":["12771146","12522101"],"confidence":"Medium","gaps":["Single-lab findings","Functional consequence of localization on Arf signaling not quantified"]},{"year":2005,"claim":"Defined the catalytic mechanism as a two-step conformational model with a composite PH–GAP substrate-binding site, and validated cellular function via CD2AP-dependent recruitment.","evidence":"Saturation kinetics, FRET, limited proteolysis, AUC; CD2AP mislocalization and siRNA with Arf1 GTP-loading readout","pmids":["16038802","15632162"],"confidence":"High","gaps":["Atomic interface between PH and GAP domains not yet defined"]},{"year":2006,"claim":"Discovered the BAR domain as a membrane-bending module regulated by Arf1·GTP, expanding ASAP1's role into endocytic recycling of EGFR.","evidence":"In vitro membrane tubulation with EM, dimerization analysis, EGFR trafficking assay with BAR deletion","pmids":["16431365"],"confidence":"High","gaps":["In vivo significance of tubulation for recycling not fully established"]},{"year":2007,"claim":"Provided rigorous catalytic kinetics and transition-state trapping, and defined ASAP1 as a coincidence detector for invadopodium/podosome formation requiring BAR, SH3, and Src phosphorylation but not GAP activity.","evidence":"Steady-state/single-turnover kinetics with AlF4- trapping; siRNA rescue with systematic domain mutants","pmids":["17112341","17893324"],"confidence":"High","gaps":["How the distinct domain inputs are integrated spatially is unresolved"]},{"year":2008,"claim":"Resolved BAR autoinhibition and dynamic PH–GAP interactions, and connected ASAP1 to Rab11/FIP3-dependent recycling endosome trafficking.","evidence":"AUC, kinetics with BAR loop mutagenesis, NMR; yeast two-hybrid/Co-IP and transferrin trafficking with FIP3","pmids":["19017632","18675341","18685082"],"confidence":"High","gaps":["How BAR autoinhibition is relieved in cells not defined"]},{"year":2011,"claim":"Identified GEFH1 as a BAR-binding negative regulator coupling Rho exchange activity to podosome assembly through ASAP1.","evidence":"Yeast two-hybrid, Co-IP, GAP assay, siRNA/overexpression","pmids":["21352810"],"confidence":"Medium","gaps":["Single lab","Mechanism of GAP inhibition by GEFH1 not defined"]},{"year":2012,"claim":"Established ASAP1 as a scaffold for ciliary receptor targeting, assembling Arf4, Rab11, Rabin8, and rhodopsin and recognizing the rhodopsin ciliary targeting signal.","evidence":"siRNA knockdown, Co-IP, photoreceptor transport assay","pmids":["22983554"],"confidence":"High","gaps":["Order of complex assembly not yet defined"]},{"year":2015,"claim":"Solved the PH domain structure revealing cooperative dual-site PIP2 binding, and showed FIP3 orchestrates the Rab11-Rabin8-Rab8 cascade by competing with rhodopsin for ASAP1.","evidence":"X-ray crystallography with mutagenesis; Co-IP competition and trafficking assays","pmids":["26365802","25673879"],"confidence":"High","gaps":["Single-lab ciliary mechanism","Generality beyond rhodopsin cargo not tested"]},{"year":2015,"claim":"Extended ASAP1 function to immune cell migration and matrix degradation, linking it to dendritic cell motility relevant to tuberculosis susceptibility.","evidence":"siRNA knockdown in primary human dendritic cells, matrix degradation and migration assays","pmids":["25774636"],"confidence":"Medium","gaps":["Single study in DCs","Molecular pathway to migration not dissected"]},{"year":2016,"claim":"Identified nonmuscle myosin 2A as a direct BAR-PH partner, positioning ASAP1 as a positive regulator of NM2A in adhesion, migration, and dorsal ruffle formation.","evidence":"In vitro binding, reciprocal Co-IP, siRNA knockdown with NM2A rescue","pmids":["26893376"],"confidence":"High","gaps":["How ASAP1 modulates NM2A contractility mechanically not defined"]},{"year":2019,"claim":"Defined the BAR domain as a direct actin filament binding and bundling module dependent on a lysine cluster, and established the Arf1 N-terminal extension as the PIP2-dependent regulatory contact controlling GAP activity.","evidence":"Actin cosedimentation, domain-swap and point mutants, cell-based actin remodeling; GAP kinetics, peptide inhibition, NMR binding","pmids":["31785555","31591270"],"confidence":"High","gaps":["Coupling of actin bundling to GAP catalysis in cells not resolved"]},{"year":2019,"claim":"Demonstrated ASAP1's requirement for mesenchymal progenitor differentiation in vivo, linking loss to growth retardation, delayed ossification, and impaired FAK/Src and PI3K/AKT signaling.","evidence":"Gene-trap mouse, in vitro differentiation assays, Western blotting for signaling components","pmids":["31246957"],"confidence":"Medium","gaps":["Single lab","Direct molecular link from ASAP1 to the signaling defects not established"]},{"year":2020,"claim":"Resolved the atomic-level allosteric switch by which cooperative PIP2 binding orients the PH domain for Arf1 contact, and characterized BAR-driven unipolar actin bundling and filament stabilization.","evidence":"NMR, neutron reflectometry, MD simulation; actin cosedimentation, TIRF/EM, cell-based projection assays","pmids":["32998886","32444496"],"confidence":"High","gaps":["Integration of bundling activity with membrane-bound GAP function in vivo unresolved"]},{"year":2021,"claim":"Showed ASAP1 regulates Mycobacterium tuberculosis uptake in macrophages through actin cytoskeleton remodeling.","evidence":"siRNA knockdown, confocal microscopy, CFU assay, F-actin staining","pmids":["34058694"],"confidence":"Medium","gaps":["Single lab","Direct effector linking ASAP1 to uptake not identified"]},{"year":2022,"claim":"Pinpointed the lysine-rich cluster (K75/K76/K79) as the structural determinant for actin binding and bundling required for cellular stress fiber maintenance.","evidence":"Structural modeling, charge mutagenesis, cosedimentation/bundling assays, siRNA rescue in U2OS cells","pmids":["35143843"],"confidence":"High","gaps":["Whether the same residues affect GAP activity not tested"]},{"year":2023,"claim":"Defined SH3-domain ligand recognition modes (MICAL1 PRM and ankyrin-B), establishing a noncanonical acidic binding pocket and expanding the ASAP1 partner network, and showed the PH domain restricts membrane-bound Arf1 conformational dynamics.","evidence":"X-ray crystallography with ITC and mutagenesis; NMR/neutron reflectometry/MD for Arf1 dynamics","pmids":["36674928","39265663","37989735"],"confidence":"High","gaps":["Functional consequence of most newly predicted SH3 partners not validated"]},{"year":2023,"claim":"Placed ASAP1 in oncogenic signaling as an Arf6 effector driving NFAT1-mediated invasion and as a regulator of IQGAP1/CDC42-EGFR-MAPK signaling promoting chemoresistance.","evidence":"Co-IP, NFAT nuclear localization, ubiquitination assays, siRNA/overexpression, xenograft and tumor models","pmids":["37500798","36792578"],"confidence":"Medium","gaps":["Single-lab cancer mechanisms","Direct enzymatic role of ASAP1 in these pathways unclear"]},{"year":2024,"claim":"Established that ASAP1 protein levels are controlled by SCF-FBXW7/CUL1-mediated ubiquitination affecting osteogenesis, and connected ASAP1 to neuronal tau seed internalization via A1AR/MSUT2.","evidence":"Co-IP, ubiquitination assays, osteogenesis and osteoporosis models; tau seeding assays with A1AR modulation","pmids":["39287759","38472475"],"confidence":"Medium","gaps":["Single-lab findings","Mechanism of ASAP1 activity modulation by A1AR not defined"]},{"year":2025,"claim":"Demonstrated coordinate requirement of GAP and BAR-domain activities for adhesion/stress fiber maintenance, identified additional cytoskeletal partners (MYO1F), and placed ASAP1 in differentiation and EMT pathways (TAZ inactivation, TGFβ/SMAD).","evidence":"Domain-rescue siRNA in multiple cell lines with Arf5 epistasis; proximity proteomics; MEKi epistasis with WWTR1; Co-IP with SMAD2/3","pmids":["40194952","41208482","39495123","40742091"],"confidence":"Medium","gaps":["Single-lab pathway placements","Direct molecular links between ASAP1 enzymatic activity and transcriptional outputs unresolved"]},{"year":null,"claim":"How ASAP1's distinct domain activities—GAP catalysis, BAR membrane bending and actin bundling, and SH3 scaffolding—are spatially and temporally coordinated within single structures (focal adhesions, podosomes, cilia) in living cells remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No integrated model of in-cell domain coordination","Physiological triggers relieving BAR autoinhibition unknown","In vivo roles of most cancer/neuronal pathways not validated genetically"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0,3,6,7,12]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[0,6,14,17]},{"term_id":"GO:0008289","term_label":"lipid binding","supporting_discovery_ids":[2,11,19,27]},{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[24,28,30,22]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[5,9,18,36]},{"term_id":"GO:0005198","term_label":"structural molecule activity","supporting_discovery_ids":[11,28]}],"localization":[{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[1,24,28]},{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[1,11,13]},{"term_id":"GO:0005768","term_label":"endosome","supporting_discovery_ids":[11,14]},{"term_id":"GO:0005929","term_label":"cilium","supporting_discovery_ids":[18,21]},{"term_id":"GO:0005815","term_label":"microtubule organizing center","supporting_discovery_ids":[14]}],"pathway":[{"term_id":"R-HSA-5653656","term_label":"Vesicle-mediated transport","supporting_discovery_ids":[11,14,18]},{"term_id":"R-HSA-1474244","term_label":"Extracellular matrix organization","supporting_discovery_ids":[1,4,13,20]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[0,6,33]},{"term_id":"R-HSA-9609507","term_label":"Protein localization","supporting_discovery_ids":[18,21]}],"complexes":["ASAP1-FIP3-Rab11 ternary complex","ciliary targeting complex (Arf4-Rab11-Rabin8-rhodopsin)"],"partners":["FAK","PYK2","CRKL","CD2AP","FIP3","NM2A","MICAL1","ARF1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9ULH1","full_name":"Arf-GAP with SH3 domain, ANK repeat and PH domain-containing protein 1","aliases":["130 kDa phosphatidylinositol 4,5-bisphosphate-dependent ARF1 GTPase-activating protein","ADP-ribosylation factor-directed GTPase-activating protein 1","ARF GTPase-activating protein 1","Development and differentiation-enhancing factor 1","DEF-1","Differentiation-enhancing factor 1","PIP2-dependent ARF1 GAP"],"length_aa":1129,"mass_kda":125.5,"function":"Possesses phosphatidylinositol 4,5-bisphosphate-dependent GTPase-activating protein activity for ARF1 (ADP ribosylation factor 1) and ARF5 and a lesser activity towards ARF6. May coordinate membrane trafficking with cell growth or actin cytoskeleton remodeling by binding to both SRC and PIP2. May function as a signal transduction protein involved in the differentiation of fibroblasts into adipocytes and possibly other cell types. Part of the ciliary targeting complex containing Rab11, ASAP1, Rabin8/RAB3IP, RAB11FIP3 and ARF4, which direct preciliary vesicle trafficking to mother centriole and ciliogenesis initiation (PubMed:25673879)","subcellular_location":"Cytoplasm; Membrane; Golgi apparatus; Golgi apparatus, trans-Golgi network","url":"https://www.uniprot.org/uniprotkb/Q9ULH1/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/ASAP1","classification":"Not Classified","n_dependent_lines":2,"n_total_lines":1208,"dependency_fraction":0.0016556291390728477},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/ASAP1","total_profiled":1310},"omim":[{"mim_id":"617413","title":"PRUNE EXOPOLYPHOSPHATASE 1; PRUNE1","url":"https://www.omim.org/entry/617413"},{"mim_id":"608651","title":"ARF GTPase-ACTIVATING PROTEIN WITH GTPase DOMAIN, ANKYRIN REPEAT, AND PLECKSTRIN HOMOLOGY DOMAIN 1; AGAP1","url":"https://www.omim.org/entry/608651"},{"mim_id":"607766","title":"ARF-GAP WITH COILED-COIL, ANKYRIN REPEAT, AND PLECKSTRIN HOMOLOGY DOMAINS 2; ACAP2","url":"https://www.omim.org/entry/607766"},{"mim_id":"607763","title":"ARF-GAP WITH COILED-COIL, ANKYRIN REPEAT, AND PLECKSTRIN HOMOLOGY DOMAINS 1; ACAP1","url":"https://www.omim.org/entry/607763"},{"mim_id":"605953","title":"ARF GTPase-ACTIVATING PROTEIN WITH SH3 DOMAIN, ANKYRIN REPEAT, AND PH DOMAIN 1; ASAP1","url":"https://www.omim.org/entry/605953"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Golgi apparatus","reliability":"Approved"},{"location":"Plasma membrane","reliability":"Approved"},{"location":"Centriolar satellite","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/ASAP1"},"hgnc":{"alias_symbol":["PAP","KIAA1249","ZG14P","CENTB4"],"prev_symbol":["DDEF1"]},"alphafold":{"accession":"Q9ULH1","domains":[{"cath_id":"1.20.1270.60","chopping":"28-275","consensus_level":"high","plddt":91.7106,"start":28,"end":275},{"cath_id":"2.30.29.30","chopping":"322-425","consensus_level":"high","plddt":87.4535,"start":322,"end":425},{"cath_id":"1.10.220.150","chopping":"431-555","consensus_level":"medium","plddt":93.5711,"start":431,"end":555},{"cath_id":"1.25.40.20","chopping":"574-710","consensus_level":"medium","plddt":91.5899,"start":574,"end":710},{"cath_id":"2.30.30.40","chopping":"1072-1128","consensus_level":"high","plddt":90.7461,"start":1072,"end":1128}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9ULH1","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9ULH1-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9ULH1-F1-predicted_aligned_error_v6.png","plddt_mean":71.31},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=ASAP1","jax_strain_url":"https://www.jax.org/strain/search?query=ASAP1"},"sequence":{"accession":"Q9ULH1","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9ULH1.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9ULH1/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9ULH1"}},"corpus_meta":[{"pmid":"9819391","id":"PMC_9819391","title":"ASAP1, a phospholipid-dependent arf GTPase-activating protein that associates with and is phosphorylated by Src.","date":"1998","source":"Molecular and cellular biology","url":"https://pubmed.ncbi.nlm.nih.gov/9819391","citation_count":212,"is_preprint":false},{"pmid":"10725410","id":"PMC_10725410","title":"The Arf GTPase-activating protein ASAP1 regulates the actin cytoskeleton.","date":"2000","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/10725410","citation_count":182,"is_preprint":false},{"pmid":"32926734","id":"PMC_32926734","title":"EIF4A3-induced circular RNA ASAP1 promotes tumorigenesis and temozolomide resistance of glioblastoma via NRAS/MEK1/ERK1-2 signaling.","date":"2021","source":"Neuro-oncology","url":"https://pubmed.ncbi.nlm.nih.gov/32926734","citation_count":170,"is_preprint":false},{"pmid":"12058076","id":"PMC_12058076","title":"The association of ASAP1, an ADP ribosylation factor-GTPase activating protein, with focal adhesion kinase contributes to the process of focal adhesion assembly.","date":"2002","source":"Molecular biology of the cell","url":"https://pubmed.ncbi.nlm.nih.gov/12058076","citation_count":142,"is_preprint":false},{"pmid":"25774636","id":"PMC_25774636","title":"Susceptibility to tuberculosis is associated with variants in the ASAP1 gene encoding a regulator of dendritic cell migration.","date":"2015","source":"Nature genetics","url":"https://pubmed.ncbi.nlm.nih.gov/25774636","citation_count":138,"is_preprint":false},{"pmid":"10734117","id":"PMC_10734117","title":"Phosphoinositide-dependent activation of the ADP-ribosylation factor GTPase-activating protein ASAP1. Evidence for the pleckstrin homology domain functioning as an allosteric site.","date":"2000","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/10734117","citation_count":109,"is_preprint":false},{"pmid":"15897555","id":"PMC_15897555","title":"DDEF1 is located in an amplified region of chromosome 8q and is overexpressed in uveal melanoma.","date":"2005","source":"Clinical cancer research : an official journal of the American Association for Cancer Research","url":"https://pubmed.ncbi.nlm.nih.gov/15897555","citation_count":108,"is_preprint":false},{"pmid":"22983554","id":"PMC_22983554","title":"The Arf GAP ASAP1 provides a platform to regulate Arf4- and Rab11-Rab8-mediated ciliary receptor targeting.","date":"2012","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/22983554","citation_count":107,"is_preprint":false},{"pmid":"17893324","id":"PMC_17893324","title":"Src-dependent phosphorylation of ASAP1 regulates podosomes.","date":"2007","source":"Molecular and cellular biology","url":"https://pubmed.ncbi.nlm.nih.gov/17893324","citation_count":88,"is_preprint":false},{"pmid":"11773070","id":"PMC_11773070","title":"DEF-1/ASAP1 is a GTPase-activating protein (GAP) for ARF1 that enhances cell motility through a GAP-dependent mechanism.","date":"2001","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/11773070","citation_count":81,"is_preprint":false},{"pmid":"18519696","id":"PMC_18519696","title":"ASAP1, a gene at 8q24, is associated with prostate cancer metastasis.","date":"2008","source":"Cancer research","url":"https://pubmed.ncbi.nlm.nih.gov/18519696","citation_count":78,"is_preprint":false},{"pmid":"16431365","id":"PMC_16431365","title":"A BAR domain in the N terminus of the Arf GAP ASAP1 affects membrane structure and trafficking of epidermal growth factor receptor.","date":"2006","source":"Current biology : CB","url":"https://pubmed.ncbi.nlm.nih.gov/16431365","citation_count":78,"is_preprint":false},{"pmid":"18685082","id":"PMC_18685082","title":"Arf GTPase-activating protein ASAP1 interacts with Rab11 effector FIP3 and regulates pericentrosomal localization of transferrin receptor-positive recycling endosome.","date":"2008","source":"Molecular biology of the cell","url":"https://pubmed.ncbi.nlm.nih.gov/18685082","citation_count":65,"is_preprint":false},{"pmid":"19017632","id":"PMC_19017632","title":"Autoinhibition of Arf GTPase-activating protein activity by the BAR domain in ASAP1.","date":"2008","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/19017632","citation_count":64,"is_preprint":false},{"pmid":"15632162","id":"PMC_15632162","title":"Mislocalization or reduced expression of Arf GTPase-activating protein ASAP1 inhibits cell spreading and migration by influencing Arf1 GTPase cycling.","date":"2005","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/15632162","citation_count":62,"is_preprint":false},{"pmid":"26365802","id":"PMC_26365802","title":"Molecular Basis for Cooperative Binding of Anionic Phospholipids to the PH Domain of the Arf GAP ASAP1.","date":"2015","source":"Structure (London, England : 1993)","url":"https://pubmed.ncbi.nlm.nih.gov/26365802","citation_count":56,"is_preprint":false},{"pmid":"27667152","id":"PMC_27667152","title":"Long non-coding RNAs, ASAP1-IT1, FAM215A, and LINC00472, in epithelial ovarian cancer.","date":"2016","source":"Gynecologic oncology","url":"https://pubmed.ncbi.nlm.nih.gov/27667152","citation_count":52,"is_preprint":false},{"pmid":"12522101","id":"PMC_12522101","title":"CrkL directs ASAP1 to peripheral focal adhesions.","date":"2003","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/12522101","citation_count":49,"is_preprint":false},{"pmid":"25673879","id":"PMC_25673879","title":"The Arf and Rab11 effector FIP3 acts synergistically with ASAP1 to direct Rabin8 in ciliary receptor targeting.","date":"2015","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/25673879","citation_count":49,"is_preprint":false},{"pmid":"12771146","id":"PMC_12771146","title":"The tyrosine kinase Pyk2 regulates Arf1 activity by phosphorylation and inhibition of the Arf-GTPase-activating protein ASAP1.","date":"2003","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/12771146","citation_count":45,"is_preprint":false},{"pmid":"17112341","id":"PMC_17112341","title":"Kinetic analysis of GTP hydrolysis catalysed by the Arf1-GTP-ASAP1 complex.","date":"2007","source":"The Biochemical journal","url":"https://pubmed.ncbi.nlm.nih.gov/17112341","citation_count":38,"is_preprint":false},{"pmid":"31246957","id":"PMC_31246957","title":"Loss of ASAP1 in mice impairs adipogenic and osteogenic differentiation of mesenchymal progenitor cells through dysregulation of FAK/Src and AKT signaling.","date":"2019","source":"PLoS genetics","url":"https://pubmed.ncbi.nlm.nih.gov/31246957","citation_count":34,"is_preprint":false},{"pmid":"32011035","id":"PMC_32011035","title":"Novel ASAP1-USP6, FAT1-USP6, SAR1A-USP6, and TNC-USP6 fusions in primary aneurysmal bone cyst.","date":"2020","source":"Genes, chromosomes & cancer","url":"https://pubmed.ncbi.nlm.nih.gov/32011035","citation_count":34,"is_preprint":false},{"pmid":"16038802","id":"PMC_16038802","title":"Regulation of ASAP1 by phospholipids is dependent on the interface between the PH and Arf GAP domains.","date":"2005","source":"Cellular signalling","url":"https://pubmed.ncbi.nlm.nih.gov/16038802","citation_count":34,"is_preprint":false},{"pmid":"26893376","id":"PMC_26893376","title":"The Arf GTPase-activating Protein, ASAP1, Binds Nonmuscle Myosin 2A to Control Remodeling of the Actomyosin Network.","date":"2016","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/26893376","citation_count":31,"is_preprint":false},{"pmid":"22682072","id":"PMC_22682072","title":"A SNP in ASAP1 gene is associated with meat quality and production traits in Nelore breed.","date":"2012","source":"Meat science","url":"https://pubmed.ncbi.nlm.nih.gov/22682072","citation_count":31,"is_preprint":false},{"pmid":"29653361","id":"PMC_29653361","title":"LncRNA ASAP1-IT1 positively modulates the development of cholangiocarcinoma via hedgehog signaling pathway.","date":"2018","source":"Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie","url":"https://pubmed.ncbi.nlm.nih.gov/29653361","citation_count":30,"is_preprint":false},{"pmid":"15212764","id":"PMC_15212764","title":"Differences between AGAP1, ASAP1 and Arf GAP1 in substrate recognition: interaction with the N-terminus of Arf1.","date":"2004","source":"Cellular signalling","url":"https://pubmed.ncbi.nlm.nih.gov/15212764","citation_count":30,"is_preprint":false},{"pmid":"25879659","id":"PMC_25879659","title":"A pooled shRNA screen for regulators of primary mammary stem and progenitor cells identifies roles for Asap1 and Prox1.","date":"2015","source":"BMC cancer","url":"https://pubmed.ncbi.nlm.nih.gov/25879659","citation_count":29,"is_preprint":false},{"pmid":"32998886","id":"PMC_32998886","title":"Membrane surface recognition by the ASAP1 PH domain and consequences for interactions with the small GTPase Arf1.","date":"2020","source":"Science advances","url":"https://pubmed.ncbi.nlm.nih.gov/32998886","citation_count":29,"is_preprint":false},{"pmid":"32235890","id":"PMC_32235890","title":"Integrative analysis of genomic amplification-dependent expression and loss-of-function screen identifies ASAP1 as a driver gene in triple-negative breast cancer progression.","date":"2020","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/32235890","citation_count":28,"is_preprint":false},{"pmid":"31785555","id":"PMC_31785555","title":"The ArfGAP ASAP1 Controls Actin Stress Fiber Organization via Its N-BAR Domain.","date":"2019","source":"iScience","url":"https://pubmed.ncbi.nlm.nih.gov/31785555","citation_count":27,"is_preprint":false},{"pmid":"27694689","id":"PMC_27694689","title":"eIF5B increases ASAP1 expression to promote HCC proliferation and invasion.","date":"2016","source":"Oncotarget","url":"https://pubmed.ncbi.nlm.nih.gov/27694689","citation_count":25,"is_preprint":false},{"pmid":"32444496","id":"PMC_32444496","title":"The BAR domain of the Arf GTPase-activating protein ASAP1 directly binds actin filaments.","date":"2020","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/32444496","citation_count":24,"is_preprint":false},{"pmid":"36792578","id":"PMC_36792578","title":"ASAP1 activates the IQGAP1/CDC42 pathway to promote tumor progression and chemotherapy resistance in gastric cancer.","date":"2023","source":"Cell death & disease","url":"https://pubmed.ncbi.nlm.nih.gov/36792578","citation_count":23,"is_preprint":false},{"pmid":"16332543","id":"PMC_16332543","title":"Mutational analysis of the Arf1*GTP/Arf GAP interface reveals an Arf1 mutant that selectively affects the Arf GAP ASAP1.","date":"2005","source":"Current biology : CB","url":"https://pubmed.ncbi.nlm.nih.gov/16332543","citation_count":23,"is_preprint":false},{"pmid":"32566028","id":"PMC_32566028","title":"Expression of ASAP1 and FAK in gastric cancer and its clinicopathological significance.","date":"2020","source":"Oncology letters","url":"https://pubmed.ncbi.nlm.nih.gov/32566028","citation_count":22,"is_preprint":false},{"pmid":"29541211","id":"PMC_29541211","title":"Lentiviral vector mediated-ASAP1 expression promotes epithelial to mesenchymal transition in ovarian cancer cells.","date":"2018","source":"Oncology letters","url":"https://pubmed.ncbi.nlm.nih.gov/29541211","citation_count":21,"is_preprint":false},{"pmid":"18675341","id":"PMC_18675341","title":"Dynamic interaction between Arf GAP and PH domains of ASAP1 in the regulation of GAP activity.","date":"2008","source":"Cellular signalling","url":"https://pubmed.ncbi.nlm.nih.gov/18675341","citation_count":21,"is_preprint":false},{"pmid":"29364481","id":"PMC_29364481","title":"Long non-coding RNA ASAP1-IT1 promotes cell proliferation, invasion and metastasis through the PTEN/AKT signaling axis in non-small cell lung cancer.","date":"2018","source":"European review for medical and pharmacological sciences","url":"https://pubmed.ncbi.nlm.nih.gov/29364481","citation_count":20,"is_preprint":false},{"pmid":"18952802","id":"PMC_18952802","title":"The AsaP1 peptidase of Aeromonas salmonicida subsp. achromogenes is a highly conserved deuterolysin metalloprotease (family M35) and a major virulence factor.","date":"2008","source":"Journal of bacteriology","url":"https://pubmed.ncbi.nlm.nih.gov/18952802","citation_count":20,"is_preprint":false},{"pmid":"37989735","id":"PMC_37989735","title":"Myr-Arf1 conformational flexibility at the membrane surface sheds light on the interactions with ArfGAP ASAP1.","date":"2023","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/37989735","citation_count":19,"is_preprint":false},{"pmid":"21352810","id":"PMC_21352810","title":"GEFH1 binds ASAP1 and regulates podosome formation.","date":"2011","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/21352810","citation_count":16,"is_preprint":false},{"pmid":"27910913","id":"PMC_27910913","title":"Loss of EGFR-ASAP1 signaling in metastatic and unresectable hepatoblastoma.","date":"2016","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/27910913","citation_count":16,"is_preprint":false},{"pmid":"33576454","id":"PMC_33576454","title":"Long non‑coding RNA ASAP1‑IT1 suppresses ovarian cancer progression by regulating Hippo/YAP signaling.","date":"2021","source":"International journal of molecular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/33576454","citation_count":15,"is_preprint":false},{"pmid":"27227929","id":"PMC_27227929","title":"No Significant Effect of ASAP1 Gene Variants on the Susceptibility to Tuberculosis in Chinese Population.","date":"2016","source":"Medicine","url":"https://pubmed.ncbi.nlm.nih.gov/27227929","citation_count":13,"is_preprint":false},{"pmid":"31591270","id":"PMC_31591270","title":"Interaction of the N terminus of ADP-ribosylation factor with the PH domain of the GTPase-activating protein ASAP1 requires phosphatidylinositol 4,5-bisphosphate.","date":"2019","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/31591270","citation_count":13,"is_preprint":false},{"pmid":"31293161","id":"PMC_31293161","title":"Probing the Broad Time Scale and Heterogeneous Conformational Dynamics in the Catalytic Core of the Arf-GAP ASAP1 via Methyl Adiabatic Relaxation Dispersion.","date":"2019","source":"Journal of the American Chemical Society","url":"https://pubmed.ncbi.nlm.nih.gov/31293161","citation_count":13,"is_preprint":false},{"pmid":"31089398","id":"PMC_31089398","title":"A Common Variant of ASAP1 Is Associated with Tuberculosis Susceptibility in the Han Chinese Population.","date":"2019","source":"Disease markers","url":"https://pubmed.ncbi.nlm.nih.gov/31089398","citation_count":11,"is_preprint":false},{"pmid":"37500798","id":"PMC_37500798","title":"Activation of NFAT by HGF and IGF-1 via ARF6 and its effector ASAP1 promotes uveal melanoma metastasis.","date":"2023","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/37500798","citation_count":11,"is_preprint":false},{"pmid":"35143843","id":"PMC_35143843","title":"A lysine-rich cluster in the N-BAR domain of ARF GTPase-activating protein ASAP1 is necessary for binding and bundling actin filaments.","date":"2022","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/35143843","citation_count":9,"is_preprint":false},{"pmid":"23811350","id":"PMC_23811350","title":"Innate and adaptive immune responses of Arctic charr (Salvelinus alpinus, L.) during infection with Aeromonas salmonicida subsp. achromogenes and the effect of the AsaP1 toxin.","date":"2013","source":"Fish & shellfish immunology","url":"https://pubmed.ncbi.nlm.nih.gov/23811350","citation_count":9,"is_preprint":false},{"pmid":"29545860","id":"PMC_29545860","title":"ASAP1 gene polymorphisms are associated with susceptibility to tuberculosis in a Chinese Xinjiang Muslim population.","date":"2018","source":"Experimental and therapeutic medicine","url":"https://pubmed.ncbi.nlm.nih.gov/29545860","citation_count":8,"is_preprint":false},{"pmid":"33194781","id":"PMC_33194781","title":"Asap1 Affects the Susceptibility of Zebrafish to Mycobacterium by Regulating Macrophage Migration.","date":"2020","source":"Frontiers in cellular and infection microbiology","url":"https://pubmed.ncbi.nlm.nih.gov/33194781","citation_count":7,"is_preprint":false},{"pmid":"36110251","id":"PMC_36110251","title":"Clinicopathological Implications of ASAP1 Expression in Hepatocellular Carcinoma.","date":"2022","source":"Pathology oncology research : POR","url":"https://pubmed.ncbi.nlm.nih.gov/36110251","citation_count":7,"is_preprint":false},{"pmid":"34715859","id":"PMC_34715859","title":"LncRNA ASAP1-IT1 enhances cancer cell stemness via regulating miR-509-3p/YAP1 axis in NSCLC.","date":"2021","source":"Cancer cell international","url":"https://pubmed.ncbi.nlm.nih.gov/34715859","citation_count":7,"is_preprint":false},{"pmid":"35181478","id":"PMC_35181478","title":"Loss of ASAP1 in the MMTV-PyMT model of luminal breast cancer activates AKT, accelerates tumorigenesis, and promotes metastasis.","date":"2022","source":"Cancer letters","url":"https://pubmed.ncbi.nlm.nih.gov/35181478","citation_count":7,"is_preprint":false},{"pmid":"36674928","id":"PMC_36674928","title":"Crystal Structure of the SH3 Domain of ASAP1 in Complex with the Proline Rich Motif (PRM) of MICAL1 Reveals a Unique SH3/PRM Interaction Mode.","date":"2023","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/36674928","citation_count":7,"is_preprint":false},{"pmid":"37875175","id":"PMC_37875175","title":"miR-212/132 attenuates OVA-induced airway inflammation by inhibiting mast cells activation through MRGPRX2 and ASAP1.","date":"2023","source":"Experimental cell research","url":"https://pubmed.ncbi.nlm.nih.gov/37875175","citation_count":6,"is_preprint":false},{"pmid":"32307199","id":"PMC_32307199","title":"Knockout of ASAP1 induces autophagy in papillary thyroid carcinoma by inhibiting the mTOR signaling pathway.","date":"2020","source":"Pathology, research and practice","url":"https://pubmed.ncbi.nlm.nih.gov/32307199","citation_count":6,"is_preprint":false},{"pmid":"38472475","id":"PMC_38472475","title":"MSUT2 regulates tau spreading via adenosinergic signaling mediated ASAP1 pathway in neurons.","date":"2024","source":"Acta neuropathologica","url":"https://pubmed.ncbi.nlm.nih.gov/38472475","citation_count":5,"is_preprint":false},{"pmid":"36777845","id":"PMC_36777845","title":"Expression profiling of circular RNA reveals a potential miR-145-5p sponge function of circ-AFF2 and circ-ASAP1 in renal cell carcinoma.","date":"2023","source":"American journal of translational research","url":"https://pubmed.ncbi.nlm.nih.gov/36777845","citation_count":5,"is_preprint":false},{"pmid":"40424906","id":"PMC_40424906","title":"ASAP1 protein macromolecule promotes the growth and development of gastric cancer cells and the remodeling of F-actin cytoskeleton: Expression of VEGF and HIF-1 α proteins.","date":"2025","source":"International journal of biological macromolecules","url":"https://pubmed.ncbi.nlm.nih.gov/40424906","citation_count":4,"is_preprint":false},{"pmid":"34058694","id":"PMC_34058694","title":"ASAP1 regulates the uptake of Mycobacterium tuberculosis H37Ra in THP1-derived macrophages by remodeling actin cytoskeleton.","date":"2021","source":"Tuberculosis (Edinburgh, Scotland)","url":"https://pubmed.ncbi.nlm.nih.gov/34058694","citation_count":4,"is_preprint":false},{"pmid":"10583691","id":"PMC_10583691","title":"Monoclonal antibodies against AsaP1, a major exotoxin of the fish pathogen Aeromonas salmonicida subsp. achromogenes, and their application in ELISA.","date":"1999","source":"Journal of applied microbiology","url":"https://pubmed.ncbi.nlm.nih.gov/10583691","citation_count":4,"is_preprint":false},{"pmid":"35712508","id":"PMC_35712508","title":"Novel Role of Long Non-Coding RNA ASAP1-IT1 in Progression of Hepatocellular Carcinoma.","date":"2022","source":"Frontiers in oncology","url":"https://pubmed.ncbi.nlm.nih.gov/35712508","citation_count":3,"is_preprint":false},{"pmid":"36249417","id":"PMC_36249417","title":"Polymorphisms in the ASAP1 and SP110 Genes and Its Association with the Susceptibility to Pulmonary Tuberculosis in a Mongolian Population.","date":"2022","source":"Journal of immunology research","url":"https://pubmed.ncbi.nlm.nih.gov/36249417","citation_count":3,"is_preprint":false},{"pmid":"23031341","id":"PMC_23031341","title":"Toxoid construction of AsaP1, a lethal toxic aspzincin metalloendopeptidase of Aeromonas salmonicida subsp. achromogenes, and studies of its activity and processing.","date":"2012","source":"Veterinary microbiology","url":"https://pubmed.ncbi.nlm.nih.gov/23031341","citation_count":3,"is_preprint":false},{"pmid":"39287759","id":"PMC_39287759","title":"CUL1 exacerbates glucocorticoid-induced osteoporosis by enhancing ASAP1 ubiquitination.","date":"2024","source":"Hormones (Athens, Greece)","url":"https://pubmed.ncbi.nlm.nih.gov/39287759","citation_count":3,"is_preprint":false},{"pmid":"39495123","id":"PMC_39495123","title":"ASAP1 and ARF1 Regulate Myogenic Differentiation in Rhabdomyosarcoma by Modulating TAZ Activity.","date":"2025","source":"Molecular cancer research : MCR","url":"https://pubmed.ncbi.nlm.nih.gov/39495123","citation_count":2,"is_preprint":false},{"pmid":"38874840","id":"PMC_38874840","title":"Inhibition of ASAP1 Modulates the Tumor Immune Microenvironment and Suppresses Lung Cancer Metastasis via the p-STAT3 Signaling Pathway.","date":"2024","source":"Cell biochemistry and biophysics","url":"https://pubmed.ncbi.nlm.nih.gov/38874840","citation_count":2,"is_preprint":false},{"pmid":"40194952","id":"PMC_40194952","title":"Actin Binding to the BAR Domain and Arf GAP Activity of ASAP1 Coordinately Control Actin Stress Fibers and Focal Adhesions.","date":"2025","source":"Biology of the cell","url":"https://pubmed.ncbi.nlm.nih.gov/40194952","citation_count":2,"is_preprint":false},{"pmid":"39763923","id":"PMC_39763923","title":"The PH domain in the ArfGAP ASAP1 drives catalytic activation through an unprecedented allosteric mechanism.","date":"2024","source":"bioRxiv : the preprint server for biology","url":"https://pubmed.ncbi.nlm.nih.gov/39763923","citation_count":2,"is_preprint":false},{"pmid":"19574642","id":"PMC_19574642","title":"Crystallization and preliminary X-ray diffraction studies of AsaP1_E294A and AsaP1_E294Q, two inactive mutants of the toxic zinc metallopeptidase AsaP1 from Aeromonas salmonicida subsp. achromogenes.","date":"2009","source":"Acta crystallographica. Section F, Structural biology and crystallization communications","url":"https://pubmed.ncbi.nlm.nih.gov/19574642","citation_count":2,"is_preprint":false},{"pmid":"25970765","id":"PMC_25970765","title":"Regulator of dendritic cell migration, ASAP1 is associated with increased susceptibility to tuberculosis.","date":"2015","source":"Clinical genetics","url":"https://pubmed.ncbi.nlm.nih.gov/25970765","citation_count":2,"is_preprint":false},{"pmid":"41208482","id":"PMC_41208482","title":"The MYO1F interactome reveals ASAP1, CD2AP and SH3KBP1 as novel adaptor proteins in podosomes and phagosomes.","date":"2025","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/41208482","citation_count":2,"is_preprint":false},{"pmid":"31894003","id":"PMC_31894003","title":"[ASAP1 knockdown reduces migration of RAW264.7 cells infected with Mycobacterium tuberculosis].","date":"2019","source":"Xi bao yu fen zi mian yi xue za zhi = Chinese journal of cellular and molecular immunology","url":"https://pubmed.ncbi.nlm.nih.gov/31894003","citation_count":1,"is_preprint":false},{"pmid":"16413287","id":"PMC_16413287","title":"Assay and functional properties of the tyrosine kinase Pyk2 in regulation of Arf1 through ASAP1 phosphorylation.","date":"2005","source":"Methods in enzymology","url":"https://pubmed.ncbi.nlm.nih.gov/16413287","citation_count":1,"is_preprint":false},{"pmid":"37686290","id":"PMC_37686290","title":"An Uncharacterised lncRNA Coded by the ASAP1 Locus Is Downregulated in Serum of Type 2 Diabetes Mellitus Patients.","date":"2023","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/37686290","citation_count":1,"is_preprint":false},{"pmid":"40742091","id":"PMC_40742091","title":"ASAP1 Promotes Epithelial to Mesenchymal Transition by Activating the TGFβ Pathway in Papillary Thyroid Cancer.","date":"2025","source":"Cancer medicine","url":"https://pubmed.ncbi.nlm.nih.gov/40742091","citation_count":0,"is_preprint":false},{"pmid":"39659924","id":"PMC_39659924","title":"ASAP1 promotes extrahepatic cholangiocarcinoma progression by regulating the Wnt/β-catenin pathway in vitro and in vivo.","date":"2024","source":"American journal of cancer research","url":"https://pubmed.ncbi.nlm.nih.gov/39659924","citation_count":0,"is_preprint":false},{"pmid":"39265663","id":"PMC_39265663","title":"New targets and designed inhibitors of ASAP Arf-GAPs derived from structural characterization of the ASAP1/440-kD ankyrin-B interaction.","date":"2024","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/39265663","citation_count":0,"is_preprint":false},{"pmid":"41856082","id":"PMC_41856082","title":"ASAP1 gene InDel variants are associated with enhanced goat resistance against Brucella infection.","date":"2026","source":"Animal bioscience","url":"https://pubmed.ncbi.nlm.nih.gov/41856082","citation_count":0,"is_preprint":false},{"pmid":"40166026","id":"PMC_40166026","title":"The PH domain in the ArfGAP ASAP1 drives catalytic activation through an unprecedented allosteric mechanism.","date":"2025","source":"Research square","url":"https://pubmed.ncbi.nlm.nih.gov/40166026","citation_count":0,"is_preprint":false},{"pmid":"41659712","id":"PMC_41659712","title":"Case Report: Novel ASAP1::BRAF fusion in a young adult with low-grade temporal lobe glioma.","date":"2026","source":"Frontiers in oncology","url":"https://pubmed.ncbi.nlm.nih.gov/41659712","citation_count":0,"is_preprint":false},{"pmid":"40470248","id":"PMC_40470248","title":"Redefining PH Domain Function: An Active Allosteric Mechanism in ASAP1-Mediated Arf1 GTP Hydrolysis.","date":"2025","source":"Research square","url":"https://pubmed.ncbi.nlm.nih.gov/40470248","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.08.08.669310","title":"Absolute Membrane Potential Recording with ASAP-Type Genetically Encoded Voltage Indicators Using Fluorescence Lifetime Imaging","date":"2025-08-12","source":"bioRxiv","url":"https://doi.org/10.1101/2025.08.08.669310","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":46345,"output_tokens":11065,"usd":0.152505,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":22032,"output_tokens":6835,"usd":0.140517,"stage2_stop_reason":"end_turn"},"total_usd":0.293022,"stage1_batch_id":"msgbatch_0179Z2pxRYBsXi2xADneafRw","stage2_batch_id":"msgbatch_01LaY9ZaVdgjWArcZo9eqQE5","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1998,\n      \"finding\": \"ASAP1 was purified and cloned as a PIP2-dependent Arf1 GTPase-activating protein (GAP). The PH, zinc finger, and ANK repeat regions together possess PIP2-dependent GAP activity on Arf1 and Arf5 in vitro. ASAP1 associates with the SH3 domains of Src family members and Crk adapter protein in vitro, coprecipitates with Src from cell lysates, and is phosphorylated on tyrosine residues in cells expressing activated Src through the same proline-rich class II Src SH3 binding site.\",\n      \"method\": \"Protein purification, in vitro GAP assay, GST pulldown, co-immunoprecipitation, in-cell phosphorylation\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — direct biochemical reconstitution of GAP activity, multiple orthogonal binding assays, replicated across multiple Arf substrates\",\n      \"pmids\": [\"9819391\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"ASAP1 localizes to focal adhesions and cycles with focal adhesion proteins during cell migration. Overexpression of ASAP1 altered focal adhesion morphology and blocked cell spreading and PDGF-induced dorsal ruffle formation; a GAP-inactive mutant had reduced effect on spreading and increased dorsal ruffle formation, demonstrating that GAP activity is required for these cytoskeletal functions.\",\n      \"method\": \"Fluorescence microscopy, cell spreading assay, PDGF-stimulation assay, overexpression of wild-type vs. catalytically inactive mutant\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct localization with functional consequence, loss-of-function and gain-of-function with defined cellular phenotype, replicated findings\",\n      \"pmids\": [\"10725410\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"PIP2 activates ASAP1 GAP activity by binding the PH domain, which acts as an allosteric site rather than merely a membrane recruitment signal. The PH domain is necessary for GAP activity even in the absence of phospholipids; PIP2 binding causes a conformational change in the Arf GAP domain. Activation and membrane recruitment can be uncoupled.\",\n      \"method\": \"In vitro GAP activity assays, limited proteolysis, domain deletion/mutation analysis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — reconstitution in vitro with mutagenesis and multiple orthogonal biochemical methods\",\n      \"pmids\": [\"10734117\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"ASAP1 (DEF-1) functions as an ARF GAP for ARF1 but not ARF6 in vivo (cell-based ARF GAP assay), unlike ACAP1, ACAP2, and ARFGAP1 which act on both. Enhancement of cell motility by ASAP1 is dependent on GAP activity, whereas inhibition of cell spreading by ASAP1 is GAP-activity independent, indicating two distinct downstream pathways.\",\n      \"method\": \"Cell-based ARF GAP assay, stable overexpression, cell motility assay, GAP-domain deletion mutant\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — cell-based GAP assay with direct comparison of substrates and domain mutants showing mechanistic separation of phenotypes\",\n      \"pmids\": [\"11773070\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"ASAP1 binds to focal adhesion kinase (FAK) via an interaction between the C-terminal SH3 domain of ASAP1 and the second proline-rich motif in the C-terminal region of FAK. This interaction contributes to focal adhesion assembly; ASAP1 variants that could not bind FAK or lacked GAP activity showed reduced inhibition of cell spreading and failed to prevent paxillin/FAK organization in focal adhesions.\",\n      \"method\": \"Affinity chromatography, yeast two-hybrid, GST pulldown, co-immunoprecipitation, cell spreading assay, overexpression of mutants\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP and pulldown confirming direct interaction, mutagenesis linking binding to cellular function\",\n      \"pmids\": [\"12058076\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"CrkL binds ASAP1 via its N-terminal SH3 domain and directs ASAP1 to peripheral focal adhesions. CrkL co-expression recruited endogenous and exogenous ASAP1 to CrkL-induced focal adhesions; an SH2-mutated CrkL that cannot localize to focal adhesions failed to recruit ASAP1.\",\n      \"method\": \"Pulldown/mass spectrometry, co-immunoprecipitation, fluorescence microscopy, overexpression of wild-type vs. SH2-mutant CrkL\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct identification in platelets, confirmed in cells with mutagenesis; single lab\",\n      \"pmids\": [\"12522101\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"Pyk2 interacts with ASAP1 through the proline-rich regions of Pyk2 and the SH3 domain of ASAP1. Pyk2 directly phosphorylates ASAP1 on tyrosine residues (Y308 and Y782) in vitro and in cells; this phosphorylation inhibits ASAP1 GAP activity toward Arf1 as measured by fluorimetric GTPase assay.\",\n      \"method\": \"Yeast two-hybrid, pulldown, co-immunoprecipitation, in vitro kinase assay, fluorimetric Arf-GTPase assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — in vitro kinase assay with mapped phosphorylation sites and direct measurement of GAP activity inhibition; single lab with multiple orthogonal methods\",\n      \"pmids\": [\"12771146\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"ASAP1 interacts with the N-terminus (amino acids 2–17) of Arf1 at an interface distinct from other Arf GAPs (AGAP1, ArfGAP1). Specific mutations in Arf1 alpha-helix 3 and switch regions (notably I46D reducing ASAP1-catalyzed hydrolysis ~10,000-fold with isolated effect on kcat) distinguish ASAP1's catalytic interface from other Arf GAPs.\",\n      \"method\": \"In vitro GAP activity assay, Arf1 mutagenesis, antibody epitope sequestration, in vivo localization studies\",\n      \"journal\": \"Cellular signalling\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — quantitative kinetic analysis with multiple point mutants and in vivo validation\",\n      \"pmids\": [\"15212764\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"ASAP1 regulation by phospholipids requires a direct interaction between its PH and Arf GAP domains; the two domains form a composite substrate-binding site. Saturation kinetics, limited proteolysis, FRET, and fluorescence spectrometry support a two-step model: conformational change upon membrane recruitment followed by a second change upon PIP2 binding.\",\n      \"method\": \"Saturation kinetics, limited proteolysis, FRET, fluorescence spectrometry, analytical ultracentrifugation\",\n      \"journal\": \"Cellular signalling\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — multiple orthogonal biophysical methods in vitro characterizing domain interaction and conformational mechanism\",\n      \"pmids\": [\"16038802\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"CD2AP (CD2-associated protein) binds ASAP1 through its N-terminal SH3 domains. Sequestration of endogenous ASAP1 to mitochondria via a CD2AP SH3-mito fusion protein inhibited cell spreading and migration in response to fibronectin and caused increased GTP loading on Arf1 and loss of paxillin from adhesions. siRNA knockdown of ASAP1 produced the same phenotypes.\",\n      \"method\": \"Affinity chromatography/mass spectrometry, co-immunoprecipitation, GST pulldown, mitochondria mislocalization strategy, siRNA knockdown, GTP-loading assay, fluorescence microscopy\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct identification of binding partner, two independent loss-of-function strategies showing same cellular phenotype and downstream Arf1 GTP loading\",\n      \"pmids\": [\"15632162\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Arf1 mutant I46D selectively abolishes ASAP1-catalyzed GTP hydrolysis (~10,000-fold reduction in kcat) while having minimal effect on AGAP1 (~3-fold). In vivo, [I46D]Arf1 acts as a constitutively active mutant at the cell periphery, disrupting ASAP1 and paxillin localization.\",\n      \"method\": \"In vitro GAP assay with kinetic analysis, in vivo localization by fluorescence microscopy, mutagenesis\",\n      \"journal\": \"Current biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — quantitative kinetics with mutagenesis and in vivo functional validation; single lab\",\n      \"pmids\": [\"16332543\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"ASAP1 contains an N-terminal BAR domain that together with the PH domain dimerizes into an extended structure that binds acidic phospholipid-containing vesicles and bends membranes to form tubular structures. This bending activity is regulated by Arf1·GTP binding to the Arf GAP domain (acting as an Arf effector). ASAP1 colocalizes with EGFR in tubular recycling structures; the BAR domain is necessary for ASAP1 function in EGFR trafficking and cell spreading.\",\n      \"method\": \"In vitro membrane tubulation assay with electron microscopy, vesicle sedimentation, dimerization analysis, live-cell imaging, EGFR trafficking assay, BAR domain deletion mutant\",\n      \"journal\": \"Current biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — in vitro reconstitution of membrane bending, EM, and cell-based trafficking/spreading assays with domain deletion mutant\",\n      \"pmids\": [\"16431365\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"ASAP1 uses Arf1-GTP as substrate with kcat ~57 s−1 and Km ~2.2 μM (steady-state). AlF4− stabilizes an Arf1-GDP·ASAP1 transition-state complex. Arg-497 mutation severely affects kcat with minimal effect on Km. Mutations of residues predicted to affect Arf1 affinity (W479, I490, R505, L511, D512) instead primarily affected kcat, supporting a conformational change in the Arf1-GTP·ASAP1 complex during catalysis.\",\n      \"method\": \"Steady-state and single-turnover kinetics, AlF4− trapping, mutagenesis, in vivo dorsal ruffle assay\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — rigorous quantitative kinetics with mutagenesis and transition-state trapping; single lab with multiple methods\",\n      \"pmids\": [\"17112341\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"ASAP1 is required for formation of invadopodia and podosomes. The BAR domain, SH3 domain, and Src phosphorylation site of ASAP1 are each required for podosome formation. The Src binding site and GAP activity are dispensable for podosome formation, suggesting ASAP1 functions as a coincidence detector integrating SH3-domain protein interactions, BAR domain scaffolding, and Src phosphorylation.\",\n      \"method\": \"siRNA knockdown, rescue with ASAP1 mutants (BAR-deleted, SH3-deleted, phosphorylation site mutant, GAP-inactive), fluorescence microscopy\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — systematic domain mutagenesis with siRNA rescue defining necessary structural elements for a specific cellular phenotype\",\n      \"pmids\": [\"17893324\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"ASAP1 directly binds FIP3 (a Rab11 and Arf6 effector) through its BAR domain as identified by yeast two-hybrid and confirmed by co-immunoprecipitation and in vitro pulldown. FIP3 binding to the BAR domain stimulates ASAP1 GAP activity against Arf1 but not Arf6. ASAP1 forms a ternary complex with Rab11 via FIP3. ASAP1 colocalizes with FIP3 in pericentrosomal recycling endosomes; ASAP1 or FIP3 depletion alters transferrin receptor localization and transferrin trafficking.\",\n      \"method\": \"Yeast two-hybrid, co-immunoprecipitation, in vitro pulldown, GAP activity assay, siRNA knockdown, fluorescence microscopy, transferrin trafficking assay\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal binding assays, functional GAP stimulation assay, and localization with trafficking phenotype\",\n      \"pmids\": [\"18685082\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"The BAR domain of ASAP1 autoinhibits GAP activity by intramolecular interaction with the PH and/or Arf GAP domains. The catalytic power of PZA (PH+GAP+Ank) is greater than BAR-PZA; the BAR domain increases Km and decreases kcat. The effect requires the N-terminal loop of the BAR domain and is not due to differential membrane association or membrane curvature changes.\",\n      \"method\": \"Sedimentation velocity analytical ultracentrifugation, in vitro GAP assay on large unilamellar vesicles, steady-state and single-turnover kinetics, BAR loop mutagenesis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — quantitative kinetic analysis with domain deletion and mutagenesis demonstrating autoinhibitory mechanism in vitro\",\n      \"pmids\": [\"19017632\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"NMR spectroscopy reveals a dynamic interaction between the PH and Arf GAP domains of ASAP1: the domains interact transiently, with the interaction partially occluding the PIP2 binding site in solution. PIP2 binding alters PH domain conformation but has little effect on Arf GAP domain structure. PH domain loop mutations at the GAP-domain interface affect PIP2 binding and both Km and kcat for Arf1 GTP hydrolysis.\",\n      \"method\": \"NMR spectroscopy, in vitro GAP kinetics, mutagenesis, analytical ultracentrifugation, lipid binding assays\",\n      \"journal\": \"Cellular signalling\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — NMR structural data combined with quantitative kinetics and mutagenesis; single lab with multiple orthogonal methods\",\n      \"pmids\": [\"18675341\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"GEFH1 (a RhoA guanine nucleotide exchange factor) binds the BAR domain of ASAP1, colocalizes with ASAP1 in podosomes, inhibits ASAP1 GAP activity, and negatively regulates podosome assembly. GEFH1 overexpression inhibited podosome assembly and a GEFH1 mutant lacking the BAR-binding domain was less effective; GEFH1 siRNA knockdown increased the rate of podosome assembly.\",\n      \"method\": \"Yeast two-hybrid, co-immunoprecipitation, fluorescence colocalization, GAP activity assay, overexpression, siRNA knockdown\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP confirming endogenous interaction, functional assays showing GAP inhibition and podosome regulation; single lab\",\n      \"pmids\": [\"21352810\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"ASAP1 is a scaffold for ciliary receptor targeting that brings together Arf4, Rab11, Rab8-GEF Rabin8, and rhodopsin. Ablation of ASAP1 abolishes ciliary targeting of rhodopsin and causes actin-rich periciliary membrane projections with mislocalized rhodopsin. ASAP1 recognizes the FR ciliary targeting signal of rhodopsin; rhodopsin FR-AA mutant fails to interact with Rab8 and cannot cross the periciliary diffusion barrier.\",\n      \"method\": \"siRNA knockdown, fluorescence microscopy, co-immunoprecipitation, transport assay in photoreceptors\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — loss-of-function with defined trafficking phenotype, identification of binding signal on cargo, multiple interaction partners confirmed\",\n      \"pmids\": [\"22983554\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Crystal structure of the ASAP1 PH domain was solved in unliganded and dibutyryl-PIP2-bound forms. PIP2 contacts both a canonical site (C site) and an atypical site (A site); PIP2 dependence of vesicle binding and GAP activity is sigmoidal (cooperative), distinct from the hyperbolic binding seen for PLC-δ1 PH domain. Mutations in either the C or A site reduced PIP2-dependent vesicle binding and GAP activity, supporting cooperative binding mechanism for rapid switching.\",\n      \"method\": \"X-ray crystallography, vesicle binding assay, in vitro GAP activity assay, site-directed mutagenesis\",\n      \"journal\": \"Structure\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure with functional validation by mutagenesis and cooperative binding analysis\",\n      \"pmids\": [\"26365802\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"ASAP1-depleted dendritic cells show impaired matrix degradation and migration. ASAP1 is involved in actin and membrane remodeling associated with podosomes in dendritic cells; genetic variants reducing ASAP1 expression in M. tuberculosis-infected dendritic cells may impair their migration.\",\n      \"method\": \"siRNA knockdown of ASAP1 in primary human dendritic cells, matrix degradation assay, migration assay\",\n      \"journal\": \"Nature genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct loss-of-function in primary cells with defined functional phenotype; single study in DCs\",\n      \"pmids\": [\"25774636\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"FIP3 competes with rhodopsin for binding to ASAP1 and displaces it from the ternary complex with Arf4-GTP and ASAP1. FIP3 promotes Rab11a activity and coordinates ASAP1 and Rab11a interactions with Rabin8, facilitating orderly Rab11-Rabin8-Rab8 cascade assembly for ciliary receptor trafficking. Ablation of FIP3 abolishes ciliary targeting similarly to ASAP1 ablation.\",\n      \"method\": \"Co-immunoprecipitation, competition binding assay, siRNA knockdown, fluorescence microscopy, trafficking assay\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP competition and loss-of-function in single lab; extends previous ciliary trafficking mechanism\",\n      \"pmids\": [\"25673879\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"The ASAP1 BAR domain together with the PH domain directly binds nonmuscle myosin 2A (NM2A) in vitro. ASAP1 and NM2A co-immunoprecipitate and colocalize in cells. ASAP1 knockdown reduced colocalization of NM2A and F-actin. Knockdown of either ASAP1 or NM2A produced similar defects in focal adhesions, cell migration, spreading, and circular dorsal ruffles. Exogenous NM2A rescued ASAP1-knockdown CDR defects, positioning ASAP1 as a positive regulator of NM2A.\",\n      \"method\": \"In vitro binding assay, co-immunoprecipitation, siRNA knockdown, rescue experiments, fluorescence microscopy\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct in vitro binding, reciprocal Co-IP, and epistasis by rescue experiments; single lab, multiple orthogonal methods\",\n      \"pmids\": [\"26893376\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"PIP2 controls binding of the N-terminal extension (residues 2–17) of ARF1 to the PH domain of ASAP1 and thereby regulates GAP activity. Deletion of the ARF1 N-terminus ([Δ17]ARF1) makes GAP activity largely PIP2-independent. A peptide of residues 2–17 inhibits GAP activity and binds PIP2-dependently to the PH domain including a 17-amino acid interdomain linker N-terminal to the first β-strand. Mutations in the linker or C-terminal α-helix of the PH domain decrease both ARF1 N-terminal binding and GAP activity, and reduce cellular actin remodeling.\",\n      \"method\": \"In vitro GAP assay with truncation and point mutants, peptide inhibition assay, NMR binding assay, cell-based actin remodeling assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — quantitative in vitro kinetics with mutagenesis, NMR binding validation, and cell-based functional assay; single lab, multiple orthogonal methods\",\n      \"pmids\": [\"31591270\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"ASAP1 depletion causes defects in actin stress fiber organization. The BAR-PH fragment is sufficient to affect actin; the N-BAR domain of ASAP1 directly binds and bundles actin filaments in vitro, whereas the Arf GAP and C-terminal SH3 domain reduce BAR-PH binding and bundling. Overexpression of ASAP1 enhanced actin remodeling; replacing the ASAP1 BAR domain with the ACAP1 BAR domain abolished actin effects.\",\n      \"method\": \"siRNA knockdown, overexpression, actin co-sedimentation, domain swap mutants, fluorescence microscopy\",\n      \"journal\": \"iScience\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct in vitro actin binding and bundling assays combined with domain mutants and cell-based phenotype; single lab\",\n      \"pmids\": [\"31785555\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"NMR methyl-geoHARD analysis of the ASAP1 ZA (PH-ArfGAP) domain reveals wide-range conformational dynamics (kex 10²–10⁵ s⁻¹) in the hydrophobic interior, including collective and local motions that may correlate with catalytic function and substrate recognition.\",\n      \"method\": \"Methyl-TROSY NMR, adiabatic relaxation dispersion, CPMG relaxation dispersion\",\n      \"journal\": \"Journal of the American Chemical Society\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 1 / Weak — rigorous NMR technique but functional correlation remains speculative; single lab, dynamics-only study\",\n      \"pmids\": [\"31293161\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Loss of ASAP1 in mice (gene-trap) impairs adipogenic and osteogenic differentiation of mesenchymal progenitor cells, causing growth retardation and delayed ossification. Mechanistically, FAK/Src and PI3K/AKT signaling is compromised in ASAP1-null MEFs, leading to impaired adipogenic and osteogenic differentiation.\",\n      \"method\": \"Gene-trap mouse model, in vitro differentiation assays, Western blotting for FAK/Src and PI3K/AKT pathway components\",\n      \"journal\": \"PLoS genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic loss-of-function with defined differentiation phenotype and pathway readout; single lab\",\n      \"pmids\": [\"31246957\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Binding of multiple PI(4,5)P2 molecules to the ASAP1 PH domain triggers a functionally relevant allosteric conformational switch and maintains the PH domain in a defined orientation that allows critical contacts with Arf1 at the membrane, as determined by combining NMR, neutron reflectometry, and molecular dynamics simulation.\",\n      \"method\": \"NMR, neutron reflectometry, molecular dynamics simulation\",\n      \"journal\": \"Science advances\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — three complementary structural/biophysical methods providing atomic-level mechanism; single integrated study\",\n      \"pmids\": [\"32998886\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"The ASAP1 N-BAR domain directly binds F-actin, homodimerization of ASAP1 aligns F-actin in predominantly unipolar bundles, and ASAP1 stabilizes actin filaments against depolymerization. The N-BAR domain moderately reduces spontaneous G-actin polymerization. Overexpression of ASAP1 BAR-PH tandem induced actin-filled cellular projections; an ASAP1 construct lacking the N-BAR domain failed to induce projections.\",\n      \"method\": \"Actin cosedimentation, polymerization and depolymerization assays, TIRF microscopy, confocal microscopy, electron microscopy, overexpression in fibroblasts\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — direct in vitro binding reconstitution with multiple structural and cell-based assays; multiple orthogonal methods\",\n      \"pmids\": [\"32444496\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Knockdown of ASAP1 in THP1-derived macrophages increased efficiency of Mycobacterium tuberculosis H37Ra entry and enhanced F-actin aggregation and vinculin/paxillin-rich puncta formation, identifying ASAP1 as a regulator of Mtb uptake through actin cytoskeleton remodeling.\",\n      \"method\": \"siRNA knockdown, fluorescence confocal microscopy, colony forming unit assay, F-actin staining\",\n      \"journal\": \"Tuberculosis\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function with defined entry and actin phenotype; single lab, single method type\",\n      \"pmids\": [\"34058694\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"A lysine-rich cluster (K75, K76, K79) in the N-BAR domain of ASAP1 is required for binding and bundling actin filaments. Charge-neutralizing or charge-reversing mutations at these positions reduced BAR-PH binding to F-actin and abrogated actin bundle formation in vitro and cellular actin remodeling in U2OS cells; [K75E, K76E, K79E] full-length ASAP1 did not rescue endogenous ASAP1 knockdown-induced reduction of stress fibers.\",\n      \"method\": \"Structural modeling, mutagenesis, actin co-sedimentation, in vitro bundling assay, cell-based actin remodeling assay, siRNA rescue\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — mutagenesis with in vitro reconstitution and cell-based rescue assay; single lab, multiple orthogonal methods\",\n      \"pmids\": [\"35143843\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Membrane-bound active Arf1 (Myr-Arf1) explores large conformational dynamics with its G domain oscillating between membrane-associated and membrane-distal conformations. Interaction with the ASAP1 PH domain restricts Arf1 G domain motions and locks it in a conformation exposing functionally relevant regions for catalysis.\",\n      \"method\": \"NMR, neutron reflectometry, molecular dynamics simulations\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — three complementary structural/biophysical methods; mechanistic insight into ASAP1–Arf1 interaction at membrane\",\n      \"pmids\": [\"37989735\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Crystal structure of the ASAP1 SH3 domain in complex with the MICAL1 proline-rich motif (PRM) revealed a unique binding mode: ASAP1 SH3 contains two negatively charged patches that recognize the 'xPx+Px+' sequence in MICAL1 PRM, yielding sub-μM affinity. This binding pocket (termed SH3AGS) is also found in GRAF and SKAP1 SH3 domains.\",\n      \"method\": \"X-ray crystallography, ITC/binding affinity measurement, mutagenesis\",\n      \"journal\": \"International journal of molecular sciences\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — crystal structure with biochemical binding validation; single lab\",\n      \"pmids\": [\"36674928\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"ASAP1 acts as an effector for ARF6 and mediates HGF/IGF-1 signaling to promote nuclear localization and transcriptional activity of NFAT1 in uveal melanoma. HGF and IGF-1 hyperactivate ARF6, which then interacts with ASAP1 to induce NFAT1 nuclear translocation; inhibition of ASAP1 or NFAT impairs cellular invasiveness and reduces metastasis in a xenograft model.\",\n      \"method\": \"Co-immunoprecipitation (ARF6-ASAP1), NFAT nuclear localization assay, siRNA knockdown, xenograft mouse model\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP for ARF6-ASAP1 interaction, functional loss-of-function with defined signaling phenotype and in vivo validation; single lab\",\n      \"pmids\": [\"37500798\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"ASAP1 activates the IQGAP1/CDC42 pathway by inhibiting ubiquitin-mediated degradation of IQGAP1, thereby enhancing CDC42 activity. Activated CDC42 upregulates the EGFR-MAPK pathway to promote chemotherapy resistance in gastric cancer.\",\n      \"method\": \"siRNA/overexpression, Co-IP, ubiquitination assay, Western blotting for CDC42/EGFR/MAPK pathway, in vitro and in vivo tumor assays\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP and ubiquitination assay identifying mechanism; single lab with multiple assays\",\n      \"pmids\": [\"36792578\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"CUL1 promotes ubiquitination and degradation of ASAP1 via the SCF-FBXW7 complex, suppressing osteoblast proliferation and osteogenesis. CUL1 silencing moderated Dex-induced inhibition of proliferation and osteogenesis by restoring ASAP1 levels.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, CUL1 siRNA knockdown, osteogenesis assay, mouse osteoporosis model\",\n      \"journal\": \"Hormones\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP and ubiquitination assay identifying the E3 complex; single lab with in vivo validation\",\n      \"pmids\": [\"39287759\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"The SH3 domain of ASAP1 binds a 12-residue positively charged peptide from the neuronal scaffold protein 440 kDa ankyrin-B via a noncanonical SH3-ligand binding mode. The crystal structure of ASAP1-SH3 in complex with this gAnkB peptide defined a consensus ASAP1-SH3 binding motif, enabling identification of novel binding partners including Clasp1/2, ALS2, β-Pix, DAPK3, PHIP, and Limk1.\",\n      \"method\": \"Crystal structure determination, ITC/binding affinity measurement, mutagenesis, in silico database search\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — crystal structure with quantitative binding and mutagenesis; single lab with multiple orthogonal methods\",\n      \"pmids\": [\"39265663\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"MSUT2 regulates tau seed internalization into neurons via adenosine receptor 1 (A1AR)-mediated modulation of ASAP1 activity. Down-regulation or inhibition of A1AR modulates ASAP1 activity, reducing internalization of pathogenic tau seeds and tau pathology in neuron cultures and mouse models.\",\n      \"method\": \"siRNA knockdown, tau seeding assay, A1AR inhibitor treatment, neuron culture and mouse model of tau pathology\",\n      \"journal\": \"Acta neuropathologica\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function genetics with defined tau seeding phenotype and pathway placement; single lab\",\n      \"pmids\": [\"38472475\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"ASAP1 and ARF1 are necessary for myogenic differentiation in FN-RMS. Loss of ASAP1 or ARF1/ARF5 (GAP substrates) blocks differentiation and prevents MEK-inhibition-induced inactivation of TAZ (WWTR1), a pro-proliferative transcriptional co-activator. Dual knockdown of ASAP1 and WWTR1 rescued MEKi-induced differentiation, placing ASAP1 upstream of TAZ inactivation.\",\n      \"method\": \"siRNA knockdown, MEK inhibitor treatment, Western blotting for TAZ phosphorylation/activation, myogenic transcription factor expression assay, epistasis rescue experiment\",\n      \"journal\": \"Molecular cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — epistasis experiments placing ASAP1 upstream of TAZ in differentiation pathway; single lab with multiple knockdown conditions\",\n      \"pmids\": [\"39495123\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"MYO1F interacts with ASAP1 through an SH3-domain-dependent interaction (proximity labeling proteomics, structural modeling, mutagenesis), and ASAP1 colocalizes with MYO1F at actin-rich podosomes and phagocytic cups in macrophages and microglia.\",\n      \"method\": \"Proximity labeling/proteomics, structural modeling, mutagenesis, immunofluorescence colocalization\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — proximity proteomics with structural and mutagenesis validation; functional consequence not directly tested for ASAP1 specifically\",\n      \"pmids\": [\"41208482\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"ASAP1 interacts with the SMAD2/3 complex and forms a positive feedback loop with TGFβ signaling, promoting EMT and cell invasiveness in papillary thyroid cancer cells. ASAP1 knockdown reduced p-SMAD2 levels; co-immunoprecipitation confirmed ASAP1-SMAD2/3 interaction.\",\n      \"method\": \"Co-immunoprecipitation, immunofluorescence, Western blotting for p-SMAD2, luciferase reporter assay, lentiviral knockdown/overexpression\",\n      \"journal\": \"Cancer medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP confirming physical interaction and TGFβ pathway assays; single lab\",\n      \"pmids\": [\"40742091\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Both the Arf GAP domain activity and the BAR domain actin/NM2A-binding activity of ASAP1 are required coordinately to maintain focal adhesions and actin stress fibers; neither domain alone is sufficient. Arf5 (a GAP substrate) loss-of-function phenocopies ASAP1 knockdown on SFs and FAs.\",\n      \"method\": \"siRNA knockdown, rescue with domain mutants (GAP-inactive, BAR-deleted), dominant negative and GTPase-deficient Arf5 mutants, fluorescence microscopy in four cell lines\",\n      \"journal\": \"Biology of the cell\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — systematic domain rescue in multiple cell lines with epistatic Arf5 experiments; single lab\",\n      \"pmids\": [\"40194952\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"The PH domain of ASAP1 enhances GAP activity by >7 orders of magnitude by acting as an active catalytic component, not merely a membrane recruitment signal. NMR and MD simulations show the PH domain directly contacts Arf·GTP at the membrane and allosterically drives conformational rearrangements of the GTP binding site to facilitate charge stabilization and accelerate GTP hydrolysis; mathematical modeling indicates this allosteric contribution equals membrane recruitment in importance.\",\n      \"method\": \"NMR, molecular dynamics simulation, kinetic assays, mutagenesis, mathematical modeling\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — rigorous structural and kinetic study with mutagenesis; preprint, not yet peer-reviewed\",\n      \"pmids\": [\"39763923\"],\n      \"is_preprint\": true\n    }\n  ],\n  \"current_model\": \"ASAP1 is a multi-domain scaffold protein that acts as a PIP2-dependent Arf GTPase-activating protein (primarily for Arf1/Arf5), in which the PH domain allosterically activates catalysis by directly engaging Arf·GTP at the membrane surface, the BAR domain autoinhibits GAP activity intramolecularly and independently bundles actin filaments via a lysine-rich cluster, and the SH3 domain mediates interactions with Src, FAK, Pyk2, CrkL, and other proline-rich partners; Src and Pyk2 phosphorylate ASAP1 to modulate its GAP activity and localization, while ASAP1 coordinates focal adhesion dynamics, invadopodium/podosome formation, circular dorsal ruffle generation, endocytic recycling, and ciliary receptor trafficking by integrating signals from phospholipids, Arf GTPases, Rab proteins, and the actomyosin cytoskeleton.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"ASAP1 is a multi-domain scaffold and PIP2-dependent Arf GTPase-activating protein that integrates phospholipid signals, Arf GTPases, and the actomyosin cytoskeleton to control focal adhesion dynamics, invadopodium/podosome formation, dorsal ruffle generation, endocytic recycling, and ciliary receptor trafficking [#0, #1, #14, #18]. It was originally purified as a PIP2-dependent GAP acting on Arf1 and Arf5 but not Arf6, with catalysis requiring the PH domain in concert with the Arf GAP domain [#0, #3, #7]. PIP2 binding to the PH domain is not merely a recruitment cue but an allosteric activator: cooperative binding at canonical and atypical sites drives a conformational switch that orients the PH domain to engage Arf·GTP at the membrane and accelerate hydrolysis, restricting the conformational motions of membrane-bound Arf1 to expose its catalytic regions [#2, #19, #27, #31]. The N-terminal BAR domain serves dual roles—it autoinhibits GAP activity through intramolecular contacts with the PH/GAP domains, and it independently dimerizes to bend membranes, bind and bundle actin filaments via a lysine-rich cluster (K75/K76/K79), and bind nonmuscle myosin 2A, thereby coupling membrane remodeling to actin organization [#11, #15, #22, #24, #28, #30]. GAP activity and BAR-domain actin/NM2A binding act coordinately to maintain focal adhesions and stress fibers, with Arf5 loss phenocopying ASAP1 depletion [#41]. The C-terminal SH3 domain, through a noncanonical acidic binding pocket, mediates interactions with proline-rich partners including FAK, Pyk2, and MICAL1, and Pyk2 phosphorylates ASAP1 to inhibit its GAP activity [#4, #6, #32, #36]. ASAP1 is recruited to focal adhesions by CrkL and CD2AP and nucleates a ciliary trafficking complex with Arf4, Rab11, FIP3, and Rabin8 that targets rhodopsin to the cilium [#5, #9, #14, #18, #21]. In disease contexts, ASAP1 functions as an Arf6 effector promoting NFAT1-driven invasion in uveal melanoma and participates in TGFβ/SMAD and IQGAP1/CDC42 signaling in cancers [#33, #34, #40].\",\n  \"teleology\": [\n    {\n      \"year\": 1998,\n      \"claim\": \"Established ASAP1's defining biochemical activity—answering whether it was an enzyme by showing it is a PIP2-dependent Arf GAP physically coupled to Src-family signaling.\",\n      \"evidence\": \"Protein purification, in vitro GAP assay, GST pulldown, and in-cell phosphorylation\",\n      \"pmids\": [\"9819391\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not resolve how PIP2 stimulates catalysis\", \"Substrate specificity among Arf isoforms not yet defined\"]\n    },\n    {\n      \"year\": 2000,\n      \"claim\": \"Linked the enzymatic activity to cellular function by localizing ASAP1 to focal adhesions and showing GAP activity is required for cell spreading and dorsal ruffle control.\",\n      \"evidence\": \"Fluorescence microscopy and cell spreading/PDGF assays with wild-type vs catalytically inactive mutant\",\n      \"pmids\": [\"10725410\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"GAP-independent functions hinted but not mechanistically separated\"]\n    },\n    {\n      \"year\": 2000,\n      \"claim\": \"Resolved the role of PIP2 by showing the PH domain is an allosteric activator distinct from a membrane recruitment signal.\",\n      \"evidence\": \"In vitro GAP assays, limited proteolysis, and domain deletion analysis\",\n      \"pmids\": [\"10734117\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Atomic structure of the conformational change unresolved\"]\n    },\n    {\n      \"year\": 2001,\n      \"claim\": \"Defined in vivo substrate specificity (Arf1 not Arf6) and dissected two independent downstream pathways—GAP-dependent motility versus GAP-independent spreading inhibition.\",\n      \"evidence\": \"Cell-based ARF GAP assay, stable overexpression, motility assay, GAP-domain deletion mutant\",\n      \"pmids\": [\"11773070\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Effectors of the GAP-independent spreading pathway not identified\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Identified FAK as a direct SH3-domain partner linking ASAP1 to focal adhesion assembly.\",\n      \"evidence\": \"Yeast two-hybrid, reciprocal Co-IP/pulldown, cell spreading assay with binding/GAP mutants\",\n      \"pmids\": [\"12058076\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether FAK binding regulates GAP activity not tested\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Established phospho-regulation and adapter recruitment—Pyk2 phosphorylates ASAP1 to inhibit GAP activity, and CrkL directs it to peripheral adhesions.\",\n      \"evidence\": \"Yeast two-hybrid, Co-IP, in vitro kinase assay with mapped sites, fluorimetric GTPase assay; CrkL recruitment with SH2-mutant\",\n      \"pmids\": [\"12771146\", \"12522101\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-lab findings\", \"Functional consequence of localization on Arf signaling not quantified\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Defined the catalytic mechanism as a two-step conformational model with a composite PH–GAP substrate-binding site, and validated cellular function via CD2AP-dependent recruitment.\",\n      \"evidence\": \"Saturation kinetics, FRET, limited proteolysis, AUC; CD2AP mislocalization and siRNA with Arf1 GTP-loading readout\",\n      \"pmids\": [\"16038802\", \"15632162\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Atomic interface between PH and GAP domains not yet defined\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Discovered the BAR domain as a membrane-bending module regulated by Arf1·GTP, expanding ASAP1's role into endocytic recycling of EGFR.\",\n      \"evidence\": \"In vitro membrane tubulation with EM, dimerization analysis, EGFR trafficking assay with BAR deletion\",\n      \"pmids\": [\"16431365\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo significance of tubulation for recycling not fully established\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Provided rigorous catalytic kinetics and transition-state trapping, and defined ASAP1 as a coincidence detector for invadopodium/podosome formation requiring BAR, SH3, and Src phosphorylation but not GAP activity.\",\n      \"evidence\": \"Steady-state/single-turnover kinetics with AlF4- trapping; siRNA rescue with systematic domain mutants\",\n      \"pmids\": [\"17112341\", \"17893324\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How the distinct domain inputs are integrated spatially is unresolved\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Resolved BAR autoinhibition and dynamic PH–GAP interactions, and connected ASAP1 to Rab11/FIP3-dependent recycling endosome trafficking.\",\n      \"evidence\": \"AUC, kinetics with BAR loop mutagenesis, NMR; yeast two-hybrid/Co-IP and transferrin trafficking with FIP3\",\n      \"pmids\": [\"19017632\", \"18675341\", \"18685082\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How BAR autoinhibition is relieved in cells not defined\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Identified GEFH1 as a BAR-binding negative regulator coupling Rho exchange activity to podosome assembly through ASAP1.\",\n      \"evidence\": \"Yeast two-hybrid, Co-IP, GAP assay, siRNA/overexpression\",\n      \"pmids\": [\"21352810\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab\", \"Mechanism of GAP inhibition by GEFH1 not defined\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Established ASAP1 as a scaffold for ciliary receptor targeting, assembling Arf4, Rab11, Rabin8, and rhodopsin and recognizing the rhodopsin ciliary targeting signal.\",\n      \"evidence\": \"siRNA knockdown, Co-IP, photoreceptor transport assay\",\n      \"pmids\": [\"22983554\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Order of complex assembly not yet defined\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Solved the PH domain structure revealing cooperative dual-site PIP2 binding, and showed FIP3 orchestrates the Rab11-Rabin8-Rab8 cascade by competing with rhodopsin for ASAP1.\",\n      \"evidence\": \"X-ray crystallography with mutagenesis; Co-IP competition and trafficking assays\",\n      \"pmids\": [\"26365802\", \"25673879\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Single-lab ciliary mechanism\", \"Generality beyond rhodopsin cargo not tested\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Extended ASAP1 function to immune cell migration and matrix degradation, linking it to dendritic cell motility relevant to tuberculosis susceptibility.\",\n      \"evidence\": \"siRNA knockdown in primary human dendritic cells, matrix degradation and migration assays\",\n      \"pmids\": [\"25774636\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single study in DCs\", \"Molecular pathway to migration not dissected\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Identified nonmuscle myosin 2A as a direct BAR-PH partner, positioning ASAP1 as a positive regulator of NM2A in adhesion, migration, and dorsal ruffle formation.\",\n      \"evidence\": \"In vitro binding, reciprocal Co-IP, siRNA knockdown with NM2A rescue\",\n      \"pmids\": [\"26893376\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How ASAP1 modulates NM2A contractility mechanically not defined\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Defined the BAR domain as a direct actin filament binding and bundling module dependent on a lysine cluster, and established the Arf1 N-terminal extension as the PIP2-dependent regulatory contact controlling GAP activity.\",\n      \"evidence\": \"Actin cosedimentation, domain-swap and point mutants, cell-based actin remodeling; GAP kinetics, peptide inhibition, NMR binding\",\n      \"pmids\": [\"31785555\", \"31591270\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Coupling of actin bundling to GAP catalysis in cells not resolved\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Demonstrated ASAP1's requirement for mesenchymal progenitor differentiation in vivo, linking loss to growth retardation, delayed ossification, and impaired FAK/Src and PI3K/AKT signaling.\",\n      \"evidence\": \"Gene-trap mouse, in vitro differentiation assays, Western blotting for signaling components\",\n      \"pmids\": [\"31246957\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab\", \"Direct molecular link from ASAP1 to the signaling defects not established\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Resolved the atomic-level allosteric switch by which cooperative PIP2 binding orients the PH domain for Arf1 contact, and characterized BAR-driven unipolar actin bundling and filament stabilization.\",\n      \"evidence\": \"NMR, neutron reflectometry, MD simulation; actin cosedimentation, TIRF/EM, cell-based projection assays\",\n      \"pmids\": [\"32998886\", \"32444496\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Integration of bundling activity with membrane-bound GAP function in vivo unresolved\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Showed ASAP1 regulates Mycobacterium tuberculosis uptake in macrophages through actin cytoskeleton remodeling.\",\n      \"evidence\": \"siRNA knockdown, confocal microscopy, CFU assay, F-actin staining\",\n      \"pmids\": [\"34058694\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab\", \"Direct effector linking ASAP1 to uptake not identified\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Pinpointed the lysine-rich cluster (K75/K76/K79) as the structural determinant for actin binding and bundling required for cellular stress fiber maintenance.\",\n      \"evidence\": \"Structural modeling, charge mutagenesis, cosedimentation/bundling assays, siRNA rescue in U2OS cells\",\n      \"pmids\": [\"35143843\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether the same residues affect GAP activity not tested\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Defined SH3-domain ligand recognition modes (MICAL1 PRM and ankyrin-B), establishing a noncanonical acidic binding pocket and expanding the ASAP1 partner network, and showed the PH domain restricts membrane-bound Arf1 conformational dynamics.\",\n      \"evidence\": \"X-ray crystallography with ITC and mutagenesis; NMR/neutron reflectometry/MD for Arf1 dynamics\",\n      \"pmids\": [\"36674928\", \"39265663\", \"37989735\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Functional consequence of most newly predicted SH3 partners not validated\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Placed ASAP1 in oncogenic signaling as an Arf6 effector driving NFAT1-mediated invasion and as a regulator of IQGAP1/CDC42-EGFR-MAPK signaling promoting chemoresistance.\",\n      \"evidence\": \"Co-IP, NFAT nuclear localization, ubiquitination assays, siRNA/overexpression, xenograft and tumor models\",\n      \"pmids\": [\"37500798\", \"36792578\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-lab cancer mechanisms\", \"Direct enzymatic role of ASAP1 in these pathways unclear\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Established that ASAP1 protein levels are controlled by SCF-FBXW7/CUL1-mediated ubiquitination affecting osteogenesis, and connected ASAP1 to neuronal tau seed internalization via A1AR/MSUT2.\",\n      \"evidence\": \"Co-IP, ubiquitination assays, osteogenesis and osteoporosis models; tau seeding assays with A1AR modulation\",\n      \"pmids\": [\"39287759\", \"38472475\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-lab findings\", \"Mechanism of ASAP1 activity modulation by A1AR not defined\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Demonstrated coordinate requirement of GAP and BAR-domain activities for adhesion/stress fiber maintenance, identified additional cytoskeletal partners (MYO1F), and placed ASAP1 in differentiation and EMT pathways (TAZ inactivation, TGFβ/SMAD).\",\n      \"evidence\": \"Domain-rescue siRNA in multiple cell lines with Arf5 epistasis; proximity proteomics; MEKi epistasis with WWTR1; Co-IP with SMAD2/3\",\n      \"pmids\": [\"40194952\", \"41208482\", \"39495123\", \"40742091\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-lab pathway placements\", \"Direct molecular links between ASAP1 enzymatic activity and transcriptional outputs unresolved\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How ASAP1's distinct domain activities—GAP catalysis, BAR membrane bending and actin bundling, and SH3 scaffolding—are spatially and temporally coordinated within single structures (focal adhesions, podosomes, cilia) in living cells remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No integrated model of in-cell domain coordination\", \"Physiological triggers relieving BAR autoinhibition unknown\", \"In vivo roles of most cancer/neuronal pathways not validated genetically\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0, 3, 6, 7, 12]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [0, 6, 14, 17]},\n      {\"term_id\": \"GO:0008289\", \"supporting_discovery_ids\": [2, 11, 19, 27]},\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [24, 28, 30, 22]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [5, 9, 18, 36]},\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [11, 28]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [1, 24, 28]},\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [1, 11, 13]},\n      {\"term_id\": \"GO:0005768\", \"supporting_discovery_ids\": [11, 14]},\n      {\"term_id\": \"GO:0005929\", \"supporting_discovery_ids\": [18, 21]},\n      {\"term_id\": \"GO:0005815\", \"supporting_discovery_ids\": [14]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-5653656\", \"supporting_discovery_ids\": [11, 14, 18]},\n      {\"term_id\": \"R-HSA-1474244\", \"supporting_discovery_ids\": [1, 4, 13, 20]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [0, 6, 33]},\n      {\"term_id\": \"R-HSA-9609507\", \"supporting_discovery_ids\": [18, 21]}\n    ],\n    \"complexes\": [\n      \"ASAP1-FIP3-Rab11 ternary complex\",\n      \"ciliary targeting complex (Arf4-Rab11-Rabin8-rhodopsin)\"\n    ],\n    \"partners\": [\n      \"FAK\",\n      \"Pyk2\",\n      \"CrkL\",\n      \"CD2AP\",\n      \"FIP3\",\n      \"NM2A\",\n      \"MICAL1\",\n      \"ARF1\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":8,"faith_total":8,"faith_pct":100.0}}