{"gene":"MICAL1","run_date":"2026-06-10T02:59:50","timeline":{"discoveries":[{"year":2011,"finding":"Mical directly oxidizes the methionine 44 (Met44) residue within the D-loop of actin filament subunits, simultaneously severing filaments and decreasing polymerization. This post-translational oxidation is the biochemical mechanism underlying Mical-mediated actin filament disassembly.","method":"In vitro biochemical assays with purified Mical protein and actin; site-directed mutagenesis; mass spectrometry identification of Met44 oxidation","journal":"Science","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro reconstitution with purified proteins, site-directed mutagenesis, mass spectrometry, replicated across multiple papers","pmids":["22116028"],"is_preprint":false},{"year":2010,"finding":"Mical directly binds F-actin and disassembles both individual and bundled actin filaments using its redox enzymatic activity. Mical is both necessary and sufficient for semaphorin-plexin-mediated F-actin reorganization in vivo, linking plexin receptors to actin dynamics.","method":"Purified protein binding and disassembly assays in vitro; genetic loss-of-function and gain-of-function in Drosophila; pharmacological inhibition of redox activity","journal":"Nature","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — purified protein biochemistry, in vivo genetics, multiple orthogonal methods, replicated","pmids":["20148037"],"is_preprint":false},{"year":2013,"finding":"The methionine sulfoxide reductase SelR (MsrB) specifically reduces Mical-oxidized actin (Met44-R-sulfoxide) back to methionine, restoring normal actin polymerization properties, establishing a reversible redox actin regulatory system. SelR opposes Mical activity and semaphorin-plexin repulsion in vivo.","method":"Genetic epistasis in Drosophila; in vitro enzymatic reduction assays with purified SelR and Mical-oxidized actin; stereospecificity analysis","journal":"Nature Cell Biology","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — in vitro reconstitution, genetic epistasis, stereospecificity established, multiple methods","pmids":["24212093"],"is_preprint":false},{"year":2005,"finding":"The N-terminal region of MICAL-1 is a FAD-containing flavoprotein monooxygenase domain structurally similar to p-hydroxybenzoate hydroxylase (PHBH). The flavin ring can switch between two discrete positions coupled to opening of a channel to the active site, suggesting a protein substrate. Crystal structure resolved at 1.45 Å.","method":"X-ray crystallography at 1.45 Å resolution of mouse MICAL-1 residues 1–489","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1 / Strong — high-resolution crystal structure with functional validation of conformational switch","pmids":["16275925"],"is_preprint":false},{"year":2005,"finding":"The N-terminal flavoenzyme domain of MICAL performs an NADPH-dependent reaction reducing molecular oxygen to H2O2 (Km,NADPH = 222 μM; kcat = 77 sec−1). H2O2 production was proposed as a signaling molecule in axon guidance.","method":"Biochemical enzyme kinetics assay with purified MICAL flavodomain; measurement of H2O2 production","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro enzymatic reconstitution with kinetic characterization, single lab","pmids":["16275926"],"is_preprint":false},{"year":2008,"finding":"MICAL enzymatic activity is autoinhibited by its C-terminal domain. CRMP (collapsin response mediator protein) and Plexin physically associate with non-enzymatic and enzymatic domains of MICAL respectively, and together release MICAL enzymatic autoinhibition. Semaphorin signaling promotes the CRMP–MICAL physical association.","method":"Co-immunoprecipitation; domain mapping; enzyme activity assays with truncation and domain mutants; cell-based assays","journal":"The Journal of Neuroscience","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — reciprocal co-IP and activity assays, single lab, two orthogonal methods","pmids":["18305261"],"is_preprint":false},{"year":2011,"finding":"MICAL-1 interacts with the hydrophobic motif of NDR1/2 kinases and negatively regulates MST-NDR kinase signaling by competing with MST1 for NDR binding, thereby inhibiting NDR-dependent proapoptotic signaling. Overexpression of MICAL-1 reduces NDR activation; knockdown augments it.","method":"Proteomics/mass spectrometry screen; co-immunoprecipitation; kinase activity assays; siRNA knockdown and overexpression with apoptosis readout","journal":"Molecular and Cellular Biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — proteomic identification, co-IP, enzymatic assay, single lab with multiple orthogonal methods","pmids":["21730291"],"is_preprint":false},{"year":2012,"finding":"Human MICAL1 is autoinhibited by its C-terminal coiled-coil region. All three human MICALs (MICAL1, MICAL2, MICAL3) regulate actin stress fibers in non-neural cells and require ROS generation for their actin-regulatory function.","method":"siRNA knockdown; overexpression of truncation mutants; fluorescence microscopy of actin structures; ROS measurement","journal":"Journal of Cell Science","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — KD/OE with defined phenotype, domain mapping, single lab","pmids":["22331357"],"is_preprint":false},{"year":2016,"finding":"Mical-mediated oxidation of actin (Met44/Met47) improves cofilin binding to filaments, and the combined effect dramatically accelerates F-actin disassembly compared to either alone. This synergism is necessary and sufficient for F-actin disassembly in vivo and magnifies effects on axon guidance and semaphorin-plexin repulsion.","method":"In vitro actin disassembly assays with purified Mical and cofilin; TIRF microscopy; Drosophila genetic assays","journal":"Nature Cell Biology","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — in vitro reconstitution, in vivo genetics, TIRF imaging, multiple orthogonal methods","pmids":["27454820"],"is_preprint":false},{"year":2017,"finding":"Mical stereospecifically oxidizes actin M44 and M47 residues. CryoEM reconstruction at 3.9 Å reveals oxidation reorients the M44 side chain and induces a new intermolecular interaction M47-O-T351 that promotes Mox-actin instability. Mox-actin can undergo extremely fast disassembly (84 subunits/s) and allows cofilin-mediated severing even in the presence of inorganic phosphate.","method":"Near-atomic resolution cryo-EM; single filament TIRF microscopy; site-directed mutagenesis","journal":"Nature Communications","confidence":"High","confidence_rationale":"Tier 1 / Strong — cryo-EM structure, mutagenesis, single-filament imaging, multiple orthogonal methods","pmids":["29259197"],"is_preprint":false},{"year":2021,"finding":"MICAL1-induced oxidation of actin filaments suppresses their protection from cofilin by: increasing cofilin binding and severing rates by several orders of magnitude; allowing non-activated phosphomimetic S3D-cofilin to bind and sever oxidized filaments; and causing tropomyosin Tpm1.8 to lose its ability to protect filaments from cofilin severing.","method":"Single filament experiments in vitro; fluorescence microscopy; TIRF assays with purified proteins","journal":"EMBO Reports","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution with purified proteins, single-filament imaging, multiple conditions tested","pmids":["33393173"],"is_preprint":false},{"year":2002,"finding":"MICAL1 was identified as a novel protein interacting with the CasL SH3 domain via its PPKPP proline-rich sequence. MICAL1 also associates with vimentin intermediate filaments through its C-terminal region, and colocalizes with vimentin and CasL at the perinuclear area.","method":"Far Western screening; co-immunoprecipitation; immunofluorescence colocalization","journal":"The Journal of Biological Chemistry","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — co-IP and pulldown, immunofluorescence, single lab, two orthogonal methods","pmids":["11827972"],"is_preprint":false},{"year":2003,"finding":"MICAL-1 (isoforms 1a and 1b) specifically interacts with active GTP-bound Rab1 in a nucleotide-dependent manner, with the interaction domain mapped to the C-terminus of MICAL-1, which also mediates binding to vimentin. MICAL-1 displays predominantly cytosolic localization by cell fractionation.","method":"Yeast two-hybrid; GST pulldown; cell fractionation; domain mapping","journal":"Biochemical and Biophysical Research Communications","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — yeast two-hybrid confirmed by GST pulldown, cell fractionation, single lab","pmids":["12788069"],"is_preprint":false},{"year":2014,"finding":"Vertebrate MICAL-1 regulates the targeting of secretory vesicles containing IgCAM cell adhesion molecules to the neuronal growth cone membrane through its ability to control the actin cytoskeleton via redox chemistry, thereby maintaining appropriate IgCAM cell surface levels. This is required for lamina-specific mossy fibre axon targeting in the mouse hippocampus in vivo.","method":"MICAL-1 knockout mouse; live imaging; cell surface biotinylation assays; immunofluorescence","journal":"Nature Communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — knockout mouse with defined in vivo phenotype, cell biological assays, multiple methods","pmids":["25007825"],"is_preprint":false},{"year":2016,"finding":"Human MICAL1 catalytic properties are regulated by its CH, LIM, and C-terminal domains. The C-terminal domain causes ~10-fold decrease in kcat, establishing an inactive/active conformational equilibrium (autoinhibition). F-actin lowers Km,NADPH and increases kcat for all MICAL forms, and actin depolymerization may be mediated by H2O2 produced by the NADPH oxidase reaction rather than direct actin methionine hydroxylation.","method":"Enzyme kinetics with purified full-length and truncated MICAL1 forms; NADPH oxidase assays; F-actin depolymerization assays","journal":"Archives of Biochemistry and Biophysics","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — in vitro enzyme kinetics with multiple truncation mutants, single lab","pmids":["26845023"],"is_preprint":false},{"year":2018,"finding":"Rab8 in its active GTP-bound state binds MICAL1 and stabilizes its active conformation, causing a specific ~4-fold increase in kcat of the NADPH oxidase reaction. SAXS measurements support a 1:1 complex between full-length MICAL1 and Rab8 with Kd ~8 μM. Free full-length MICAL1 adopts an autoinhibited conformation in which the C-terminal region interferes with catalytic conformational changes.","method":"Enzyme kinetics; small-angle X-ray scattering (SAXS); SAXS-based structural modeling","journal":"Protein Science","confidence":"Medium","confidence_rationale":"Tier 1–2 / Moderate — kinetics + SAXS structure, single lab, two orthogonal methods","pmids":["30242933"],"is_preprint":false},{"year":2024,"finding":"Cryo-EM structure of human MICAL1 at 3.1 Å resolution reveals that autoinhibition is mediated by an intramolecular interaction between the N-terminal catalytic domain and C-terminal coiled-coil domain, blocking F-actin interaction. Allosteric changes in the coiled-coil domain and binding of the tripartite CH-L2α1-LIM assembly to the coiled-coil domain are required for MICAL1 activation.","method":"Cryo-EM structure determination; biochemical binding assays; functional cell-based assays; mutagenesis","journal":"Nature Communications","confidence":"High","confidence_rationale":"Tier 1 / Strong — cryo-EM structure at 3.1 Å, biochemical validation, mutagenesis, multiple orthogonal methods","pmids":["39532862"],"is_preprint":false},{"year":2020,"finding":"MICAL1 oxidizes Met308, a conserved residue in the calmodulin-binding domain of CaMKII, reducing CaM binding and CaMKII activity. Loss of MICAL1 in mice causes cardiac arrhythmias and premature death due to CaMKII hyperactivation, establishing CaMKII as a non-actin substrate of MICAL1.","method":"MICAL1 knockout mouse model; biochemical assays for CaMKII activity and CaM binding; site-specific mutagenesis; Drosophila genetic model; human iPSC-derived cardiomyocytes","journal":"The Journal of Clinical Investigation","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — KO mouse with defined phenotype, biochemical assays, mutagenesis, multiple species validation","pmids":["32749237"],"is_preprint":false},{"year":2022,"finding":"PAK1 (a CDC42 GTPase effector) associates with MICAL1 via its N-terminal catalytic monooxygenase and calponin homology domains, and phosphorylates MICAL1 on two serine residues, leading to accelerated F-actin disassembly. Extracellular ligand stimulation leads to PAK-dependent MICAL1 phosphorylation, linking external signals to MICAL1 activation.","method":"Co-immunoprecipitation; mass spectrometry phosphoproteomics; in vitro kinase assays; F-actin disassembly assays","journal":"Cell Reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP, kinase assay, MS phosphoproteomics, single lab with multiple methods","pmids":["36198272"],"is_preprint":false},{"year":2021,"finding":"The unconventional myosin Myo15 physically interacts with Mical and uses its motor and MyTH4-FERM domains to broaden Mical's distribution, thereby spatiotemporally positioning and expanding Mical-mediated F-actin disassembly and cellular remodeling in response to Semaphorin/Plexin signals.","method":"Protein-protein interaction assays; Drosophila genetic epistasis; imaging of cellular remodeling","journal":"Science Advances","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — genetic and biochemical interaction data, single lab, two methods","pmids":["33980493"],"is_preprint":false},{"year":2018,"finding":"Two MICAL-1 mutations causing autosomal dominant lateral temporal epilepsy (p.Gly150Ser in the monooxygenase domain and p.Ala1065fs frameshift in the C-terminal domain) both significantly increased MICAL-1 oxidoreductase activity and induced cell contraction in COS7 cells, demonstrating gain-of-function effects on MICAL-1 enzymatic activity leading to dysregulation of F-actin dynamics.","method":"Cell-based oxidoreductase activity assays; cell contraction assay in COS7 cells; whole exome sequencing","journal":"Annals of Neurology","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — cell-based enzymatic assay with patient mutations, single lab","pmids":["29394500"],"is_preprint":false},{"year":2022,"finding":"Mical oxidizes Tau at Cys322 via its monooxygenase domain, altering Tau interactions with microtubules and the actin cytoskeleton, and greatly affecting Tau aggregation propensity and Tau-associated toxicity in a Drosophila Tauopathy model. MICAL1 co-localizes with Tau in Pick bodies in human Tauopathy samples.","method":"Drosophila genetic interaction studies; Mical inhibitor treatment; monooxygenase domain-specific mutation; Tau cysteine mutant transgenes; mass spectrometry quantification of cysteine oxidation","journal":"Acta Neuropathologica Communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic interactions, domain-specific mutation, MS validation, single lab","pmids":["35379354"],"is_preprint":false},{"year":2024,"finding":"MICAL1 locally depolymerizes cortical actin at HIV-1 budding sites to promote viral budding and release. MICAL1 directly disassembles branched actin networks. The Rab35 activator of MICAL1 is also recruited at budding sites and functions in the same pathway. MICAL1 depletion prevents timely ESCRT scission machinery recruitment.","method":"Superresolution microscopy; MICAL1 siRNA depletion; F-actin disassembly assays; in vitro branched-actin network assays; ESCRT recruitment imaging","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — KD with defined phenotype, in vitro actin assay, imaging, single lab","pmids":["39556735"],"is_preprint":false},{"year":2025,"finding":"MICAL1 is a shear-activated regulator that promotes local F-actin disassembly around the GPIb-IX-V complex in platelets, enabling its translocation to lipid rafts and reinforcing VWF binding under high-shear conditions. MICAL1-deficient platelets display impaired VWF-GPIbα adhesion, increased deformability under shear, and defective thrombus formation in vivo.","method":"MICAL1-knockout mice; microfluidics shear assay; TIRF/superresolution microscopy; lipid raft fractionation; in vivo thrombosis model","journal":"Nature Communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — KO mouse, biochemical fractionation, microfluidics, multiple methods, single lab","pmids":["40783397"],"is_preprint":false},{"year":2023,"finding":"F-actin bundled with fascin is extensively disassembled by Mical; moreover, Mical-oxidized actin is poorly bundled by fascin, amplifying disassembly. Cofilin synergizes with Mical to dramatically amplify disassembly of bundled F-actin beyond the sum of their individual effects. Fascin bundling dampens Mical's disassembly activity, establishing a balance between crosslinking and oxidative disassembly.","method":"Biochemical actin bundling/disassembly assays with purified proteins; structural imaging; Drosophila genetic epistasis assays; high-resolution cellular imaging","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — in vitro reconstitution, structural assays, in vivo genetics, multiple orthogonal methods","pmids":["37725655"],"is_preprint":false},{"year":2023,"finding":"The SH3 domain of ASAP1 binds to the proline-rich motif (PRM) of MICAL1 with sub-μM affinity. Crystal structure of the ASAP1-SH3/MICAL1-PRM complex reveals a unique binding mode where two negatively charged patches in the SH3 pocket recognize an 'xPx+Px+' sequence in MICAL1-PRM, differing from typical SH3/PRM interactions.","method":"Crystal structure determination; isothermal titration calorimetry (ITC); mutagenesis","journal":"International Journal of Molecular Sciences","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — crystal structure with binding affinity measurement, single lab","pmids":["36674928"],"is_preprint":false},{"year":2028,"finding":"MICAL1 co-immunoprecipitates with RAB35, and activated RAB35 (GTP-bound) increases MICAL1 interaction. Silencing of RAB35 reduced ROS generation and Akt phosphorylation downstream of MICAL1 in breast cancer cells.","method":"Co-immunoprecipitation; Rac1 pulldown assay; ROS measurement; siRNA knockdown","journal":"BMC Cancer","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single co-IP, single lab, no direct enzymatic characterization of RAB35-MICAL1 interaction","pmids":["27430308"],"is_preprint":false},{"year":2019,"finding":"NEDD9 interacts with MICAL1 under hypoxic conditions, and NEDD9 stabilizes MICAL1 protein. Silencing NEDD9 reduced MICAL1 stability. MICAL1 overexpression selectively increased GTP-Rac1 (but not Cdc42 or RhoA), implicating MICAL1 in Rac1-dependent migration.","method":"Co-immunoprecipitation; siRNA knockdown; Rac1/Cdc42/RhoA pulldown activity assay","journal":"Frontiers in Pharmacology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single co-IP, single lab, limited mechanistic follow-up","pmids":["31019460"],"is_preprint":false},{"year":2024,"finding":"PlexinA1 forms a complex with MICAL1 and promotes Rac1 activation and ROS production, which in turn prevents MICAL1 ubiquitin/proteasome-mediated degradation and stabilizes MICAL1 protein.","method":"Co-immunoprecipitation; immunofluorescence; siRNA-mediated knockdown; Rac1 pulldown assay; ROS measurement; ubiquitination assay","journal":"Biochimica et Biophysica Acta. Molecular Basis of Disease","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single co-IP, single lab, no direct in vitro reconstitution of the regulatory pathway","pmids":["38508474"],"is_preprint":false},{"year":2009,"finding":"Sox14 transcription factor directly regulates expression of mical in Drosophila, establishing a genetic pathway where Sox14 → Mical governs dendrite severing during pruning. Overexpression of Mical significantly rescues pruning defects in sox14 mutants.","method":"Genetic epistasis; loss-of-function and gain-of-function in Drosophila; chromatin immunoprecipitation (Sox14 binding to mical promoter)","journal":"Nature Neuroscience","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis with rescue experiment, transcriptional target validation, single lab","pmids":["19881505"],"is_preprint":false}],"current_model":"MICAL1 is a multidomain FAD-containing flavoprotein monooxygenase that, upon activation by semaphorin-plexin signaling or upstream regulators (including RAB8, PAK1, and RAB35), uses NADPH to stereospecifically oxidize methionine residues 44 and 47 of actin filaments, causing severing, depolymerization, and reduced reassembly; this activity is autoinhibited by intramolecular interactions between its N-terminal catalytic domain and C-terminal coiled-coil domain (as resolved by cryo-EM), is synergized by cofilin, is reversed by the methionine sulfoxide reductase SelR/MsrB, and extends beyond actin to substrates including CaMKII (M308 oxidation) and Tau (C322 oxidation), with roles in axon guidance, cytokinesis, platelet function, cardiac stress responses, and epilepsy."},"narrative":{"mechanistic_narrative":"MICAL1 is a multidomain FAD-dependent flavoprotein monooxygenase that couples redox chemistry to actin cytoskeleton remodeling, serving as the effector that links semaphorin-plexin signaling to F-actin reorganization in vivo [PMID:20148037, PMID:16275925]. Its N-terminal flavoenzyme domain is structurally related to p-hydroxybenzoate hydroxylase and uses NADPH and molecular oxygen to drive catalysis, with a flavin ring that switches between positions coupled to opening of a substrate channel [PMID:16275925, PMID:16275926]. The enzyme directly binds and disassembles individual and bundled F-actin by stereospecifically oxidizing actin methionine residues Met44 and Met47; this post-translational oxidation reorients the Met44 side chain and induces a new intermolecular contact that destabilizes the filament, simultaneously severing it and blocking reassembly [PMID:22116028, PMID:29259197]. Oxidation is reversed by the methionine sulfoxide reductase SelR/MsrB, which restores normal polymerization and establishes a reversible redox switch on actin [PMID:24212093]. MICAL1-mediated oxidation also primes filaments for accelerated cofilin binding and severing — overriding the protection normally conferred by tropomyosin and fascin bundling — so that cofilin synergizes with MICAL1 to dramatically amplify disassembly [PMID:27454820, PMID:33393173, PMID:37725655]. MICAL1 catalytic activity is held in an autoinhibited state by an intramolecular interaction between the N-terminal catalytic domain and the C-terminal coiled-coil region, resolved by cryo-EM, and is relieved by upstream regulators including GTP-bound Rab8, PAK1-mediated phosphorylation, and CRMP/Plexin association downstream of semaphorin signaling [PMID:18305261, PMID:30242933, PMID:39532862, PMID:36198272]. Its substrate range extends beyond actin: MICAL1 oxidizes Met308 of CaMKII to reduce calmodulin binding and kinase activity, and oxidizes Cys322 of Tau to alter cytoskeletal interactions and aggregation [PMID:32749237, PMID:35379354]. Through these activities MICAL1 controls axon guidance and growth-cone IgCAM trafficking, dendrite pruning, cytokinesis-related membrane events, HIV-1 budding, shear-dependent platelet adhesion, and cardiac CaMKII homeostasis [PMID:25007825, PMID:32749237, PMID:39556735, PMID:40783397, PMID:19881505]. Gain-of-function MICAL1 mutations that elevate its oxidoreductase activity cause autosomal dominant lateral temporal epilepsy [PMID:29394500].","teleology":[{"year":2002,"claim":"Established the first physical partners of MICAL1, placing it at cytoskeletal adaptor junctions before its enzymatic role was known.","evidence":"Far Western screening, co-IP, and immunofluorescence in cells identifying CasL SH3 and vimentin interactions","pmids":["11827972"],"confidence":"Medium","gaps":["No enzymatic function assigned at this stage","Functional consequence of vimentin/CasL binding unresolved"]},{"year":2003,"claim":"Linked MICAL1 to Rab GTPase signaling by showing nucleotide-dependent binding to active Rab1, implicating it in vesicle-related processes.","evidence":"Yeast two-hybrid, GST pulldown, and cell fractionation with domain mapping","pmids":["12788069"],"confidence":"Medium","gaps":["No catalytic consequence of Rab1 binding tested","Cytosolic localization not connected to a pathway"]},{"year":2005,"claim":"Defined MICAL1 as a FAD monooxygenase with a PHBH-like fold and a conformational flavin switch, and showed it produces H2O2 from NADPH/O2, framing the enzymatic basis of its cytoskeletal effects.","evidence":"1.45 Å X-ray crystallography of the mouse flavodomain plus enzyme kinetics measuring H2O2 production","pmids":["16275925","16275926"],"confidence":"High","gaps":["Physiological protein substrate not yet identified","Whether H2O2 or direct oxidation drives actin disassembly unresolved"]},{"year":2008,"claim":"Resolved how MICAL is switched on, showing C-terminal autoinhibition relieved by CRMP and Plexin binding downstream of semaphorin signaling.","evidence":"Co-IP, domain mapping, and activity assays with truncation mutants in cells","pmids":["18305261"],"confidence":"Medium","gaps":["Structural basis of autoinhibition not defined","Single lab"]},{"year":2009,"claim":"Placed Mical in a transcriptional pathway for developmental remodeling by identifying Sox14 as a direct upstream activator driving dendrite pruning.","evidence":"Drosophila genetic epistasis, rescue, and ChIP of Sox14 at the mical promoter","pmids":["19881505"],"confidence":"Medium","gaps":["Vertebrate transcriptional regulation not addressed","Downstream actin events inferred from genetics"]},{"year":2011,"claim":"Identified the core catalytic mechanism — direct oxidation of actin Met44 that simultaneously severs and depolymerizes filaments — defining MICAL as an actin-modifying enzyme.","evidence":"In vitro biochemistry with purified protein, mutagenesis, and mass spectrometry","pmids":["22116028"],"confidence":"High","gaps":["Reversibility of the modification not yet shown","Additional oxidized residues not yet mapped"]},{"year":2011,"claim":"Expanded MICAL1's reach beyond the cytoskeleton by showing it competes with MST1 for NDR kinase binding to suppress proapoptotic signaling.","evidence":"Proteomic screen, co-IP, kinase assays, and knockdown/overexpression apoptosis readouts","pmids":["21730291"],"confidence":"Medium","gaps":["Redox activity not required for the scaffold function","Single lab"]},{"year":2013,"claim":"Established MICAL-driven actin oxidation as a reversible redox switch by identifying SelR/MsrB as the stereospecific reductase opposing MICAL in vivo.","evidence":"Drosophila genetic epistasis and in vitro stereospecific reduction of Mical-oxidized actin","pmids":["24212093"],"confidence":"High","gaps":["Regulation of the MICAL/SelR balance in vivo not detailed"]},{"year":2014,"claim":"Connected MICAL1 redox actin control to growth-cone biology, showing it governs IgCAM vesicle targeting and lamina-specific axon targeting in mouse.","evidence":"MICAL1 knockout mouse, surface biotinylation, and live imaging","pmids":["25007825"],"confidence":"High","gaps":["Direct vesicle-actin coupling mechanism not fully resolved"]},{"year":2016,"claim":"Demonstrated cofilin synergy with MICAL oxidation and refined the residues to Met44/Met47, explaining how oxidation potentiates accelerated disassembly.","evidence":"In vitro disassembly assays, TIRF microscopy, and Drosophila genetics; plus kinetic dissection of domain autoinhibition and the H2O2-versus-direct-oxidation question","pmids":["27454820","26845023"],"confidence":"High","gaps":["Relative contributions of direct oxidation versus H2O2 still debated between studies","Mechanism of cofilin enhancement structurally undefined at the time"]},{"year":2017,"claim":"Provided near-atomic structural mechanism for how oxidation destabilizes filaments via Met44 reorientation and a new M47-O-T351 contact enabling ultrafast disassembly.","evidence":"3.9 Å cryo-EM, single-filament TIRF, and mutagenesis","pmids":["29259197"],"confidence":"High","gaps":["Structure of the full autoinhibited enzyme not yet solved"]},{"year":2018,"claim":"Showed how upstream signals activate MICAL1, with GTP-Rab8 stabilizing the active conformation and relieving C-terminal autoinhibition.","evidence":"Enzyme kinetics and SAXS modeling of the 1:1 MICAL1-Rab8 complex; epilepsy mutations shown to be enzymatic gain-of-function","pmids":["30242933","29394500"],"confidence":"Medium","gaps":["SAXS provides low-resolution architecture only","Mechanism connecting elevated activity to seizures untested in neurons"]},{"year":2020,"claim":"Established a non-actin substrate, showing MICAL1 oxidizes CaMKII Met308 to limit kinase activity and protect against cardiac arrhythmia.","evidence":"MICAL1 knockout mouse, CaMKII biochemistry, mutagenesis, Drosophila and iPSC-cardiomyocyte models","pmids":["32749237"],"confidence":"High","gaps":["Spatial regulation of MICAL1-CaMKII oxidation in cardiomyocytes not mapped"]},{"year":2021,"claim":"Mechanistically explained how oxidation overrides filament protection, allowing even phosphomimetic cofilin to sever and stripping tropomyosin protection.","evidence":"Single-filament TIRF assays with purified proteins; plus identification of Myo15 as a spatial positioner of Mical activity","pmids":["33393173","33980493"],"confidence":"High","gaps":["In vivo significance of tropomyosin override not directly tested","Myo15-Mical coupling shown in Drosophila only"]},{"year":2022,"claim":"Identified PAK1 phosphorylation as an extracellular-signal-driven activation input and added Tau Cys322 as another oxidation substrate relevant to tauopathy.","evidence":"Co-IP, phosphoproteomics, kinase and disassembly assays; Drosophila tauopathy genetics with MS of cysteine oxidation","pmids":["36198272","35379354"],"confidence":"Medium","gaps":["PAK1 phospho-sites not structurally placed in the autoinhibition model","Human relevance of Tau oxidation beyond colocalization unestablished"]},{"year":2023,"claim":"Defined the interplay between bundling and oxidative disassembly, showing fascin dampens while cofilin amplifies MICAL action, and characterized a distinct ASAP1-SH3/MICAL1-PRM binding mode.","evidence":"In vitro bundling/disassembly assays and Drosophila genetics; crystal structure and ITC of the ASAP1-SH3/MICAL1-PRM complex","pmids":["37725655","36674928"],"confidence":"High","gaps":["Functional consequence of ASAP1 binding to MICAL1 in cells not defined"]},{"year":2024,"claim":"Solved the human autoinhibited enzyme structure, defining the N-terminal catalytic / C-terminal coiled-coil intramolecular lock and the tripartite CH-L2α1-LIM assembly required for activation.","evidence":"3.1 Å cryo-EM with biochemical binding assays, mutagenesis, and cell-based assays; plus role in HIV-1 budding via cortical actin depolymerization","pmids":["39532862","39556735"],"confidence":"High","gaps":["How each upstream regulator engages the activation assembly not fully mapped","Rab35 recruitment mechanism at budding sites incompletely defined"]},{"year":2025,"claim":"Extended MICAL1 function to mechanotransduction in platelets, showing shear-activated F-actin disassembly enables GPIb-IX-V translocation to lipid rafts and stable VWF adhesion.","evidence":"MICAL1 knockout mice, microfluidics shear assays, superresolution imaging, lipid raft fractionation, and in vivo thrombosis","pmids":["40783397"],"confidence":"Medium","gaps":["Molecular sensor coupling shear to MICAL1 activation unidentified","Single lab"]},{"year":null,"claim":"How the multiple activation inputs (Rab8, Rab35, PAK1, CRMP/Plexin, NEDD9, PlexinA1) are integrated to spatially and temporally tune MICAL1 oxidative activity, and how substrate selection between actin, CaMKII, and Tau is governed, remain unresolved.","evidence":"","pmids":[],"confidence":"Low","gaps":["No unified model of competitive/cooperative activation","Substrate-selectivity determinants unknown","Several Rac1/ROS regulatory links rest on single co-IP studies"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0016491","term_label":"oxidoreductase activity","supporting_discovery_ids":[0,3,4,9,14,17,21]},{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0,9,17,21]},{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[1,8,10,24]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[6]}],"localization":[{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[12]},{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[1,7,11]},{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[22,23]}],"pathway":[{"term_id":"R-HSA-1266738","term_label":"Developmental Biology","supporting_discovery_ids":[1,13,29]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[5,15,18]},{"term_id":"R-HSA-109582","term_label":"Hemostasis","supporting_discovery_ids":[23]}],"complexes":[],"partners":["RAB8","PAK1","CRMP","PLXNA1","NDR1","CASL","ASAP1","RAB1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q8TDZ2","full_name":"[F-actin]-monooxygenase MICAL1","aliases":["Molecule interacting with CasL protein 1","MICAL-1","NEDD9-interacting protein with calponin homology and LIM domains"],"length_aa":1067,"mass_kda":117.9,"function":"Monooxygenase that promotes depolymerization of F-actin by mediating oxidation of specific methionine residues on actin to form methionine-sulfoxide, resulting in actin filament disassembly and preventing repolymerization (PubMed:29343822). In the absence of actin, it also functions as a NADPH oxidase producing H(2)O(2) (PubMed:21864500, PubMed:26845023, PubMed:29343822). Acts as a cytoskeletal regulator that connects NEDD9 to intermediate filaments. Also acts as a negative regulator of apoptosis via its interaction with STK38 and STK38L; acts by antagonizing STK38 and STK38L activation by MST1/STK4. Involved in regulation of lamina-specific connectivity in the nervous system such as the development of lamina-restricted hippocampal connections. Through redox regulation of the actin cytoskeleton controls the intracellular distribution of secretory vesicles containing L1/neurofascin/NgCAM family proteins in neurons, thereby regulating their cell surface levels (By similarity). May act as Rab effector protein and play a role in vesicle trafficking. Promotes endosomal tubule extension by associating with RAB8 (RAB8A or RAB8B), RAB10 and GRAF (GRAF1/ARHGAP26 or GRAF2/ARHGAP10) on the endosomal membrane which may connect GRAFs to Rabs, thereby participating in neosynthesized Rab8-Rab10-Rab11-dependent protein export (PubMed:32344433)","subcellular_location":"Cytoplasm; Cytoplasm, cytoskeleton; Endosome membrane; Midbody","url":"https://www.uniprot.org/uniprotkb/Q8TDZ2/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/MICAL1","classification":"Not Classified","n_dependent_lines":1,"n_total_lines":1208,"dependency_fraction":0.0008278145695364238},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/MICAL1","total_profiled":1310},"omim":[{"mim_id":"608882","title":"MICROTUBULE-ASSOCIATED MONOOXYGENASE, CALPONIN AND LIM DOMAINS-CONTAINING, 3; 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This post-translational oxidation is the biochemical mechanism underlying Mical-mediated actin filament disassembly.\",\n      \"method\": \"In vitro biochemical assays with purified Mical protein and actin; site-directed mutagenesis; mass spectrometry identification of Met44 oxidation\",\n      \"journal\": \"Science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro reconstitution with purified proteins, site-directed mutagenesis, mass spectrometry, replicated across multiple papers\",\n      \"pmids\": [\"22116028\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Mical directly binds F-actin and disassembles both individual and bundled actin filaments using its redox enzymatic activity. Mical is both necessary and sufficient for semaphorin-plexin-mediated F-actin reorganization in vivo, linking plexin receptors to actin dynamics.\",\n      \"method\": \"Purified protein binding and disassembly assays in vitro; genetic loss-of-function and gain-of-function in Drosophila; pharmacological inhibition of redox activity\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — purified protein biochemistry, in vivo genetics, multiple orthogonal methods, replicated\",\n      \"pmids\": [\"20148037\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"The methionine sulfoxide reductase SelR (MsrB) specifically reduces Mical-oxidized actin (Met44-R-sulfoxide) back to methionine, restoring normal actin polymerization properties, establishing a reversible redox actin regulatory system. SelR opposes Mical activity and semaphorin-plexin repulsion in vivo.\",\n      \"method\": \"Genetic epistasis in Drosophila; in vitro enzymatic reduction assays with purified SelR and Mical-oxidized actin; stereospecificity analysis\",\n      \"journal\": \"Nature Cell Biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — in vitro reconstitution, genetic epistasis, stereospecificity established, multiple methods\",\n      \"pmids\": [\"24212093\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"The N-terminal region of MICAL-1 is a FAD-containing flavoprotein monooxygenase domain structurally similar to p-hydroxybenzoate hydroxylase (PHBH). The flavin ring can switch between two discrete positions coupled to opening of a channel to the active site, suggesting a protein substrate. Crystal structure resolved at 1.45 Å.\",\n      \"method\": \"X-ray crystallography at 1.45 Å resolution of mouse MICAL-1 residues 1–489\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — high-resolution crystal structure with functional validation of conformational switch\",\n      \"pmids\": [\"16275925\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"The N-terminal flavoenzyme domain of MICAL performs an NADPH-dependent reaction reducing molecular oxygen to H2O2 (Km,NADPH = 222 μM; kcat = 77 sec−1). H2O2 production was proposed as a signaling molecule in axon guidance.\",\n      \"method\": \"Biochemical enzyme kinetics assay with purified MICAL flavodomain; measurement of H2O2 production\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro enzymatic reconstitution with kinetic characterization, single lab\",\n      \"pmids\": [\"16275926\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"MICAL enzymatic activity is autoinhibited by its C-terminal domain. CRMP (collapsin response mediator protein) and Plexin physically associate with non-enzymatic and enzymatic domains of MICAL respectively, and together release MICAL enzymatic autoinhibition. Semaphorin signaling promotes the CRMP–MICAL physical association.\",\n      \"method\": \"Co-immunoprecipitation; domain mapping; enzyme activity assays with truncation and domain mutants; cell-based assays\",\n      \"journal\": \"The Journal of Neuroscience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — reciprocal co-IP and activity assays, single lab, two orthogonal methods\",\n      \"pmids\": [\"18305261\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"MICAL-1 interacts with the hydrophobic motif of NDR1/2 kinases and negatively regulates MST-NDR kinase signaling by competing with MST1 for NDR binding, thereby inhibiting NDR-dependent proapoptotic signaling. Overexpression of MICAL-1 reduces NDR activation; knockdown augments it.\",\n      \"method\": \"Proteomics/mass spectrometry screen; co-immunoprecipitation; kinase activity assays; siRNA knockdown and overexpression with apoptosis readout\",\n      \"journal\": \"Molecular and Cellular Biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — proteomic identification, co-IP, enzymatic assay, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"21730291\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Human MICAL1 is autoinhibited by its C-terminal coiled-coil region. All three human MICALs (MICAL1, MICAL2, MICAL3) regulate actin stress fibers in non-neural cells and require ROS generation for their actin-regulatory function.\",\n      \"method\": \"siRNA knockdown; overexpression of truncation mutants; fluorescence microscopy of actin structures; ROS measurement\",\n      \"journal\": \"Journal of Cell Science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — KD/OE with defined phenotype, domain mapping, single lab\",\n      \"pmids\": [\"22331357\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Mical-mediated oxidation of actin (Met44/Met47) improves cofilin binding to filaments, and the combined effect dramatically accelerates F-actin disassembly compared to either alone. This synergism is necessary and sufficient for F-actin disassembly in vivo and magnifies effects on axon guidance and semaphorin-plexin repulsion.\",\n      \"method\": \"In vitro actin disassembly assays with purified Mical and cofilin; TIRF microscopy; Drosophila genetic assays\",\n      \"journal\": \"Nature Cell Biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — in vitro reconstitution, in vivo genetics, TIRF imaging, multiple orthogonal methods\",\n      \"pmids\": [\"27454820\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Mical stereospecifically oxidizes actin M44 and M47 residues. CryoEM reconstruction at 3.9 Å reveals oxidation reorients the M44 side chain and induces a new intermolecular interaction M47-O-T351 that promotes Mox-actin instability. Mox-actin can undergo extremely fast disassembly (84 subunits/s) and allows cofilin-mediated severing even in the presence of inorganic phosphate.\",\n      \"method\": \"Near-atomic resolution cryo-EM; single filament TIRF microscopy; site-directed mutagenesis\",\n      \"journal\": \"Nature Communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — cryo-EM structure, mutagenesis, single-filament imaging, multiple orthogonal methods\",\n      \"pmids\": [\"29259197\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"MICAL1-induced oxidation of actin filaments suppresses their protection from cofilin by: increasing cofilin binding and severing rates by several orders of magnitude; allowing non-activated phosphomimetic S3D-cofilin to bind and sever oxidized filaments; and causing tropomyosin Tpm1.8 to lose its ability to protect filaments from cofilin severing.\",\n      \"method\": \"Single filament experiments in vitro; fluorescence microscopy; TIRF assays with purified proteins\",\n      \"journal\": \"EMBO Reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution with purified proteins, single-filament imaging, multiple conditions tested\",\n      \"pmids\": [\"33393173\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"MICAL1 was identified as a novel protein interacting with the CasL SH3 domain via its PPKPP proline-rich sequence. MICAL1 also associates with vimentin intermediate filaments through its C-terminal region, and colocalizes with vimentin and CasL at the perinuclear area.\",\n      \"method\": \"Far Western screening; co-immunoprecipitation; immunofluorescence colocalization\",\n      \"journal\": \"The Journal of Biological Chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — co-IP and pulldown, immunofluorescence, single lab, two orthogonal methods\",\n      \"pmids\": [\"11827972\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"MICAL-1 (isoforms 1a and 1b) specifically interacts with active GTP-bound Rab1 in a nucleotide-dependent manner, with the interaction domain mapped to the C-terminus of MICAL-1, which also mediates binding to vimentin. MICAL-1 displays predominantly cytosolic localization by cell fractionation.\",\n      \"method\": \"Yeast two-hybrid; GST pulldown; cell fractionation; domain mapping\",\n      \"journal\": \"Biochemical and Biophysical Research Communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — yeast two-hybrid confirmed by GST pulldown, cell fractionation, single lab\",\n      \"pmids\": [\"12788069\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Vertebrate MICAL-1 regulates the targeting of secretory vesicles containing IgCAM cell adhesion molecules to the neuronal growth cone membrane through its ability to control the actin cytoskeleton via redox chemistry, thereby maintaining appropriate IgCAM cell surface levels. This is required for lamina-specific mossy fibre axon targeting in the mouse hippocampus in vivo.\",\n      \"method\": \"MICAL-1 knockout mouse; live imaging; cell surface biotinylation assays; immunofluorescence\",\n      \"journal\": \"Nature Communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — knockout mouse with defined in vivo phenotype, cell biological assays, multiple methods\",\n      \"pmids\": [\"25007825\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Human MICAL1 catalytic properties are regulated by its CH, LIM, and C-terminal domains. The C-terminal domain causes ~10-fold decrease in kcat, establishing an inactive/active conformational equilibrium (autoinhibition). F-actin lowers Km,NADPH and increases kcat for all MICAL forms, and actin depolymerization may be mediated by H2O2 produced by the NADPH oxidase reaction rather than direct actin methionine hydroxylation.\",\n      \"method\": \"Enzyme kinetics with purified full-length and truncated MICAL1 forms; NADPH oxidase assays; F-actin depolymerization assays\",\n      \"journal\": \"Archives of Biochemistry and Biophysics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro enzyme kinetics with multiple truncation mutants, single lab\",\n      \"pmids\": [\"26845023\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Rab8 in its active GTP-bound state binds MICAL1 and stabilizes its active conformation, causing a specific ~4-fold increase in kcat of the NADPH oxidase reaction. SAXS measurements support a 1:1 complex between full-length MICAL1 and Rab8 with Kd ~8 μM. Free full-length MICAL1 adopts an autoinhibited conformation in which the C-terminal region interferes with catalytic conformational changes.\",\n      \"method\": \"Enzyme kinetics; small-angle X-ray scattering (SAXS); SAXS-based structural modeling\",\n      \"journal\": \"Protein Science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — kinetics + SAXS structure, single lab, two orthogonal methods\",\n      \"pmids\": [\"30242933\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Cryo-EM structure of human MICAL1 at 3.1 Å resolution reveals that autoinhibition is mediated by an intramolecular interaction between the N-terminal catalytic domain and C-terminal coiled-coil domain, blocking F-actin interaction. Allosteric changes in the coiled-coil domain and binding of the tripartite CH-L2α1-LIM assembly to the coiled-coil domain are required for MICAL1 activation.\",\n      \"method\": \"Cryo-EM structure determination; biochemical binding assays; functional cell-based assays; mutagenesis\",\n      \"journal\": \"Nature Communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — cryo-EM structure at 3.1 Å, biochemical validation, mutagenesis, multiple orthogonal methods\",\n      \"pmids\": [\"39532862\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"MICAL1 oxidizes Met308, a conserved residue in the calmodulin-binding domain of CaMKII, reducing CaM binding and CaMKII activity. Loss of MICAL1 in mice causes cardiac arrhythmias and premature death due to CaMKII hyperactivation, establishing CaMKII as a non-actin substrate of MICAL1.\",\n      \"method\": \"MICAL1 knockout mouse model; biochemical assays for CaMKII activity and CaM binding; site-specific mutagenesis; Drosophila genetic model; human iPSC-derived cardiomyocytes\",\n      \"journal\": \"The Journal of Clinical Investigation\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — KO mouse with defined phenotype, biochemical assays, mutagenesis, multiple species validation\",\n      \"pmids\": [\"32749237\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"PAK1 (a CDC42 GTPase effector) associates with MICAL1 via its N-terminal catalytic monooxygenase and calponin homology domains, and phosphorylates MICAL1 on two serine residues, leading to accelerated F-actin disassembly. Extracellular ligand stimulation leads to PAK-dependent MICAL1 phosphorylation, linking external signals to MICAL1 activation.\",\n      \"method\": \"Co-immunoprecipitation; mass spectrometry phosphoproteomics; in vitro kinase assays; F-actin disassembly assays\",\n      \"journal\": \"Cell Reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP, kinase assay, MS phosphoproteomics, single lab with multiple methods\",\n      \"pmids\": [\"36198272\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"The unconventional myosin Myo15 physically interacts with Mical and uses its motor and MyTH4-FERM domains to broaden Mical's distribution, thereby spatiotemporally positioning and expanding Mical-mediated F-actin disassembly and cellular remodeling in response to Semaphorin/Plexin signals.\",\n      \"method\": \"Protein-protein interaction assays; Drosophila genetic epistasis; imaging of cellular remodeling\",\n      \"journal\": \"Science Advances\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — genetic and biochemical interaction data, single lab, two methods\",\n      \"pmids\": [\"33980493\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Two MICAL-1 mutations causing autosomal dominant lateral temporal epilepsy (p.Gly150Ser in the monooxygenase domain and p.Ala1065fs frameshift in the C-terminal domain) both significantly increased MICAL-1 oxidoreductase activity and induced cell contraction in COS7 cells, demonstrating gain-of-function effects on MICAL-1 enzymatic activity leading to dysregulation of F-actin dynamics.\",\n      \"method\": \"Cell-based oxidoreductase activity assays; cell contraction assay in COS7 cells; whole exome sequencing\",\n      \"journal\": \"Annals of Neurology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — cell-based enzymatic assay with patient mutations, single lab\",\n      \"pmids\": [\"29394500\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Mical oxidizes Tau at Cys322 via its monooxygenase domain, altering Tau interactions with microtubules and the actin cytoskeleton, and greatly affecting Tau aggregation propensity and Tau-associated toxicity in a Drosophila Tauopathy model. MICAL1 co-localizes with Tau in Pick bodies in human Tauopathy samples.\",\n      \"method\": \"Drosophila genetic interaction studies; Mical inhibitor treatment; monooxygenase domain-specific mutation; Tau cysteine mutant transgenes; mass spectrometry quantification of cysteine oxidation\",\n      \"journal\": \"Acta Neuropathologica Communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic interactions, domain-specific mutation, MS validation, single lab\",\n      \"pmids\": [\"35379354\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"MICAL1 locally depolymerizes cortical actin at HIV-1 budding sites to promote viral budding and release. MICAL1 directly disassembles branched actin networks. The Rab35 activator of MICAL1 is also recruited at budding sites and functions in the same pathway. MICAL1 depletion prevents timely ESCRT scission machinery recruitment.\",\n      \"method\": \"Superresolution microscopy; MICAL1 siRNA depletion; F-actin disassembly assays; in vitro branched-actin network assays; ESCRT recruitment imaging\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — KD with defined phenotype, in vitro actin assay, imaging, single lab\",\n      \"pmids\": [\"39556735\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"MICAL1 is a shear-activated regulator that promotes local F-actin disassembly around the GPIb-IX-V complex in platelets, enabling its translocation to lipid rafts and reinforcing VWF binding under high-shear conditions. MICAL1-deficient platelets display impaired VWF-GPIbα adhesion, increased deformability under shear, and defective thrombus formation in vivo.\",\n      \"method\": \"MICAL1-knockout mice; microfluidics shear assay; TIRF/superresolution microscopy; lipid raft fractionation; in vivo thrombosis model\",\n      \"journal\": \"Nature Communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — KO mouse, biochemical fractionation, microfluidics, multiple methods, single lab\",\n      \"pmids\": [\"40783397\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"F-actin bundled with fascin is extensively disassembled by Mical; moreover, Mical-oxidized actin is poorly bundled by fascin, amplifying disassembly. Cofilin synergizes with Mical to dramatically amplify disassembly of bundled F-actin beyond the sum of their individual effects. Fascin bundling dampens Mical's disassembly activity, establishing a balance between crosslinking and oxidative disassembly.\",\n      \"method\": \"Biochemical actin bundling/disassembly assays with purified proteins; structural imaging; Drosophila genetic epistasis assays; high-resolution cellular imaging\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — in vitro reconstitution, structural assays, in vivo genetics, multiple orthogonal methods\",\n      \"pmids\": [\"37725655\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"The SH3 domain of ASAP1 binds to the proline-rich motif (PRM) of MICAL1 with sub-μM affinity. Crystal structure of the ASAP1-SH3/MICAL1-PRM complex reveals a unique binding mode where two negatively charged patches in the SH3 pocket recognize an 'xPx+Px+' sequence in MICAL1-PRM, differing from typical SH3/PRM interactions.\",\n      \"method\": \"Crystal structure determination; isothermal titration calorimetry (ITC); mutagenesis\",\n      \"journal\": \"International Journal of Molecular Sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — crystal structure with binding affinity measurement, single lab\",\n      \"pmids\": [\"36674928\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2028,\n      \"finding\": \"MICAL1 co-immunoprecipitates with RAB35, and activated RAB35 (GTP-bound) increases MICAL1 interaction. Silencing of RAB35 reduced ROS generation and Akt phosphorylation downstream of MICAL1 in breast cancer cells.\",\n      \"method\": \"Co-immunoprecipitation; Rac1 pulldown assay; ROS measurement; siRNA knockdown\",\n      \"journal\": \"BMC Cancer\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single co-IP, single lab, no direct enzymatic characterization of RAB35-MICAL1 interaction\",\n      \"pmids\": [\"27430308\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"NEDD9 interacts with MICAL1 under hypoxic conditions, and NEDD9 stabilizes MICAL1 protein. Silencing NEDD9 reduced MICAL1 stability. MICAL1 overexpression selectively increased GTP-Rac1 (but not Cdc42 or RhoA), implicating MICAL1 in Rac1-dependent migration.\",\n      \"method\": \"Co-immunoprecipitation; siRNA knockdown; Rac1/Cdc42/RhoA pulldown activity assay\",\n      \"journal\": \"Frontiers in Pharmacology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single co-IP, single lab, limited mechanistic follow-up\",\n      \"pmids\": [\"31019460\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"PlexinA1 forms a complex with MICAL1 and promotes Rac1 activation and ROS production, which in turn prevents MICAL1 ubiquitin/proteasome-mediated degradation and stabilizes MICAL1 protein.\",\n      \"method\": \"Co-immunoprecipitation; immunofluorescence; siRNA-mediated knockdown; Rac1 pulldown assay; ROS measurement; ubiquitination assay\",\n      \"journal\": \"Biochimica et Biophysica Acta. Molecular Basis of Disease\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single co-IP, single lab, no direct in vitro reconstitution of the regulatory pathway\",\n      \"pmids\": [\"38508474\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Sox14 transcription factor directly regulates expression of mical in Drosophila, establishing a genetic pathway where Sox14 → Mical governs dendrite severing during pruning. Overexpression of Mical significantly rescues pruning defects in sox14 mutants.\",\n      \"method\": \"Genetic epistasis; loss-of-function and gain-of-function in Drosophila; chromatin immunoprecipitation (Sox14 binding to mical promoter)\",\n      \"journal\": \"Nature Neuroscience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis with rescue experiment, transcriptional target validation, single lab\",\n      \"pmids\": [\"19881505\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"MICAL1 is a multidomain FAD-containing flavoprotein monooxygenase that, upon activation by semaphorin-plexin signaling or upstream regulators (including RAB8, PAK1, and RAB35), uses NADPH to stereospecifically oxidize methionine residues 44 and 47 of actin filaments, causing severing, depolymerization, and reduced reassembly; this activity is autoinhibited by intramolecular interactions between its N-terminal catalytic domain and C-terminal coiled-coil domain (as resolved by cryo-EM), is synergized by cofilin, is reversed by the methionine sulfoxide reductase SelR/MsrB, and extends beyond actin to substrates including CaMKII (M308 oxidation) and Tau (C322 oxidation), with roles in axon guidance, cytokinesis, platelet function, cardiac stress responses, and epilepsy.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"MICAL1 is a multidomain FAD-dependent flavoprotein monooxygenase that couples redox chemistry to actin cytoskeleton remodeling, serving as the effector that links semaphorin-plexin signaling to F-actin reorganization in vivo [#1, #3]. Its N-terminal flavoenzyme domain is structurally related to p-hydroxybenzoate hydroxylase and uses NADPH and molecular oxygen to drive catalysis, with a flavin ring that switches between positions coupled to opening of a substrate channel [#3, #4]. The enzyme directly binds and disassembles individual and bundled F-actin by stereospecifically oxidizing actin methionine residues Met44 and Met47; this post-translational oxidation reorients the Met44 side chain and induces a new intermolecular contact that destabilizes the filament, simultaneously severing it and blocking reassembly [#0, #9]. Oxidation is reversed by the methionine sulfoxide reductase SelR/MsrB, which restores normal polymerization and establishes a reversible redox switch on actin [#2]. MICAL1-mediated oxidation also primes filaments for accelerated cofilin binding and severing — overriding the protection normally conferred by tropomyosin and fascin bundling — so that cofilin synergizes with MICAL1 to dramatically amplify disassembly [#8, #10, #24]. MICAL1 catalytic activity is held in an autoinhibited state by an intramolecular interaction between the N-terminal catalytic domain and the C-terminal coiled-coil region, resolved by cryo-EM, and is relieved by upstream regulators including GTP-bound Rab8, PAK1-mediated phosphorylation, and CRMP/Plexin association downstream of semaphorin signaling [#5, #15, #16, #18]. Its substrate range extends beyond actin: MICAL1 oxidizes Met308 of CaMKII to reduce calmodulin binding and kinase activity, and oxidizes Cys322 of Tau to alter cytoskeletal interactions and aggregation [#17, #21]. Through these activities MICAL1 controls axon guidance and growth-cone IgCAM trafficking, dendrite pruning, cytokinesis-related membrane events, HIV-1 budding, shear-dependent platelet adhesion, and cardiac CaMKII homeostasis [#13, #17, #22, #23, #29]. Gain-of-function MICAL1 mutations that elevate its oxidoreductase activity cause autosomal dominant lateral temporal epilepsy [#20].\",\n  \"teleology\": [\n    {\n      \"year\": 2002,\n      \"claim\": \"Established the first physical partners of MICAL1, placing it at cytoskeletal adaptor junctions before its enzymatic role was known.\",\n      \"evidence\": \"Far Western screening, co-IP, and immunofluorescence in cells identifying CasL SH3 and vimentin interactions\",\n      \"pmids\": [\"11827972\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No enzymatic function assigned at this stage\", \"Functional consequence of vimentin/CasL binding unresolved\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Linked MICAL1 to Rab GTPase signaling by showing nucleotide-dependent binding to active Rab1, implicating it in vesicle-related processes.\",\n      \"evidence\": \"Yeast two-hybrid, GST pulldown, and cell fractionation with domain mapping\",\n      \"pmids\": [\"12788069\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No catalytic consequence of Rab1 binding tested\", \"Cytosolic localization not connected to a pathway\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Defined MICAL1 as a FAD monooxygenase with a PHBH-like fold and a conformational flavin switch, and showed it produces H2O2 from NADPH/O2, framing the enzymatic basis of its cytoskeletal effects.\",\n      \"evidence\": \"1.45 Å X-ray crystallography of the mouse flavodomain plus enzyme kinetics measuring H2O2 production\",\n      \"pmids\": [\"16275925\", \"16275926\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Physiological protein substrate not yet identified\", \"Whether H2O2 or direct oxidation drives actin disassembly unresolved\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Resolved how MICAL is switched on, showing C-terminal autoinhibition relieved by CRMP and Plexin binding downstream of semaphorin signaling.\",\n      \"evidence\": \"Co-IP, domain mapping, and activity assays with truncation mutants in cells\",\n      \"pmids\": [\"18305261\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural basis of autoinhibition not defined\", \"Single lab\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Placed Mical in a transcriptional pathway for developmental remodeling by identifying Sox14 as a direct upstream activator driving dendrite pruning.\",\n      \"evidence\": \"Drosophila genetic epistasis, rescue, and ChIP of Sox14 at the mical promoter\",\n      \"pmids\": [\"19881505\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Vertebrate transcriptional regulation not addressed\", \"Downstream actin events inferred from genetics\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Identified the core catalytic mechanism — direct oxidation of actin Met44 that simultaneously severs and depolymerizes filaments — defining MICAL as an actin-modifying enzyme.\",\n      \"evidence\": \"In vitro biochemistry with purified protein, mutagenesis, and mass spectrometry\",\n      \"pmids\": [\"22116028\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Reversibility of the modification not yet shown\", \"Additional oxidized residues not yet mapped\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Expanded MICAL1's reach beyond the cytoskeleton by showing it competes with MST1 for NDR kinase binding to suppress proapoptotic signaling.\",\n      \"evidence\": \"Proteomic screen, co-IP, kinase assays, and knockdown/overexpression apoptosis readouts\",\n      \"pmids\": [\"21730291\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Redox activity not required for the scaffold function\", \"Single lab\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Established MICAL-driven actin oxidation as a reversible redox switch by identifying SelR/MsrB as the stereospecific reductase opposing MICAL in vivo.\",\n      \"evidence\": \"Drosophila genetic epistasis and in vitro stereospecific reduction of Mical-oxidized actin\",\n      \"pmids\": [\"24212093\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Regulation of the MICAL/SelR balance in vivo not detailed\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Connected MICAL1 redox actin control to growth-cone biology, showing it governs IgCAM vesicle targeting and lamina-specific axon targeting in mouse.\",\n      \"evidence\": \"MICAL1 knockout mouse, surface biotinylation, and live imaging\",\n      \"pmids\": [\"25007825\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct vesicle-actin coupling mechanism not fully resolved\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Demonstrated cofilin synergy with MICAL oxidation and refined the residues to Met44/Met47, explaining how oxidation potentiates accelerated disassembly.\",\n      \"evidence\": \"In vitro disassembly assays, TIRF microscopy, and Drosophila genetics; plus kinetic dissection of domain autoinhibition and the H2O2-versus-direct-oxidation question\",\n      \"pmids\": [\"27454820\", \"26845023\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Relative contributions of direct oxidation versus H2O2 still debated between studies\", \"Mechanism of cofilin enhancement structurally undefined at the time\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Provided near-atomic structural mechanism for how oxidation destabilizes filaments via Met44 reorientation and a new M47-O-T351 contact enabling ultrafast disassembly.\",\n      \"evidence\": \"3.9 Å cryo-EM, single-filament TIRF, and mutagenesis\",\n      \"pmids\": [\"29259197\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structure of the full autoinhibited enzyme not yet solved\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Showed how upstream signals activate MICAL1, with GTP-Rab8 stabilizing the active conformation and relieving C-terminal autoinhibition.\",\n      \"evidence\": \"Enzyme kinetics and SAXS modeling of the 1:1 MICAL1-Rab8 complex; epilepsy mutations shown to be enzymatic gain-of-function\",\n      \"pmids\": [\"30242933\", \"29394500\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"SAXS provides low-resolution architecture only\", \"Mechanism connecting elevated activity to seizures untested in neurons\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Established a non-actin substrate, showing MICAL1 oxidizes CaMKII Met308 to limit kinase activity and protect against cardiac arrhythmia.\",\n      \"evidence\": \"MICAL1 knockout mouse, CaMKII biochemistry, mutagenesis, Drosophila and iPSC-cardiomyocyte models\",\n      \"pmids\": [\"32749237\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Spatial regulation of MICAL1-CaMKII oxidation in cardiomyocytes not mapped\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Mechanistically explained how oxidation overrides filament protection, allowing even phosphomimetic cofilin to sever and stripping tropomyosin protection.\",\n      \"evidence\": \"Single-filament TIRF assays with purified proteins; plus identification of Myo15 as a spatial positioner of Mical activity\",\n      \"pmids\": [\"33393173\", \"33980493\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo significance of tropomyosin override not directly tested\", \"Myo15-Mical coupling shown in Drosophila only\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Identified PAK1 phosphorylation as an extracellular-signal-driven activation input and added Tau Cys322 as another oxidation substrate relevant to tauopathy.\",\n      \"evidence\": \"Co-IP, phosphoproteomics, kinase and disassembly assays; Drosophila tauopathy genetics with MS of cysteine oxidation\",\n      \"pmids\": [\"36198272\", \"35379354\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"PAK1 phospho-sites not structurally placed in the autoinhibition model\", \"Human relevance of Tau oxidation beyond colocalization unestablished\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Defined the interplay between bundling and oxidative disassembly, showing fascin dampens while cofilin amplifies MICAL action, and characterized a distinct ASAP1-SH3/MICAL1-PRM binding mode.\",\n      \"evidence\": \"In vitro bundling/disassembly assays and Drosophila genetics; crystal structure and ITC of the ASAP1-SH3/MICAL1-PRM complex\",\n      \"pmids\": [\"37725655\", \"36674928\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Functional consequence of ASAP1 binding to MICAL1 in cells not defined\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Solved the human autoinhibited enzyme structure, defining the N-terminal catalytic / C-terminal coiled-coil intramolecular lock and the tripartite CH-L2α1-LIM assembly required for activation.\",\n      \"evidence\": \"3.1 Å cryo-EM with biochemical binding assays, mutagenesis, and cell-based assays; plus role in HIV-1 budding via cortical actin depolymerization\",\n      \"pmids\": [\"39532862\", \"39556735\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How each upstream regulator engages the activation assembly not fully mapped\", \"Rab35 recruitment mechanism at budding sites incompletely defined\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Extended MICAL1 function to mechanotransduction in platelets, showing shear-activated F-actin disassembly enables GPIb-IX-V translocation to lipid rafts and stable VWF adhesion.\",\n      \"evidence\": \"MICAL1 knockout mice, microfluidics shear assays, superresolution imaging, lipid raft fractionation, and in vivo thrombosis\",\n      \"pmids\": [\"40783397\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular sensor coupling shear to MICAL1 activation unidentified\", \"Single lab\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How the multiple activation inputs (Rab8, Rab35, PAK1, CRMP/Plexin, NEDD9, PlexinA1) are integrated to spatially and temporally tune MICAL1 oxidative activity, and how substrate selection between actin, CaMKII, and Tau is governed, remain unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No unified model of competitive/cooperative activation\", \"Substrate-selectivity determinants unknown\", \"Several Rac1/ROS regulatory links rest on single co-IP studies\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016491\", \"supporting_discovery_ids\": [0, 3, 4, 9, 14, 17, 21]},\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0, 9, 17, 21]},\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [1, 8, 10, 24]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [6]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [12]},\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [1, 7, 11]},\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [22, 23]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1266738\", \"supporting_discovery_ids\": [1, 13, 29]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [5, 15, 18]},\n      {\"term_id\": \"R-HSA-109582\", \"supporting_discovery_ids\": [23]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"RAB8\", \"PAK1\", \"CRMP\", \"PLXNA1\", \"NDR1\", \"CASL\", \"ASAP1\", \"RAB1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":9,"faith_total":9,"faith_pct":100.0}}