{"gene":"MYO5C","run_date":"2026-06-10T05:19:52","timeline":{"discoveries":[{"year":2002,"finding":"Myo5c tail domain (dominant-negative) colocalizes with and perturbs a membrane compartment containing the transferrin receptor and Rab8, and causes transferrin accumulation in that compartment, indicating Myo5c functions in transferrin/Rab8-associated membrane trafficking in epithelial cells.","method":"GFP-Myo5c tail dominant-negative expression in HeLa cells with immunolocalization","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — dominant-negative approach with colocalization, single lab but two markers (transferrin receptor and Rab8) provide convergent evidence","pmids":["11870218"],"is_preprint":false},{"year":2008,"finding":"Myo5c associates with mature secretory vesicles (by confocal and immunogold EM) in lacrimal acinar cells; dominant-negative Myo5c tail expression reduces carbachol-stimulated secretion of vesicle content markers (secretory component, syncollin-GFP) and impairs compound vesicle fusion, indicating Myo5c facilitates apical exocytosis of secretory vesicles.","method":"Confocal fluorescence and immunogold EM; adenovirus-mediated dominant-negative GFP-Myo5c-tail expression; measurement of secretory vesicle content release and vesicle diameter","journal":"American journal of physiology. Cell physiology","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (EM localization, dominant-negative functional assay, two independent secretory markers, vesicle morphometry), independently consistent with PMID:19741097","pmids":["18434623"],"is_preprint":false},{"year":2009,"finding":"Endogenous Myo5c localizes to two distinct compartments in MCF-7 cells: small puncta that colocalize with secretory granule markers (chromogranin A, Rab27b) and slender tubules labeled by Rab8a. Myo5c puncta move slowly (~30 nm/s) in a microtubule-independent manner while tubules move rapidly (~440 nm/s) in a microtubule-dependent manner. Dominant-negative Myo5c tail dramatically perturbs the distribution of granule markers, establishing a functional role in secretory granule trafficking.","method":"Live-cell GFP imaging, TIRF microscopy, immunofluorescence colocalization, dominant-negative tail expression, microtubule depolymerization experiments","journal":"Molecular biology of the cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (live imaging, TIRF, co-localization with multiple markers, dominant-negative functional readout), single lab but comprehensive","pmids":["19741097"],"is_preprint":false},{"year":2009,"finding":"Myo5c colocalizes with Rab8 in HepG2 cells; expression of dominant-negative Myo5c tail reduces release of dengue virus 2 (DV2) from cells without affecting intracellular viral production, and is accompanied by increased Rab8 expression, indicating Myo5c/Rab8-mediated membrane trafficking is required for DV2 egress.","method":"Fluorescent immunostaining/colocalization, dominant-negative Myo5c tail stable transfection, plaque assay for viral titer, flow cytometry for Rab8 expression","journal":"Intervirology","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — dominant-negative approach with functional readout (plaque assay) and Rab8 quantification, single lab with multiple measurements","pmids":["19641326"],"is_preprint":false},{"year":2014,"finding":"Myo5c is a low duty ratio, non-processive motor as a single dimer, but coupling of two Myo5c-HMM dimers via a DNA scaffold enables processive movement along actin filaments in 30–36 nm steps. Steady-state ATPase and ADP dissociation kinetics show that the Myo5c-HMM dimer has 6-fold lower Km for actin than the single-headed S1, indicating inter-head cooperativity.","method":"Steady-state ATPase assay, ADP dissociation kinetics, nanometer-precision single-molecule tracking of DNA-scaffold-linked Myo5c dimers","journal":"Scientific reports","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution with ATPase kinetics and single-molecule motility assays, single lab but multiple orthogonal biochemical and biophysical methods","pmids":["24809456"],"is_preprint":false},{"year":2024,"finding":"Human Myo5c-HMM motor activity is modulated by tropomyosin isoforms (Tpm1.6, Tpm1.8, Tpm3.1 alter maximum actin-activated ATPase and motile activity) and inhibited by pentabromopseudilin (PBP) with IC50 ~280 nM, predicted to bind near the actin and nucleotide binding site. Myo5c is composed of a motor domain, neck with six IQ motifs binding calmodulin or EF-hand light chains, coiled-coil dimerization region, and C-terminal globular tail domain.","method":"Recombinant human Myo5c-HMM purification and reconstitution with CaM/light chains; actin-activated ATPase assay with tropomyosin cofilaments; in vitro motility assay; computational docking of PBP; immunohistochemical localization in rat and human tissue","journal":"Frontiers in physiology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution with multiple ATPase and motility assays plus small-molecule inhibitor characterization, single lab but multiple orthogonal functional readouts","pmids":["38606007"],"is_preprint":false},{"year":2025,"finding":"myo5c knockdown in HL60 cells enhanced neutrophil extracellular trap (NET) formation upon stimulation, suggesting Myo5c negatively regulates neutrophil activation. In vivo, neutrophil Myo5c expression was inversely correlated with IL-6 levels and markers of neutrophil and platelet activation after cardiopulmonary bypass.","method":"myo5c knockout in HL60 cells (neutrophil cell line) with NET formation assay; RNA sequencing of patient neutrophils; correlation analyses","journal":"Frontiers in cardiovascular medicine","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single KO cell line experiment for NET phenotype, no mechanistic pathway established, limited follow-up of molecular mechanism","pmids":["40495980"],"is_preprint":false}],"current_model":"MYO5C encodes a class V actin-based motor protein (myosin Vc) expressed predominantly in epithelial and glandular tissues, where it associates with Rab8a-positive tubular membranes and Rab27b/chromogranin A-positive secretory granules, powers slow (~30 nm/s) microtubule-independent movement of secretory granules and rapid (~440 nm/s) microtubule-dependent movement of tubular carriers, and is required for regulated exocytosis of secretory vesicles; as an individual dimer it is a low duty-ratio non-processive motor, but coupling of two dimers enables processive stepping in ~30–36 nm increments along actin filaments, and its ATPase/motile activity is modulated by tropomyosin isoforms and inhibited by pentabromopseudilin."},"narrative":{"mechanistic_narrative":"MYO5C encodes myosin Vc, a class V actin-based motor that drives membrane trafficking and regulated exocytosis in epithelial and glandular cells [PMID:19741097]. The motor associates with two distinct membrane compartments: chromogranin A/Rab27b-positive secretory granules, which it moves slowly (~30 nm/s) in a microtubule-independent fashion, and Rab8a-positive tubular carriers, which move rapidly (~440 nm/s) along microtubules [PMID:19741097]. Through its Rab8-associated trafficking activity it controls transferrin/transferrin-receptor recycling compartments, and dominant-negative tail expression perturbs these compartments and granule distribution, establishing a requirement for Myo5c in secretory granule transport and apical exocytosis [PMID:11870218, PMID:19741097]. In lacrimal acinar cells Myo5c localizes to mature secretory vesicles and is required for carbachol-stimulated content release and compound vesicle fusion [PMID:18434623]. Biophysically, a single Myo5c dimer is a low duty-ratio, non-processive motor, but coupling of two dimers confers processive stepping in 30–36 nm increments, reflecting inter-head cooperativity, and its actin-activated ATPase and motility are tuned by tropomyosin isoforms and inhibited by pentabromopseudilin [PMID:24809456, PMID:38606007]. The protein comprises a motor domain, a neck with six IQ motifs binding calmodulin/EF-hand light chains, a coiled-coil dimerization region, and a C-terminal globular tail [PMID:38606007].","teleology":[{"year":2002,"claim":"Established that Myo5c participates in membrane trafficking by linking it to a Rab8/transferrin-receptor recycling compartment, the first functional clue to its cellular role.","evidence":"GFP-Myo5c tail dominant-negative expression and immunolocalization in HeLa cells","pmids":["11870218"],"confidence":"Medium","gaps":["Dominant-negative approach does not prove direct motor transport","No demonstration of direct Rab8 binding","Cargo specificity not defined"]},{"year":2008,"claim":"Showed Myo5c functions in regulated apical exocytosis by demonstrating it localizes to mature secretory vesicles and is required for stimulated content release and compound fusion.","evidence":"Confocal/immunogold EM and adenoviral dominant-negative tail expression with secretory readouts in lacrimal acinar cells","pmids":["18434623"],"confidence":"High","gaps":["Mechanism of vesicle fusion coupling unresolved","Light-chain/calcium regulation in this context not addressed"]},{"year":2009,"claim":"Resolved that Myo5c operates on two functionally distinct compartments with different motility regimes, distinguishing microtubule-independent granule movement from microtubule-dependent tubule transport.","evidence":"Live-cell GFP/TIRF imaging, colocalization with chromogranin A/Rab27b and Rab8a, and dominant-negative tail in MCF-7 cells","pmids":["19741097"],"confidence":"High","gaps":["Molecular basis of cargo selection between granules and tubules unknown","Direct Rab27b/Rab8a binding not biochemically established"]},{"year":2009,"claim":"Extended Myo5c/Rab8 trafficking to a pathophysiological readout by showing it is required for dengue virus egress without affecting intracellular replication.","evidence":"Colocalization, dominant-negative tail stable transfection, plaque assay and Rab8 flow cytometry in HepG2 cells","pmids":["19641326"],"confidence":"Medium","gaps":["Does not establish direct interaction with viral components","Step of egress pathway controlled by Myo5c undefined"]},{"year":2014,"claim":"Defined the motor mechanochemistry, showing single dimers are non-processive but paired dimers move processively with inter-head cooperativity.","evidence":"Steady-state ATPase, ADP dissociation kinetics, and single-molecule tracking of DNA-scaffold-linked Myo5c-HMM dimers","pmids":["24809456"],"confidence":"High","gaps":["Physiological mechanism for in vivo dimer coupling unknown","Cargo-induced activation not addressed"]},{"year":2024,"claim":"Characterized regulation of human Myo5c motor output by tropomyosin isoforms and identified a small-molecule inhibitor, plus its domain architecture and light-chain binding.","evidence":"Recombinant human Myo5c-HMM reconstitution, actin-activated ATPase with tropomyosin cofilaments, motility assays, PBP docking, and tissue immunohistochemistry","pmids":["38606007"],"confidence":"High","gaps":["In vivo relevance of specific tropomyosin isoforms not tested","Light-chain composition under physiological conditions undefined"]},{"year":2025,"claim":"Implicated Myo5c as a negative regulator of neutrophil activation, expanding its role beyond epithelial trafficking.","evidence":"myo5c knockout in HL60 cells with NET formation assay and RNA-seq correlation in patient neutrophils","pmids":["40495980"],"confidence":"Low","gaps":["No mechanistic pathway linking Myo5c to NET formation established","Single cell-line knockout without rescue","Correlative patient data only"]},{"year":null,"claim":"How Myo5c selectively recognizes and is activated by its distinct membrane cargoes (Rab8a tubules versus Rab27b/chromogranin granules) at the molecular level remains unresolved.","evidence":"","pmids":[],"confidence":"Low","gaps":["No direct biochemical mapping of tail-Rab interactions","Mechanism of cargo-dependent motor activation unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0003774","term_label":"cytoskeletal motor activity","supporting_discovery_ids":[2,4,5]},{"term_id":"GO:0140657","term_label":"ATP-dependent activity","supporting_discovery_ids":[4,5]},{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[4,5]}],"localization":[{"term_id":"GO:0031410","term_label":"cytoplasmic vesicle","supporting_discovery_ids":[1,2]},{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[4,5]}],"pathway":[{"term_id":"R-HSA-5653656","term_label":"Vesicle-mediated transport","supporting_discovery_ids":[0,1,2]},{"term_id":"R-HSA-9609507","term_label":"Protein localization","supporting_discovery_ids":[0,2]}],"complexes":[],"partners":["RAB8A","RAB27B","CALM1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9NQX4","full_name":"Unconventional myosin-Vc","aliases":[],"length_aa":1742,"mass_kda":202.8,"function":"May be involved in transferrin trafficking. Likely to power actin-based membrane trafficking in many physiologically crucial tissues","subcellular_location":"","url":"https://www.uniprot.org/uniprotkb/Q9NQX4/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/MYO5C","classification":"Not Classified","n_dependent_lines":0,"n_total_lines":1208,"dependency_fraction":0.0},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"CALD1","stoichiometry":0.2},{"gene":"CALM3","stoichiometry":0.2},{"gene":"CAPZB","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/MYO5C","total_profiled":1310},"omim":[{"mim_id":"610022","title":"MYOSIN VC; MYO5C","url":"https://www.omim.org/entry/610022"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Cytosol","reliability":"Supported"},{"location":"Nucleoplasm","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in many","driving_tissues":[],"url":"https://www.proteinatlas.org/search/MYO5C"},"hgnc":{"alias_symbol":["MGC74969"],"prev_symbol":[]},"alphafold":{"accession":"Q9NQX4","domains":[{"cath_id":"2.30.30,2.30.30","chopping":"2-62","consensus_level":"medium","plddt":84.1556,"start":2,"end":62},{"cath_id":"1.20.58.530","chopping":"495-561","consensus_level":"medium","plddt":86.8104,"start":495,"end":561},{"cath_id":"-","chopping":"1365-1497","consensus_level":"medium","plddt":87.8383,"start":1365,"end":1497}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9NQX4","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9NQX4-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9NQX4-F1-predicted_aligned_error_v6.png","plddt_mean":76.12},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=MYO5C","jax_strain_url":"https://www.jax.org/strain/search?query=MYO5C"},"sequence":{"accession":"Q9NQX4","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9NQX4.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9NQX4/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9NQX4"}},"corpus_meta":[{"pmid":"11870218","id":"PMC_11870218","title":"Human myosin-Vc is a novel class V myosin expressed in epithelial cells.","date":"2002","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/11870218","citation_count":149,"is_preprint":false},{"pmid":"29883837","id":"PMC_29883837","title":"Characteristics and Outcome of ROS1-Positive Non-Small Cell Lung Cancer Patients in Routine Clinical Practice.","date":"2018","source":"Journal of thoracic oncology : official publication of the International Association for the Study of Lung Cancer","url":"https://pubmed.ncbi.nlm.nih.gov/29883837","citation_count":87,"is_preprint":false},{"pmid":"19741097","id":"PMC_19741097","title":"Myosin Vc is a molecular motor that functions in secretory granule trafficking.","date":"2009","source":"Molecular biology of the cell","url":"https://pubmed.ncbi.nlm.nih.gov/19741097","citation_count":58,"is_preprint":false},{"pmid":"26126179","id":"PMC_26126179","title":"Genes associated with the progression of neurofibrillary tangles in Alzheimer's disease.","date":"2014","source":"Translational psychiatry","url":"https://pubmed.ncbi.nlm.nih.gov/26126179","citation_count":54,"is_preprint":false},{"pmid":"29325019","id":"PMC_29325019","title":"Deep molecular phenotypes link complex disorders and physiological insult to CpG methylation.","date":"2018","source":"Human molecular genetics","url":"https://pubmed.ncbi.nlm.nih.gov/29325019","citation_count":30,"is_preprint":false},{"pmid":"18434623","id":"PMC_18434623","title":"The class V myosin motor, myosin 5c, localizes to mature secretory vesicles and facilitates exocytosis in lacrimal acini.","date":"2008","source":"American journal of physiology. Cell physiology","url":"https://pubmed.ncbi.nlm.nih.gov/18434623","citation_count":27,"is_preprint":false},{"pmid":"20972619","id":"PMC_20972619","title":"A multi-gene transcriptional profiling approach to the discovery of cell signature markers.","date":"2010","source":"Cytotechnology","url":"https://pubmed.ncbi.nlm.nih.gov/20972619","citation_count":20,"is_preprint":false},{"pmid":"33622851","id":"PMC_33622851","title":"Comparative Genomics and Integrated Network Approach Unveiled Undirected Phylogeny Patterns, Co-mutational Hot Spots, Functional Cross Talk, and Regulatory Interactions in SARS-CoV-2.","date":"2021","source":"mSystems","url":"https://pubmed.ncbi.nlm.nih.gov/33622851","citation_count":17,"is_preprint":false},{"pmid":"24809456","id":"PMC_24809456","title":"Coupling of two non-processive myosin 5c dimers enables processive stepping along actin filaments.","date":"2014","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/24809456","citation_count":15,"is_preprint":false},{"pmid":"19641326","id":"PMC_19641326","title":"Myosin Vc, a member of the actin motor family associated with Rab8, is involved in the release of DV2 from HepG2 cells.","date":"2009","source":"Intervirology","url":"https://pubmed.ncbi.nlm.nih.gov/19641326","citation_count":14,"is_preprint":false},{"pmid":"34680964","id":"PMC_34680964","title":"Pendred Syndrome, or Not Pendred Syndrome? That Is the Question.","date":"2021","source":"Genes","url":"https://pubmed.ncbi.nlm.nih.gov/34680964","citation_count":10,"is_preprint":false},{"pmid":"31801962","id":"PMC_31801962","title":"Replicating associations between DNA methylation and body mass index in a longitudinal sample of older twins.","date":"2019","source":"International journal of obesity (2005)","url":"https://pubmed.ncbi.nlm.nih.gov/31801962","citation_count":10,"is_preprint":false},{"pmid":"32477400","id":"PMC_32477400","title":"Severe Phenotype in a Patient With Homozygous 15q21.2 Microdeletion Involving BCL2L10, GNB5, and MYO5C Genes, Resembling Infantile Developmental Disorder With Cardiac Arrhythmias (IDDCA).","date":"2020","source":"Frontiers in genetics","url":"https://pubmed.ncbi.nlm.nih.gov/32477400","citation_count":8,"is_preprint":false},{"pmid":"38606007","id":"PMC_38606007","title":"Motor properties of Myosin 5c are modulated by tropomyosin isoforms and inhibited by pentabromopseudilin.","date":"2024","source":"Frontiers in physiology","url":"https://pubmed.ncbi.nlm.nih.gov/38606007","citation_count":3,"is_preprint":false},{"pmid":"28665450","id":"PMC_28665450","title":"Predicting miRNA targets for head and neck squamous cell carcinoma using an ensemble method.","date":"2018","source":"The International journal of biological markers","url":"https://pubmed.ncbi.nlm.nih.gov/28665450","citation_count":3,"is_preprint":false},{"pmid":"40961947","id":"PMC_40961947","title":"Identification of novel type 1 and type 2 diabetes genes by co-localization of human islet eQTL and GWAS variants with colocRedRibbon.","date":"2025","source":"Cell genomics","url":"https://pubmed.ncbi.nlm.nih.gov/40961947","citation_count":3,"is_preprint":false},{"pmid":"22409602","id":"PMC_22409602","title":"Chromosome 15q21-22-related polymorphisms and haplotypes are associated with susceptibility to type-2 diabetic nonproliferative retinopathy.","date":"2012","source":"Genetic testing and molecular biomarkers","url":"https://pubmed.ncbi.nlm.nih.gov/22409602","citation_count":3,"is_preprint":false},{"pmid":"40495980","id":"PMC_40495980","title":"Neutrophil Myo5c gene downregulation is associated with postoperative organ dysfunction following pediatric cardiac surgery with cardiopulmonary bypass.","date":"2025","source":"Frontiers in cardiovascular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/40495980","citation_count":0,"is_preprint":false},{"pmid":"41722227","id":"PMC_41722227","title":"Research note: Integration of GWAS and RNA-seq identifies mutations linked to within feather patterning in domestic chickens.","date":"2026","source":"Poultry science","url":"https://pubmed.ncbi.nlm.nih.gov/41722227","citation_count":0,"is_preprint":false},{"pmid":"30496538","id":"PMC_30496538","title":"Duplicated Myosin V Genes in Teleosts Show Evolutionary Rate Variations among the Motor and Cargo-Binding Domains.","date":"2019","source":"Genome biology and evolution","url":"https://pubmed.ncbi.nlm.nih.gov/30496538","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":12778,"output_tokens":2132,"usd":0.035157,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":9056,"output_tokens":2572,"usd":0.05479,"stage2_stop_reason":"end_turn"},"total_usd":0.089947,"stage1_batch_id":"msgbatch_013a8chtv3AWyTqv4z4sEF4C","stage2_batch_id":"msgbatch_01AHsuW63vinsvuTgzmQ3n62","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2002,\n      \"finding\": \"Myo5c tail domain (dominant-negative) colocalizes with and perturbs a membrane compartment containing the transferrin receptor and Rab8, and causes transferrin accumulation in that compartment, indicating Myo5c functions in transferrin/Rab8-associated membrane trafficking in epithelial cells.\",\n      \"method\": \"GFP-Myo5c tail dominant-negative expression in HeLa cells with immunolocalization\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — dominant-negative approach with colocalization, single lab but two markers (transferrin receptor and Rab8) provide convergent evidence\",\n      \"pmids\": [\"11870218\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Myo5c associates with mature secretory vesicles (by confocal and immunogold EM) in lacrimal acinar cells; dominant-negative Myo5c tail expression reduces carbachol-stimulated secretion of vesicle content markers (secretory component, syncollin-GFP) and impairs compound vesicle fusion, indicating Myo5c facilitates apical exocytosis of secretory vesicles.\",\n      \"method\": \"Confocal fluorescence and immunogold EM; adenovirus-mediated dominant-negative GFP-Myo5c-tail expression; measurement of secretory vesicle content release and vesicle diameter\",\n      \"journal\": \"American journal of physiology. Cell physiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (EM localization, dominant-negative functional assay, two independent secretory markers, vesicle morphometry), independently consistent with PMID:19741097\",\n      \"pmids\": [\"18434623\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Endogenous Myo5c localizes to two distinct compartments in MCF-7 cells: small puncta that colocalize with secretory granule markers (chromogranin A, Rab27b) and slender tubules labeled by Rab8a. Myo5c puncta move slowly (~30 nm/s) in a microtubule-independent manner while tubules move rapidly (~440 nm/s) in a microtubule-dependent manner. Dominant-negative Myo5c tail dramatically perturbs the distribution of granule markers, establishing a functional role in secretory granule trafficking.\",\n      \"method\": \"Live-cell GFP imaging, TIRF microscopy, immunofluorescence colocalization, dominant-negative tail expression, microtubule depolymerization experiments\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (live imaging, TIRF, co-localization with multiple markers, dominant-negative functional readout), single lab but comprehensive\",\n      \"pmids\": [\"19741097\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Myo5c colocalizes with Rab8 in HepG2 cells; expression of dominant-negative Myo5c tail reduces release of dengue virus 2 (DV2) from cells without affecting intracellular viral production, and is accompanied by increased Rab8 expression, indicating Myo5c/Rab8-mediated membrane trafficking is required for DV2 egress.\",\n      \"method\": \"Fluorescent immunostaining/colocalization, dominant-negative Myo5c tail stable transfection, plaque assay for viral titer, flow cytometry for Rab8 expression\",\n      \"journal\": \"Intervirology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — dominant-negative approach with functional readout (plaque assay) and Rab8 quantification, single lab with multiple measurements\",\n      \"pmids\": [\"19641326\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Myo5c is a low duty ratio, non-processive motor as a single dimer, but coupling of two Myo5c-HMM dimers via a DNA scaffold enables processive movement along actin filaments in 30–36 nm steps. Steady-state ATPase and ADP dissociation kinetics show that the Myo5c-HMM dimer has 6-fold lower Km for actin than the single-headed S1, indicating inter-head cooperativity.\",\n      \"method\": \"Steady-state ATPase assay, ADP dissociation kinetics, nanometer-precision single-molecule tracking of DNA-scaffold-linked Myo5c dimers\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution with ATPase kinetics and single-molecule motility assays, single lab but multiple orthogonal biochemical and biophysical methods\",\n      \"pmids\": [\"24809456\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Human Myo5c-HMM motor activity is modulated by tropomyosin isoforms (Tpm1.6, Tpm1.8, Tpm3.1 alter maximum actin-activated ATPase and motile activity) and inhibited by pentabromopseudilin (PBP) with IC50 ~280 nM, predicted to bind near the actin and nucleotide binding site. Myo5c is composed of a motor domain, neck with six IQ motifs binding calmodulin or EF-hand light chains, coiled-coil dimerization region, and C-terminal globular tail domain.\",\n      \"method\": \"Recombinant human Myo5c-HMM purification and reconstitution with CaM/light chains; actin-activated ATPase assay with tropomyosin cofilaments; in vitro motility assay; computational docking of PBP; immunohistochemical localization in rat and human tissue\",\n      \"journal\": \"Frontiers in physiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution with multiple ATPase and motility assays plus small-molecule inhibitor characterization, single lab but multiple orthogonal functional readouts\",\n      \"pmids\": [\"38606007\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"myo5c knockdown in HL60 cells enhanced neutrophil extracellular trap (NET) formation upon stimulation, suggesting Myo5c negatively regulates neutrophil activation. In vivo, neutrophil Myo5c expression was inversely correlated with IL-6 levels and markers of neutrophil and platelet activation after cardiopulmonary bypass.\",\n      \"method\": \"myo5c knockout in HL60 cells (neutrophil cell line) with NET formation assay; RNA sequencing of patient neutrophils; correlation analyses\",\n      \"journal\": \"Frontiers in cardiovascular medicine\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single KO cell line experiment for NET phenotype, no mechanistic pathway established, limited follow-up of molecular mechanism\",\n      \"pmids\": [\"40495980\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"MYO5C encodes a class V actin-based motor protein (myosin Vc) expressed predominantly in epithelial and glandular tissues, where it associates with Rab8a-positive tubular membranes and Rab27b/chromogranin A-positive secretory granules, powers slow (~30 nm/s) microtubule-independent movement of secretory granules and rapid (~440 nm/s) microtubule-dependent movement of tubular carriers, and is required for regulated exocytosis of secretory vesicles; as an individual dimer it is a low duty-ratio non-processive motor, but coupling of two dimers enables processive stepping in ~30–36 nm increments along actin filaments, and its ATPase/motile activity is modulated by tropomyosin isoforms and inhibited by pentabromopseudilin.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"MYO5C encodes myosin Vc, a class V actin-based motor that drives membrane trafficking and regulated exocytosis in epithelial and glandular cells [#2]. The motor associates with two distinct membrane compartments: chromogranin A/Rab27b-positive secretory granules, which it moves slowly (~30 nm/s) in a microtubule-independent fashion, and Rab8a-positive tubular carriers, which move rapidly (~440 nm/s) along microtubules [#2]. Through its Rab8-associated trafficking activity it controls transferrin/transferrin-receptor recycling compartments, and dominant-negative tail expression perturbs these compartments and granule distribution, establishing a requirement for Myo5c in secretory granule transport and apical exocytosis [#0, #2]. In lacrimal acinar cells Myo5c localizes to mature secretory vesicles and is required for carbachol-stimulated content release and compound vesicle fusion [#1]. Biophysically, a single Myo5c dimer is a low duty-ratio, non-processive motor, but coupling of two dimers confers processive stepping in 30\\u201336 nm increments, reflecting inter-head cooperativity, and its actin-activated ATPase and motility are tuned by tropomyosin isoforms and inhibited by pentabromopseudilin [#4, #5]. The protein comprises a motor domain, a neck with six IQ motifs binding calmodulin/EF-hand light chains, a coiled-coil dimerization region, and a C-terminal globular tail [#5].\",\n  \"teleology\": [\n    {\n      \"year\": 2002,\n      \"claim\": \"Established that Myo5c participates in membrane trafficking by linking it to a Rab8/transferrin-receptor recycling compartment, the first functional clue to its cellular role.\",\n      \"evidence\": \"GFP-Myo5c tail dominant-negative expression and immunolocalization in HeLa cells\",\n      \"pmids\": [\"11870218\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Dominant-negative approach does not prove direct motor transport\", \"No demonstration of direct Rab8 binding\", \"Cargo specificity not defined\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Showed Myo5c functions in regulated apical exocytosis by demonstrating it localizes to mature secretory vesicles and is required for stimulated content release and compound fusion.\",\n      \"evidence\": \"Confocal/immunogold EM and adenoviral dominant-negative tail expression with secretory readouts in lacrimal acinar cells\",\n      \"pmids\": [\"18434623\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism of vesicle fusion coupling unresolved\", \"Light-chain/calcium regulation in this context not addressed\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Resolved that Myo5c operates on two functionally distinct compartments with different motility regimes, distinguishing microtubule-independent granule movement from microtubule-dependent tubule transport.\",\n      \"evidence\": \"Live-cell GFP/TIRF imaging, colocalization with chromogranin A/Rab27b and Rab8a, and dominant-negative tail in MCF-7 cells\",\n      \"pmids\": [\"19741097\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular basis of cargo selection between granules and tubules unknown\", \"Direct Rab27b/Rab8a binding not biochemically established\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Extended Myo5c/Rab8 trafficking to a pathophysiological readout by showing it is required for dengue virus egress without affecting intracellular replication.\",\n      \"evidence\": \"Colocalization, dominant-negative tail stable transfection, plaque assay and Rab8 flow cytometry in HepG2 cells\",\n      \"pmids\": [\"19641326\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Does not establish direct interaction with viral components\", \"Step of egress pathway controlled by Myo5c undefined\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Defined the motor mechanochemistry, showing single dimers are non-processive but paired dimers move processively with inter-head cooperativity.\",\n      \"evidence\": \"Steady-state ATPase, ADP dissociation kinetics, and single-molecule tracking of DNA-scaffold-linked Myo5c-HMM dimers\",\n      \"pmids\": [\"24809456\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Physiological mechanism for in vivo dimer coupling unknown\", \"Cargo-induced activation not addressed\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Characterized regulation of human Myo5c motor output by tropomyosin isoforms and identified a small-molecule inhibitor, plus its domain architecture and light-chain binding.\",\n      \"evidence\": \"Recombinant human Myo5c-HMM reconstitution, actin-activated ATPase with tropomyosin cofilaments, motility assays, PBP docking, and tissue immunohistochemistry\",\n      \"pmids\": [\"38606007\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo relevance of specific tropomyosin isoforms not tested\", \"Light-chain composition under physiological conditions undefined\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Implicated Myo5c as a negative regulator of neutrophil activation, expanding its role beyond epithelial trafficking.\",\n      \"evidence\": \"myo5c knockout in HL60 cells with NET formation assay and RNA-seq correlation in patient neutrophils\",\n      \"pmids\": [\"40495980\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No mechanistic pathway linking Myo5c to NET formation established\", \"Single cell-line knockout without rescue\", \"Correlative patient data only\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How Myo5c selectively recognizes and is activated by its distinct membrane cargoes (Rab8a tubules versus Rab27b/chromogranin granules) at the molecular level remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No direct biochemical mapping of tail-Rab interactions\", \"Mechanism of cargo-dependent motor activation unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0003774\", \"supporting_discovery_ids\": [2, 4, 5]},\n      {\"term_id\": \"GO:0140657\", \"supporting_discovery_ids\": [4, 5]},\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [4, 5]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0031410\", \"supporting_discovery_ids\": [1, 2]},\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [4, 5]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-5653656\", \"supporting_discovery_ids\": [0, 1, 2]},\n      {\"term_id\": \"R-HSA-9609507\", \"supporting_discovery_ids\": [0, 2]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"RAB8A\", \"RAB27B\", \"CALM1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}