{"gene":"TG_278870","gene_symbol":"","run_date":"2026-09-08T14:08:30","timeline":{"discoveries":[{"year":2013,"finding":"TgMyoF (class XXII) localizes in the vicinity of the apicoplast during division. Conditional knockdown of TgMyoF severely affects apicoplast turnover and leads to accumulation of the apicoplast and secretory organelles in enlarged residual bodies, establishing an essential role in apicoplast inheritance. Conditional stabilization of the MyoF tail (dominant negative heterodimer) reveals a central role in positioning of the two centrosomes prior to daughter cell formation and in apicoplast segregation.","method":"Conditional knockdown (tetracycline-regulatable), dominant-negative tail overexpression, live-cell fluorescence imaging, phenotypic analysis of apicoplast loss and centrosome positioning","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — conditional KO with defined cellular phenotype plus dominant-negative approach, replicated across multiple experiments in a single rigorous study","pmids":["23695356"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2013,"finding":"TgARO (armadillo repeats only protein) interacts with myosin F, and inhibition of actin polymerization or myosin function results in rhoptry dispersal, establishing that apical positioning of rhoptries is an actomyosin-based process dependent on MyoF.","method":"Co-immunoprecipitation (TgARO–MyoF interaction), conditional depletion of TgARO, actin/myosin inhibitor treatment, fluorescence microscopy of rhoptry positioning","journal":"Cell host & microbe","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal interaction shown by Co-IP, functional complementation with wild-type vs. acylation mutant, loss-of-function with defined invasion phenotype","pmids":["23498954"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2016,"finding":"Structural analysis shows that ARO resembles the myosin-binding domain of the C. elegans myosin chaperone UNC-45, and a conserved patch of aromatic and acidic residues on ARO denotes the putative MyoF-binding site, providing structural basis for the ARO–MyoF interaction in rhoptry positioning.","method":"Crystal structure determination of TgARO, structure-guided mutagenesis of ARM repeats, interaction assays, functional complementation with PfARO","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — crystal structure with functional validation of individual ARM repeats in a single lab; MyoF-binding site is structural prediction supported by the overall structural context but not directly tested by mutagenesis of the binding patch","pmids":["26769898"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2017,"finding":"TgUNC acts as a chaperone dedicated to the folding, assembly and function of all Toxoplasma myosins including MyoF; conditional ablation of TgUNC phenocopies MyoF loss (organelle positioning and apicoplast inheritance defects), demonstrating that MyoF function depends on TgUNC-mediated chaperone activity.","method":"Conditional knockdown of TgUNC, phenotypic analysis recapitulating known myosin phenotypes, genetic epistasis","journal":"Nature communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — conditional KO with defined phenotype recapitulating multiple myosin knockdowns; single lab, pathway placement by phenome comparison","pmids":["28593938"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2023,"finding":"Loss of TgMyoF and actin leads to accumulation of apicoplast vesicles in the cytosol and failure of the apicoplast to associate with parasite centrosomes during division. Live-cell imaging shows that the apicoplast exhibits highly dynamic branched, U-shaped and linear morphologies during division that are dependent on TgMyoF and F-actin, establishing that this actomyosin system drives apicoplast elongation and apicoplast–centrosome association.","method":"Conditional depletion of TgMyoF, live-cell fluorescence imaging of apicoplast morphology dynamics, actin perturbation experiments","journal":"mBio","confidence":"High","confidence_rationale":"Tier 2 / Strong — conditional KO with defined cellular phenotype, live-cell imaging of dynamics, two orthogonal perturbations (MyoF depletion and actin disruption) giving consistent results","pmids":["37732764"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2024,"finding":"MyoF displays a dynamic, filamentous-like organization in the parasite cytosol (not associated with Golgi, apicoplast, or dense granule surfaces). Loss of MyoF causes dramatic rearrangement of the actin cytoskeleton in interphase parasites accompanied by significantly reduced actin dynamics, while actin organization during replication and motility is unaffected, establishing MyoF as an actin-organizing protein that facilitates cargo movement via an unconventional mechanism rather than canonical cargo-motor transport.","method":"Live-cell imaging of MyoF-EmeraldFP, conditional loss-of-function, quantitative analysis of actin dynamics and cytoskeletal organization","journal":"Molecular biology of the cell","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — live-cell imaging plus conditional KO with two orthogonal readouts (localization and actin dynamics), single lab","pmids":["38416592"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2026,"finding":"Purified MyoF lacking its WD40 tail domain is dimeric and can bind and translocate actin in an in vitro motility assay but is non-processive as a single molecule. Full-length MyoF is processive as single molecules on Toxoplasma actin and jasplakinolide-stabilized actin bundles. The WD40 domain oligomerizes (shown by EM and size-exclusion chromatography) to form higher-order complexes of multiple dimers, explaining processive behavior of the full-length motor. MyoF binds microtubules through its WD40 domain and can slide actin filaments relative to microtubules, supporting a model where MyoF oligomers drive actin dynamics by translocating filaments relative to the parasite cytoskeleton.","method":"In vitro purification of recombinant MyoF constructs, in vitro actin motility assay, single-molecule imaging, negative-stain electron microscopy, quantitative size-exclusion chromatography, microtubule-binding and sliding assays, domain-deletion mutagenesis (ΔWD40 tail)","journal":"Journal of cell science","confidence":"High","confidence_rationale":"Tier 1 / Strong — multiple orthogonal in vitro biochemical and structural methods (motility assay, single-molecule imaging, EM, SEC, microtubule-sliding assay) with domain-deletion mutagenesis in one rigorous study","pmids":["41995028"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]}],"current_model":"Toxoplasma gondii Myosin F (MyoF) is a class XXII, alveolate-unique molecular motor whose WD40 β-propeller tail domain mediates oligomerization into processive higher-order complexes that translocate actin filaments relative to the parasite cytoskeleton (including relative to microtubules), thereby organizing a dynamic cytosolic F-actin network; this actomyosin activity drives apicoplast elongation and centrosome association to ensure apicoplast inheritance, mediates vesicle/organelle trafficking through the endomembrane system, and—via interaction with the rhoptry-membrane scaffold protein TgARO—positions rhoptries apically for host-cell invasion, with MyoF itself requiring the TgUNC chaperone for proper folding and assembly."},"narrative":{"mechanistic_narrative":"Toxoplasma gondii Myosin F (MyoF) is a class XXII, alveolate-specific actin-based motor that organizes a dynamic cytosolic F-actin network to drive organelle inheritance and apical secretory-organelle positioning during parasite division and invasion [PMID:23695356, PMID:37732764, PMID:41995028]. Biochemically, MyoF constructs lacking the C-terminal WD40 β-propeller tail form dimers that bind and translocate actin but are non-processive, whereas the WD40 domain oligomerizes multiple dimers into higher-order complexes that render the full-length motor processive on parasite actin filaments and bundles; the same WD40 domain binds microtubules and enables MyoF to slide actin filaments relative to microtubules [PMID:41995028]. Consistent with this filament-translocating (rather than canonical cargo-transport) mode of action, MyoF adopts a dynamic filamentous cytosolic organization, and its loss remodels the interphase actin cytoskeleton and reduces actin dynamics [PMID:38416592]. Functionally, MyoF-driven actomyosin activity elongates the apicoplast and promotes its association with the centrosomes, ensuring apicoplast inheritance and preventing accumulation of organelles in enlarged residual bodies [PMID:23695356, PMID:37732764]. Through interaction with the rhoptry-associated armadillo-repeat protein TgARO, MyoF positions rhoptries apically, an actomyosin-dependent step required for host-cell invasion [PMID:23498954, PMID:26769898]. MyoF folding and assembly depend on the dedicated myosin chaperone TgUNC, whose loss phenocopies MyoF depletion [PMID:28593938].","teleology":[{"year":2013,"claim":"Establishing that a class XXII myosin is required for organelle inheritance answered whether an actomyosin motor governs apicoplast segregation and centrosome positioning in a dividing parasite.","evidence":"Conditional knockdown and dominant-negative tail overexpression with live-cell imaging of apicoplast and centrosome phenotypes in T. gondii","pmids":["23695356"],"confidence":"High","gaps":["Did not resolve whether MyoF acts as a cargo transporter or an actin-organizing motor","Molecular partners at the apicoplast/centrosome not identified","In vitro motor activity not measured"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2013,"claim":"Linking MyoF to the rhoptry scaffold TgARO answered how apical secretory organelles are positioned, showing rhoptry localization is an actomyosin-dependent process required for invasion.","evidence":"Co-immunoprecipitation of TgARO with MyoF, conditional TgARO depletion, actin/myosin inhibitor treatment, and rhoptry-positioning microscopy in T. gondii","pmids":["23498954"],"confidence":"High","gaps":["Direct MyoF-binding interface on TgARO not mapped","Whether the interaction is direct or bridged not established"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2016,"claim":"The crystal structure of TgARO provided a structural rationale for the ARO–MyoF interaction by identifying a candidate myosin-binding surface resembling the UNC-45 myosin-binding domain.","evidence":"Crystal structure of TgARO with structure-guided ARM-repeat mutagenesis, interaction assays, and PfARO complementation in T. gondii","pmids":["26769898"],"confidence":"Medium","gaps":["Predicted MyoF-binding patch not directly tested by mutagenesis of that patch","Structure of the MyoF domain that engages ARO not determined"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2017,"claim":"Identifying TgUNC as a dedicated myosin chaperone answered how MyoF and other parasite myosins achieve proper folding and assembly.","evidence":"Conditional TgUNC knockdown with phenotypic recapitulation of myosin loss and genetic epistasis in T. gondii","pmids":["28593938"],"confidence":"Medium","gaps":["Direct chaperone binding to MyoF not biochemically demonstrated","Pathway placement rests on phenome comparison rather than direct interaction"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2023,"claim":"Live-cell imaging of apicoplast dynamics answered what MyoF actually does at the organelle, showing the actomyosin system drives apicoplast elongation and centrosome association rather than passive tethering.","evidence":"Conditional MyoF depletion, actin perturbation, and live-cell imaging of dynamic apicoplast morphologies in T. gondii","pmids":["37732764"],"confidence":"High","gaps":["Molecular link between MyoF-driven actin and the apicoplast membrane not identified","How centrosome capture is achieved mechanistically unresolved"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2024,"claim":"Characterizing MyoF's cytosolic organization and effect on actin resolved its mode of action, reframing it as an actin-organizing protein acting via an unconventional mechanism rather than a canonical cargo motor.","evidence":"Live-cell imaging of MyoF-EmeraldFP with conditional loss-of-function and quantitative analysis of actin dynamics and cytoskeletal organization in T. gondii","pmids":["38416592"],"confidence":"Medium","gaps":["The unconventional cargo-movement mechanism not defined at molecular level","Why actin organization during replication and motility is spared unexplained"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2026,"claim":"In vitro reconstitution answered how MyoF achieves processivity and organizes actin, showing WD40-mediated oligomerization builds processive higher-order motors that slide actin filaments relative to microtubules.","evidence":"Recombinant MyoF purification, in vitro motility and single-molecule assays, negative-stain EM, size-exclusion chromatography, microtubule-binding/sliding assays, and ΔWD40 domain-deletion mutagenesis","pmids":["41995028"],"confidence":"High","gaps":["Stoichiometry and regulation of oligomer assembly in vivo not established","How TgARO and TgUNC interface with the oligomeric motor biochemically not tested"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":null,"claim":"How MyoF-generated actin dynamics are physically coupled to specific cargo membranes (apicoplast, rhoptries) and how oligomerization is regulated in the living parasite remain open.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No identified membrane adaptor linking MyoF-driven actin to organelle surfaces","Regulation of WD40 oligomerization in vivo unknown","Direct biochemical demonstration of the MyoF–TgARO and MyoF–TgUNC interfaces lacking"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]}],"mechanism_profile":{"molecular_activity":[],"localization":[],"pathway":[],"complexes":["WD40-mediated higher-order oligomer of MyoF dimers [#6]"],"partners":["TGARO (ARMADILLO-REPEAT PROTEIN, RHOPTRY POSITIONING) [#1, #2]","TGUNC (MYOSIN FOLDING/ASSEMBLY CHAPERONE) [#3]","ACTIN FILAMENTS/BUNDLES (MOTOR SUBSTRATE) [#5, #6]","MICROTUBULES (WD40 TAIL BINDING) [#6]"],"other_free_text":["MyoF is a class XXII, alveolate-unique myosin [#0].","Full-length MyoF is processive as single molecules on Toxoplasma actin, whereas the WD40-truncated dimer binds and translocates actin but is non-processive [#6].","MyoF slides actin filaments relative to microtubules, consistent with a filament-translocating rather than canonical cargo-transport mechanism [#5, #6]."]},"strain_conflicts":[]},"prefetch_data":{},"corpus_meta":[{"pmid":"28593938","id":"PMC_28593938","title":"Myosin-dependent cell-cell communication controls synchronicity of division in acute and chronic stages of Toxoplasma gondii.","date":"2017","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/28593938","citation_count":90,"is_preprint":false},{"pmid":"23695356","id":"PMC_23695356","title":"Toxoplasma gondii myosin F, an essential motor for centrosomes positioning and apicoplast inheritance.","date":"2013","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/23695356","citation_count":77,"is_preprint":false},{"pmid":"23498954","id":"PMC_23498954","title":"The Toxoplasma protein ARO mediates the apical positioning of rhoptry organelles, a prerequisite for host cell invasion.","date":"2013","source":"Cell host & microbe","url":"https://pubmed.ncbi.nlm.nih.gov/23498954","citation_count":77,"is_preprint":false},{"pmid":"26769898","id":"PMC_26769898","title":"Structural and functional dissection of Toxoplasma gondii armadillo repeats only protein.","date":"2016","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/26769898","citation_count":26,"is_preprint":false},{"pmid":"37732764","id":"PMC_37732764","title":"F-actin and myosin F control apicoplast elongation dynamics which drive apicoplast-centrosome association in Toxoplasma gondii.","date":"2023","source":"mBio","url":"https://pubmed.ncbi.nlm.nih.gov/37732764","citation_count":13,"is_preprint":false},{"pmid":"38416592","id":"PMC_38416592","title":"Myosin F controls actin organization and dynamics in Toxoplasma gondii.","date":"2024","source":"Molecular biology of the cell","url":"https://pubmed.ncbi.nlm.nih.gov/38416592","citation_count":6,"is_preprint":false},{"pmid":"36711828","id":"PMC_36711828","title":"F-actin and Myosin F control apicoplast elongation dynamics which drive apicoplast-centrosome association in Toxoplasma gondii.","date":"2023","source":"bioRxiv : the preprint server for biology","url":"https://pubmed.ncbi.nlm.nih.gov/36711828","citation_count":2,"is_preprint":false},{"pmid":"41995028","id":"PMC_41995028","title":"Molecular features of myosin F adapted for driving actin flows in Toxoplasma gondii.","date":"2026","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/41995028","citation_count":0,"is_preprint":false},{"pmid":"37331576","id":"PMC_37331576","title":"Knocking down the expression of the molecular motors, myosin A, C and F genes in Toxoplasma gondii to decrease the parasite virulence.","date":"2023","source":"Experimental parasitology","url":"https://pubmed.ncbi.nlm.nih.gov/37331576","citation_count":0,"is_preprint":false},{"pmid":"41256612","id":"PMC_41256612","title":"Molecular features of Myosin F adapted for driving actin flows in Toxoplasma gondii.","date":"2025","source":"bioRxiv : the preprint server for biology","url":"https://pubmed.ncbi.nlm.nih.gov/41256612","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":6772,"output_tokens":2298,"usd":0.027393,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":7820,"output_tokens":3725,"usd":0.066113,"stage2_stop_reason":"end_turn"},"total_usd":0.093506,"stage1_batch_id":"msgbatch_015xmQKDRqnTh9anE4m7LiD6","stage2_batch_id":"msgbatch_01JLpkFTVhoQBNgqHCKFrVVU","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2013,\n      \"finding\": \"TgMyoF (class XXII) localizes in the vicinity of the apicoplast during division. Conditional knockdown of TgMyoF severely affects apicoplast turnover and leads to accumulation of the apicoplast and secretory organelles in enlarged residual bodies, establishing an essential role in apicoplast inheritance. Conditional stabilization of the MyoF tail (dominant negative heterodimer) reveals a central role in positioning of the two centrosomes prior to daughter cell formation and in apicoplast segregation.\",\n      \"method\": \"Conditional knockdown (tetracycline-regulatable), dominant-negative tail overexpression, live-cell fluorescence imaging, phenotypic analysis of apicoplast loss and centrosome positioning\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — conditional KO with defined cellular phenotype plus dominant-negative approach, replicated across multiple experiments in a single rigorous study\",\n      \"pmids\": [\"23695356\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"TgARO (armadillo repeats only protein) interacts with myosin F, and inhibition of actin polymerization or myosin function results in rhoptry dispersal, establishing that apical positioning of rhoptries is an actomyosin-based process dependent on MyoF.\",\n      \"method\": \"Co-immunoprecipitation (TgARO–MyoF interaction), conditional depletion of TgARO, actin/myosin inhibitor treatment, fluorescence microscopy of rhoptry positioning\",\n      \"journal\": \"Cell host & microbe\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal interaction shown by Co-IP, functional complementation with wild-type vs. acylation mutant, loss-of-function with defined invasion phenotype\",\n      \"pmids\": [\"23498954\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Structural analysis shows that ARO resembles the myosin-binding domain of the C. elegans myosin chaperone UNC-45, and a conserved patch of aromatic and acidic residues on ARO denotes the putative MyoF-binding site, providing structural basis for the ARO–MyoF interaction in rhoptry positioning.\",\n      \"method\": \"Crystal structure determination of TgARO, structure-guided mutagenesis of ARM repeats, interaction assays, functional complementation with PfARO\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — crystal structure with functional validation of individual ARM repeats in a single lab; MyoF-binding site is structural prediction supported by the overall structural context but not directly tested by mutagenesis of the binding patch\",\n      \"pmids\": [\"26769898\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"TgUNC acts as a chaperone dedicated to the folding, assembly and function of all Toxoplasma myosins including MyoF; conditional ablation of TgUNC phenocopies MyoF loss (organelle positioning and apicoplast inheritance defects), demonstrating that MyoF function depends on TgUNC-mediated chaperone activity.\",\n      \"method\": \"Conditional knockdown of TgUNC, phenotypic analysis recapitulating known myosin phenotypes, genetic epistasis\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — conditional KO with defined phenotype recapitulating multiple myosin knockdowns; single lab, pathway placement by phenome comparison\",\n      \"pmids\": [\"28593938\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Loss of TgMyoF and actin leads to accumulation of apicoplast vesicles in the cytosol and failure of the apicoplast to associate with parasite centrosomes during division. Live-cell imaging shows that the apicoplast exhibits highly dynamic branched, U-shaped and linear morphologies during division that are dependent on TgMyoF and F-actin, establishing that this actomyosin system drives apicoplast elongation and apicoplast–centrosome association.\",\n      \"method\": \"Conditional depletion of TgMyoF, live-cell fluorescence imaging of apicoplast morphology dynamics, actin perturbation experiments\",\n      \"journal\": \"mBio\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — conditional KO with defined cellular phenotype, live-cell imaging of dynamics, two orthogonal perturbations (MyoF depletion and actin disruption) giving consistent results\",\n      \"pmids\": [\"37732764\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"MyoF displays a dynamic, filamentous-like organization in the parasite cytosol (not associated with Golgi, apicoplast, or dense granule surfaces). Loss of MyoF causes dramatic rearrangement of the actin cytoskeleton in interphase parasites accompanied by significantly reduced actin dynamics, while actin organization during replication and motility is unaffected, establishing MyoF as an actin-organizing protein that facilitates cargo movement via an unconventional mechanism rather than canonical cargo-motor transport.\",\n      \"method\": \"Live-cell imaging of MyoF-EmeraldFP, conditional loss-of-function, quantitative analysis of actin dynamics and cytoskeletal organization\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — live-cell imaging plus conditional KO with two orthogonal readouts (localization and actin dynamics), single lab\",\n      \"pmids\": [\"38416592\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"Purified MyoF lacking its WD40 tail domain is dimeric and can bind and translocate actin in an in vitro motility assay but is non-processive as a single molecule. Full-length MyoF is processive as single molecules on Toxoplasma actin and jasplakinolide-stabilized actin bundles. The WD40 domain oligomerizes (shown by EM and size-exclusion chromatography) to form higher-order complexes of multiple dimers, explaining processive behavior of the full-length motor. MyoF binds microtubules through its WD40 domain and can slide actin filaments relative to microtubules, supporting a model where MyoF oligomers drive actin dynamics by translocating filaments relative to the parasite cytoskeleton.\",\n      \"method\": \"In vitro purification of recombinant MyoF constructs, in vitro actin motility assay, single-molecule imaging, negative-stain electron microscopy, quantitative size-exclusion chromatography, microtubule-binding and sliding assays, domain-deletion mutagenesis (ΔWD40 tail)\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — multiple orthogonal in vitro biochemical and structural methods (motility assay, single-molecule imaging, EM, SEC, microtubule-sliding assay) with domain-deletion mutagenesis in one rigorous study\",\n      \"pmids\": [\"41995028\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    }\n  ],\n  \"current_model\": \"Toxoplasma gondii Myosin F (MyoF) is a class XXII, alveolate-unique molecular motor whose WD40 β-propeller tail domain mediates oligomerization into processive higher-order complexes that translocate actin filaments relative to the parasite cytoskeleton (including relative to microtubules), thereby organizing a dynamic cytosolic F-actin network; this actomyosin activity drives apicoplast elongation and centrosome association to ensure apicoplast inheritance, mediates vesicle/organelle trafficking through the endomembrane system, and—via interaction with the rhoptry-membrane scaffold protein TgARO—positions rhoptries apically for host-cell invasion, with MyoF itself requiring the TgUNC chaperone for proper folding and assembly.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"Toxoplasma gondii Myosin F (MyoF) is a class XXII, alveolate-specific actin-based motor that organizes a dynamic cytosolic F-actin network to drive organelle inheritance and apical secretory-organelle positioning during parasite division and invasion [#0, #4, #6]. Biochemically, MyoF constructs lacking the C-terminal WD40 \\u03b2-propeller tail form dimers that bind and translocate actin but are non-processive, whereas the WD40 domain oligomerizes multiple dimers into higher-order complexes that render the full-length motor processive on parasite actin filaments and bundles; the same WD40 domain binds microtubules and enables MyoF to slide actin filaments relative to microtubules [#6]. Consistent with this filament-translocating (rather than canonical cargo-transport) mode of action, MyoF adopts a dynamic filamentous cytosolic organization, and its loss remodels the interphase actin cytoskeleton and reduces actin dynamics [#5]. Functionally, MyoF-driven actomyosin activity elongates the apicoplast and promotes its association with the centrosomes, ensuring apicoplast inheritance and preventing accumulation of organelles in enlarged residual bodies [#0, #4]. Through interaction with the rhoptry-associated armadillo-repeat protein TgARO, MyoF positions rhoptries apically, an actomyosin-dependent step required for host-cell invasion [#1, #2]. MyoF folding and assembly depend on the dedicated myosin chaperone TgUNC, whose loss phenocopies MyoF depletion [#3].\",\n  \"teleology\": [\n    {\n      \"year\": 2013,\n      \"claim\": \"Establishing that a class XXII myosin is required for organelle inheritance answered whether an actomyosin motor governs apicoplast segregation and centrosome positioning in a dividing parasite.\",\n      \"evidence\": \"Conditional knockdown and dominant-negative tail overexpression with live-cell imaging of apicoplast and centrosome phenotypes in T. gondii\",\n      \"pmids\": [\"23695356\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not resolve whether MyoF acts as a cargo transporter or an actin-organizing motor\", \"Molecular partners at the apicoplast/centrosome not identified\", \"In vitro motor activity not measured\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Linking MyoF to the rhoptry scaffold TgARO answered how apical secretory organelles are positioned, showing rhoptry localization is an actomyosin-dependent process required for invasion.\",\n      \"evidence\": \"Co-immunoprecipitation of TgARO with MyoF, conditional TgARO depletion, actin/myosin inhibitor treatment, and rhoptry-positioning microscopy in T. gondii\",\n      \"pmids\": [\"23498954\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct MyoF-binding interface on TgARO not mapped\", \"Whether the interaction is direct or bridged not established\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"The crystal structure of TgARO provided a structural rationale for the ARO\\u2013MyoF interaction by identifying a candidate myosin-binding surface resembling the UNC-45 myosin-binding domain.\",\n      \"evidence\": \"Crystal structure of TgARO with structure-guided ARM-repeat mutagenesis, interaction assays, and PfARO complementation in T. gondii\",\n      \"pmids\": [\"26769898\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Predicted MyoF-binding patch not directly tested by mutagenesis of that patch\", \"Structure of the MyoF domain that engages ARO not determined\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Identifying TgUNC as a dedicated myosin chaperone answered how MyoF and other parasite myosins achieve proper folding and assembly.\",\n      \"evidence\": \"Conditional TgUNC knockdown with phenotypic recapitulation of myosin loss and genetic epistasis in T. gondii\",\n      \"pmids\": [\"28593938\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct chaperone binding to MyoF not biochemically demonstrated\", \"Pathway placement rests on phenome comparison rather than direct interaction\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Live-cell imaging of apicoplast dynamics answered what MyoF actually does at the organelle, showing the actomyosin system drives apicoplast elongation and centrosome association rather than passive tethering.\",\n      \"evidence\": \"Conditional MyoF depletion, actin perturbation, and live-cell imaging of dynamic apicoplast morphologies in T. gondii\",\n      \"pmids\": [\"37732764\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular link between MyoF-driven actin and the apicoplast membrane not identified\", \"How centrosome capture is achieved mechanistically unresolved\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Characterizing MyoF's cytosolic organization and effect on actin resolved its mode of action, reframing it as an actin-organizing protein acting via an unconventional mechanism rather than a canonical cargo motor.\",\n      \"evidence\": \"Live-cell imaging of MyoF-EmeraldFP with conditional loss-of-function and quantitative analysis of actin dynamics and cytoskeletal organization in T. gondii\",\n      \"pmids\": [\"38416592\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"The unconventional cargo-movement mechanism not defined at molecular level\", \"Why actin organization during replication and motility is spared unexplained\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"In vitro reconstitution answered how MyoF achieves processivity and organizes actin, showing WD40-mediated oligomerization builds processive higher-order motors that slide actin filaments relative to microtubules.\",\n      \"evidence\": \"Recombinant MyoF purification, in vitro motility and single-molecule assays, negative-stain EM, size-exclusion chromatography, microtubule-binding/sliding assays, and \\u0394WD40 domain-deletion mutagenesis\",\n      \"pmids\": [\"41995028\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Stoichiometry and regulation of oligomer assembly in vivo not established\", \"How TgARO and TgUNC interface with the oligomeric motor biochemically not tested\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How MyoF-generated actin dynamics are physically coupled to specific cargo membranes (apicoplast, rhoptries) and how oligomerization is regulated in the living parasite remain open.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No identified membrane adaptor linking MyoF-driven actin to organelle surfaces\", \"Regulation of WD40 oligomerization in vivo unknown\", \"Direct biochemical demonstration of the MyoF\\u2013TgARO and MyoF\\u2013TgUNC interfaces lacking\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0000146\", \"supporting_discovery_ids\": [6], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]},\n      {\"term_id\": \"GO:0051015\", \"supporting_discovery_ids\": [5, 6], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]},\n      {\"term_id\": \"GO:0008017\", \"supporting_discovery_ids\": [6], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]},\n      {\"term_id\": \"GO:0042803\", \"supporting_discovery_ids\": [6], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [5], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]},\n      {\"term_id\": \"GO:0009536\", \"supporting_discovery_ids\": [0], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"GO:0007059\", \"supporting_discovery_ids\": [0, 4], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]},\n      {\"term_id\": \"GO:0030036\", \"supporting_discovery_ids\": [5], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]},\n      {\"term_id\": \"GO:0051656\", \"supporting_discovery_ids\": [1, 4], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]}\n    ],\n    \"complexes\": [\n      \"WD40-mediated higher-order oligomer of MyoF dimers [#6]\"\n    ],\n    \"partners\": [\n      \"TgARO (armadillo-repeat protein, rhoptry positioning) [#1, #2]\",\n      \"TgUNC (myosin folding/assembly chaperone) [#3]\",\n      \"Actin filaments/bundles (motor substrate) [#5, #6]\",\n      \"Microtubules (WD40 tail binding) [#6]\"\n    ],\n    \"other_free_text\": [\n      \"MyoF is a class XXII, alveolate-unique myosin [#0].\",\n      \"Full-length MyoF is processive as single molecules on Toxoplasma actin, whereas the WD40-truncated dimer binds and translocates actin but is non-processive [#6].\",\n      \"MyoF slides actin filaments relative to microtubules, consistent with a filament-translocating rather than canonical cargo-transport mechanism [#5, #6].\"\n    ]\n  },\n  \"strain_conflicts\": []\n}\n```","audit_flag":null,"strains":[{"strain":"ME49","gene_id":"TGME49_278870","name":null,"product":"myosin F","uniprot_acc":"A0A125YRU2","chromosome":"TGME49_chrXII","start_pos":5793484},{"strain":"GT1","gene_id":"TGGT1_278870","name":null,"product":"myosin F","uniprot_acc":null,"chromosome":"TGGT1_chrXII","start_pos":5709485},{"strain":"RH88","gene_id":"TGRH88_065010","name":null,"product":"myosin F","uniprot_acc":null,"chromosome":"CM023093","start_pos":5872812},{"strain":"VEG","gene_id":"TGVEG_278870","name":null,"product":"myosin F","uniprot_acc":"B9QML2","chromosome":"TGVEG_chrXII","start_pos":5799662}]}