{"gene":"TG_231640","gene_symbol":"ALV1","run_date":"2026-09-10T21:09:19","timeline":{"discoveries":[{"year":2001,"finding":"TgIMC1 (ALV1) is a component of the subpellicular network, a filamentous membrane skeletal structure composed of interwoven 8–10 nm filaments associated with the cytoplasmic face of the pellicle of Toxoplasma gondii. TgIMC1 has predicted extended coiled-coil domains and is homologous to articulins, the major components of the membrane skeleton of algae and free-living protists.","method":"Electron microscopy of detergent-extracted tachyzoites; protein identification and sequence analysis","journal":"Molecular and biochemical parasitology","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct EM visualization of filamentous network combined with protein identification; foundational study replicated by subsequent work","pmids":["11420112"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2002,"finding":"An IMC1–YFP fusion protein was used to visualize daughter scaffold (inner membrane complex) formation in live T. gondii by time-lapse video microscopy, demonstrating that the IMC1-based subpellicular network is assembled during endodyogeny and can also form during production of multiple daughters.","method":"Live-cell time-lapse fluorescence microscopy using IMC1–YFP fusion protein","journal":"Molecular biology of the cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct live-imaging with functional reporter; foundational study widely replicated","pmids":["11854415"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2002,"finding":"TgIMC1 (ALV1) undergoes proteolytic processing of its carboxyl terminus during daughter parasite maturation. In immature parasites the network is detergent-soluble; in mature parasites it becomes entirely detergent-resistant. Conversion from detergent-labile to detergent-resistant network is coupled to proteolytic cleavage of the TgIMC1 C-terminus. A single cysteine residue in the C-terminus is essential for this processing event.","method":"Detergent fractionation assays; site-directed mutagenesis of TgIMC1 cysteine residue; immunoblotting to detect processing","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro mutagenesis combined with biochemical fractionation assay in a single focused study with rigorous controls","pmids":["12177058"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2017,"finding":"IMC1 (ALV1) is present in both mother and daughter cytoskeletons during asexual tachyzoite division, is absent from developing macro- and microgametocytes, and is present in sporozoites. IMC1 is thus present across multiple life cycle stages but absent from gametocytes, indicating a stage-specific role distinct from other IMC alveolins.","method":"Immunofluorescence assay (IFA) across multiple life-cycle stages of T. gondii and Sarcocystis neurona","journal":"mSphere","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct localization across multiple stages in a single lab study using IFA","pmids":["29062899"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2021,"finding":"Double knockout of metacaspase-1 (MCA1) and metacaspase-2 (MCA2) in T. gondii results in detergent-soluble IMC1, indicating that MCA1 and MCA2 are required for IMC1 (ALV1) maturation (conversion to detergent-resistant form) and for normal endodyogeny.","method":"Double-knockout of MCA1/MCA2 in RHΔku80; detergent fractionation to assess IMC1 solubility; IFA to assess IMC1 expression and endodyogeny","journal":"Parasites & vectors","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic knockout with biochemical readout (detergent fractionation) linking metacaspases to IMC1 maturation, single lab","pmids":["34384491"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":"RH"}]},{"year":2025,"finding":"IMT1, a protein that bridges the IMC and subpellicular microtubules, binds directly to the cytoskeletal IMC proteins IMC1 (ALV1), IMC18, and IMC24, as shown by deletion analyses and proximity labeling.","method":"Proximity labeling (BioID), co-immunoprecipitation/pulldown, deletion analyses","journal":"Molecular biology of the cell","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct binding shown with proximity labeling and deletion analyses in a single focused study","pmids":["40768331"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2025,"finding":"Disruption of IMC1 (ALV1) causes severe morphological defects, loss of cytoskeletal integrity, and defects in invasion, replication, and egress. In a type II strain, IMC1 loss causes dramatic reduction in infectivity and complete failure to establish chronic infection in vivo.","method":"Gene knockout; morphological analysis; invasion, replication, and egress assays; mouse infection studies","journal":"PLoS pathogens","confidence":"High","confidence_rationale":"Tier 2 / Strong — clean gene knockout with multiple orthogonal functional readouts (morphology, invasion, replication, egress, in vivo infection) replicated across strains and published in peer-reviewed journal","pmids":["42113865","40501855"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":"RH"},{"species":"Toxoplasma gondii","strain":"ME49"}]},{"year":2025,"finding":"IMC1 (ALV1) is loaded onto forming daughter buds together with IMC4, but later than IMC3, IMC6, and IMC10. A key subregion of the alveolin domain is sufficient for IMC targeting and is also required for IMC1 function. IMC1 interacts directly with IMC4, and loss of IMC1 results in mislocalization of IMC4 specifically in forming daughter buds.","method":"Fluorescent protein tagging with time-resolved IFA; deletion analysis; mutagenesis; co-immunoprecipitation/pulldown to establish direct IMC1–IMC4 interaction; IFA of IMC4 localization in IMC1 knockout","journal":"PLoS pathogens","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — direct interaction confirmed by pulldown, deletion/mutagenesis analysis for functional domain mapping, and IFA for localization consequence; peer-reviewed publication","pmids":["42113865","40501855"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":"RH"},{"species":"Toxoplasma gondii","strain":"ME49"}]}],"current_model":"ALV1/IMC1 is the founding alveolin and a central structural component of the Toxoplasma gondii subpellicular network, forming 8–10 nm filaments on the cytoplasmic face of the IMC; it is loaded onto daughter buds during endodyogeny (with IMC4, but after IMC3/IMC6/IMC10), undergoes metacaspase-dependent proteolytic processing of its cysteine-containing C-terminus to transition from a detergent-labile to a detergent-resistant mature network, directly binds IMC4 (whose localization in daughter buds depends on IMC1) and the IMC-microtubule bridging protein IMT1, and is essential for cytoskeletal integrity, invasion, replication, egress, and establishment of chronic infection in vivo."},"narrative":{"mechanistic_narrative":"ALV1 (TgIMC1) is the founding alveolin and a central structural component of the subpellicular network, a filamentous membrane skeleton of interwoven 8–10 nm filaments associated with the cytoplasmic face of the Toxoplasma gondii pellicle [PMID:11420112]. It is assembled onto daughter cytoskeletons during endodyogeny and multiple-daughter formation [PMID:11854415], being loaded onto forming daughter buds together with IMC4 but later than IMC3, IMC6, and IMC10; a subregion of its alveolin domain is sufficient for IMC targeting and required for its function, and it binds IMC4 directly, with IMC1 loss mislocalizing IMC4 in daughter buds [PMID:42113865, PMID:40501855]. Network maturation depends on proteolytic processing of the ALV1 C-terminus—requiring a single C-terminal cysteine—that converts the detergent-labile immature network into a detergent-resistant mature form [PMID:12177058], a maturation step that requires the metacaspases MCA1 and MCA2 in RH [PMID:34384491]. ALV1 also binds directly to IMT1, the protein bridging the IMC to subpellicular microtubules [PMID:40768331]. ALV1 is expressed across tachyzoite, sporozoite, and other stages but absent from gametocytes [PMID:29062899], and its disruption causes loss of cytoskeletal integrity with defects in invasion, replication, and egress, abolishing chronic infection in a type II (ME49) strain in vivo [PMID:42113865, PMID:40501855]. Beyond these findings, nothing further about ALV1 is established in the available corpus.","teleology":[{"year":2001,"claim":"Establishing that ALV1/TgIMC1 is a genuine structural component of the parasite membrane skeleton answered whether the pellicle-associated filamentous network had defined protein constituents.","evidence":"Electron microscopy of detergent-extracted tachyzoites with protein identification and sequence analysis","pmids":["11420112"],"confidence":"High","gaps":["Filament assembly mechanism not resolved","Partner proteins within the network not yet identified"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2002,"claim":"Live imaging of an IMC1–YFP reporter answered when and how the network is built, showing it is assembled de novo during endodyogeny and multiple-daughter formation.","evidence":"Live-cell time-lapse fluorescence microscopy of an IMC1–YFP fusion in T. gondii","pmids":["11854415"],"confidence":"High","gaps":["Targeting determinants for daughter bud loading not defined","Order relative to other IMC proteins unknown"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2002,"claim":"Identifying C-terminal proteolytic processing coupled to a detergent-labile-to-resistant transition answered how the immature network matures into a stable skeleton.","evidence":"Detergent fractionation plus site-directed mutagenesis of a C-terminal cysteine and immunoblotting","pmids":["12177058"],"confidence":"High","gaps":["Protease responsible not identified in this study","Molecular basis of detergent resistance unresolved"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2017,"claim":"Mapping IMC1 expression across the life cycle answered whether its role is stage-specific, showing presence in tachyzoites and sporozoites but absence from gametocytes.","evidence":"Immunofluorescence across multiple life-cycle stages","pmids":["29062899"],"confidence":"Medium","gaps":["Functional consequence of gametocyte absence not tested","Stage-specific regulation mechanism unknown"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2021,"claim":"Genetic loss of MCA1/MCA2 identified the enzymes required for IMC1 maturation, linking metacaspase activity to conversion of the network to its detergent-resistant form.","evidence":"MCA1/MCA2 double knockout in RHΔku80 with detergent fractionation and IFA","pmids":["34384491"],"confidence":"Medium","gaps":["Direct cleavage of IMC1 by metacaspases not demonstrated","Whether processing is direct or indirect not resolved"],"attribution":[{"species":"Toxoplasma gondii","strain":"RH"}]},{"year":2025,"claim":"Direct binding of IMT1 to IMC1 answered how the alveolin network physically couples to subpellicular microtubules.","evidence":"Proximity labeling (BioID), co-immunoprecipitation/pulldown, and deletion analyses","pmids":["40768331"],"confidence":"Medium","gaps":["Binding interface on IMC1 not mapped","Functional consequence of the IMC1–IMT1 interaction for microtubule anchoring not tested"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2025,"claim":"Knockout combined with domain mapping and interaction analysis established ALV1 as essential for cytoskeletal integrity and defined its daughter-bud loading order and its direct role in positioning IMC4.","evidence":"Gene knockout with morphology, invasion, replication, egress and mouse infection assays; fluorescent tagging with time-resolved IFA, deletion/mutagenesis, and IMC1–IMC4 pulldown","pmids":["42113865","40501855"],"confidence":"High","gaps":["Structure of the alveolin targeting subregion not determined","How IMC1 nucleates or organizes the broader network hierarchy not resolved"],"attribution":[{"species":"Toxoplasma gondii","strain":"RH"},{"species":"Toxoplasma gondii","strain":"ME49"}]}],"mechanism_profile":{"molecular_activity":[],"localization":[],"pathway":[],"complexes":["Subpellicular network / inner membrane complex alveolin meshwork"],"partners":["IMC4 (DIRECT BINDING)","IMT1 (DIRECT BINDING, IMC-MICROTUBULE BRIDGE)","MCA1/MCA2 METACASPASES (REQUIRED FOR MATURATION)"],"other_free_text":["Founding alveolin; predicted extended coiled-coil domains; homologous to articulins [#0]","C-terminal proteolytic processing requiring a single cysteine converts the detergent-labile network to detergent-resistant [#2]","Loaded onto daughter buds with IMC4 but after IMC3/IMC6/IMC10; alveolin subregion sufficient for IMC targeting [#7]","Loss mislocalizes IMC4 in forming daughter buds [#7]","Expressed in tachyzoites and sporozoites but absent from gametocytes [#3]"]},"strain_conflicts":[]},"prefetch_data":{},"corpus_meta":[{"pmid":"18408052","id":"PMC_18408052","title":"A novel actin-related protein is associated with daughter cell formation in Toxoplasma gondii.","date":"2008","source":"Eukaryotic cell","url":"https://pubmed.ncbi.nlm.nih.gov/18408052","citation_count":19,"is_preprint":false},{"pmid":"11420112","id":"PMC_11420112","title":"Characterization of the subpellicular network, a filamentous membrane skeletal component in the parasite Toxoplasma gondii.","date":"2001","source":"Molecular and biochemical parasitology","url":"https://pubmed.ncbi.nlm.nih.gov/11420112","citation_count":200,"is_preprint":false},{"pmid":"11854415","id":"PMC_11854415","title":"Daughter cell assembly in the protozoan parasite Toxoplasma gondii.","date":"2002","source":"Molecular biology of the cell","url":"https://pubmed.ncbi.nlm.nih.gov/11854415","citation_count":151,"is_preprint":false},{"pmid":"12177058","id":"PMC_12177058","title":"Proteolytic processing of TgIMC1 during maturation of the membrane skeleton of Toxoplasma gondii.","date":"2002","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/12177058","citation_count":70,"is_preprint":false},{"pmid":"29062899","id":"PMC_29062899","title":"Differential Roles for Inner Membrane Complex Proteins across Toxoplasma gondii and Sarcocystis neurona Development.","date":"2017","source":"mSphere","url":"https://pubmed.ncbi.nlm.nih.gov/29062899","citation_count":50,"is_preprint":false},{"pmid":"24533291","id":"PMC_24533291","title":"Identification of differentially expressed proteins in sulfadiazine resistant and sensitive strains of Toxoplasma gondii using difference-gel electrophoresis (DIGE).","date":"2013","source":"International journal for parasitology. Drugs and drug resistance","url":"https://pubmed.ncbi.nlm.nih.gov/24533291","citation_count":42,"is_preprint":false},{"pmid":"18315523","id":"PMC_18315523","title":"Toxoplasma gondii Hsp20 is a stripe-arranged chaperone-like protein associated with the outer leaflet of the inner membrane complex.","date":"2008","source":"Biology of the cell","url":"https://pubmed.ncbi.nlm.nih.gov/18315523","citation_count":34,"is_preprint":false},{"pmid":"34384491","id":"PMC_34384491","title":"Requirement of Toxoplasma gondii metacaspases for IMC1 maturation, endodyogeny and virulence in mice.","date":"2021","source":"Parasites & vectors","url":"https://pubmed.ncbi.nlm.nih.gov/34384491","citation_count":5,"is_preprint":false},{"pmid":"40849645","id":"PMC_40849645","title":"Research on inner membrane complex protein 1: a novel nanovaccines against Toxoplasma gondii.","date":"2025","source":"BMC veterinary research","url":"https://pubmed.ncbi.nlm.nih.gov/40849645","citation_count":3,"is_preprint":false},{"pmid":"40768331","id":"PMC_40768331","title":"Function and interactions of a protein bridge between the inner membrane complex and subpellicular microtubules in Toxoplasma gondii.","date":"2025","source":"Molecular biology of the cell","url":"https://pubmed.ncbi.nlm.nih.gov/40768331","citation_count":2,"is_preprint":false},{"pmid":"40501855","id":"PMC_40501855","title":"Toxoplasma IMC1 is a central component of the subpellicular network and plays critical roles in parasite morphology, replication, and infectivity.","date":"2025","source":"bioRxiv : the preprint server for biology","url":"https://pubmed.ncbi.nlm.nih.gov/40501855","citation_count":0,"is_preprint":false},{"pmid":"42113865","id":"PMC_42113865","title":"Toxoplasma IMC1 is a central component of the subpellicular network and plays critical roles in parasite morphology, replication, and infectivity.","date":"2026","source":"PLoS pathogens","url":"https://pubmed.ncbi.nlm.nih.gov/42113865","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":2837,"output_tokens":601,"usd":0.008763,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":5327,"output_tokens":1586,"usd":0.033142,"stage2_stop_reason":"end_turn"},"total_usd":0.09193,"stage1_batch_id":"msgbatch_01WgHz6kceqm573AGLyz57at","stage2_batch_id":"msgbatch_01LK4wDYj5zYquahv7gi9893","note":"batch pricing = 50% of standard","skipped":true,"corrective_rerun":"2026-09-10 wrong-gene rerun (round-4 rules)","corrective_reason":"re-read with subject anchor"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2001,\n      \"finding\": \"TgIMC1 (ALV1) is a component of the subpellicular network, a filamentous membrane skeletal structure composed of interwoven 8–10 nm filaments associated with the cytoplasmic face of the pellicle of Toxoplasma gondii. TgIMC1 has predicted extended coiled-coil domains and is homologous to articulins, the major components of the membrane skeleton of algae and free-living protists.\",\n      \"method\": \"Electron microscopy of detergent-extracted tachyzoites; protein identification and sequence analysis\",\n      \"journal\": \"Molecular and biochemical parasitology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct EM visualization of filamentous network combined with protein identification; foundational study replicated by subsequent work\",\n      \"pmids\": [\"11420112\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"An IMC1–YFP fusion protein was used to visualize daughter scaffold (inner membrane complex) formation in live T. gondii by time-lapse video microscopy, demonstrating that the IMC1-based subpellicular network is assembled during endodyogeny and can also form during production of multiple daughters.\",\n      \"method\": \"Live-cell time-lapse fluorescence microscopy using IMC1–YFP fusion protein\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct live-imaging with functional reporter; foundational study widely replicated\",\n      \"pmids\": [\"11854415\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"TgIMC1 (ALV1) undergoes proteolytic processing of its carboxyl terminus during daughter parasite maturation. In immature parasites the network is detergent-soluble; in mature parasites it becomes entirely detergent-resistant. Conversion from detergent-labile to detergent-resistant network is coupled to proteolytic cleavage of the TgIMC1 C-terminus. A single cysteine residue in the C-terminus is essential for this processing event.\",\n      \"method\": \"Detergent fractionation assays; site-directed mutagenesis of TgIMC1 cysteine residue; immunoblotting to detect processing\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro mutagenesis combined with biochemical fractionation assay in a single focused study with rigorous controls\",\n      \"pmids\": [\"12177058\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"IMC1 (ALV1) is present in both mother and daughter cytoskeletons during asexual tachyzoite division, is absent from developing macro- and microgametocytes, and is present in sporozoites. IMC1 is thus present across multiple life cycle stages but absent from gametocytes, indicating a stage-specific role distinct from other IMC alveolins.\",\n      \"method\": \"Immunofluorescence assay (IFA) across multiple life-cycle stages of T. gondii and Sarcocystis neurona\",\n      \"journal\": \"mSphere\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct localization across multiple stages in a single lab study using IFA\",\n      \"pmids\": [\"29062899\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Double knockout of metacaspase-1 (MCA1) and metacaspase-2 (MCA2) in T. gondii results in detergent-soluble IMC1, indicating that MCA1 and MCA2 are required for IMC1 (ALV1) maturation (conversion to detergent-resistant form) and for normal endodyogeny.\",\n      \"method\": \"Double-knockout of MCA1/MCA2 in RHΔku80; detergent fractionation to assess IMC1 solubility; IFA to assess IMC1 expression and endodyogeny\",\n      \"journal\": \"Parasites & vectors\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic knockout with biochemical readout (detergent fractionation) linking metacaspases to IMC1 maturation, single lab\",\n      \"pmids\": [\"34384491\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"RH\"}]\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"IMT1, a protein that bridges the IMC and subpellicular microtubules, binds directly to the cytoskeletal IMC proteins IMC1 (ALV1), IMC18, and IMC24, as shown by deletion analyses and proximity labeling.\",\n      \"method\": \"Proximity labeling (BioID), co-immunoprecipitation/pulldown, deletion analyses\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct binding shown with proximity labeling and deletion analyses in a single focused study\",\n      \"pmids\": [\"40768331\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Disruption of IMC1 (ALV1) causes severe morphological defects, loss of cytoskeletal integrity, and defects in invasion, replication, and egress. In a type II strain, IMC1 loss causes dramatic reduction in infectivity and complete failure to establish chronic infection in vivo.\",\n      \"method\": \"Gene knockout; morphological analysis; invasion, replication, and egress assays; mouse infection studies\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — clean gene knockout with multiple orthogonal functional readouts (morphology, invasion, replication, egress, in vivo infection) replicated across strains and published in peer-reviewed journal\",\n      \"pmids\": [\"42113865\", \"40501855\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"RH\"}, {\"species\": \"Toxoplasma gondii\", \"strain\": \"ME49\"}]\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"IMC1 (ALV1) is loaded onto forming daughter buds together with IMC4, but later than IMC3, IMC6, and IMC10. A key subregion of the alveolin domain is sufficient for IMC targeting and is also required for IMC1 function. IMC1 interacts directly with IMC4, and loss of IMC1 results in mislocalization of IMC4 specifically in forming daughter buds.\",\n      \"method\": \"Fluorescent protein tagging with time-resolved IFA; deletion analysis; mutagenesis; co-immunoprecipitation/pulldown to establish direct IMC1–IMC4 interaction; IFA of IMC4 localization in IMC1 knockout\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — direct interaction confirmed by pulldown, deletion/mutagenesis analysis for functional domain mapping, and IFA for localization consequence; peer-reviewed publication\",\n      \"pmids\": [\"42113865\", \"40501855\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"RH\"}, {\"species\": \"Toxoplasma gondii\", \"strain\": \"ME49\"}]\n    }\n  ],\n  \"current_model\": \"ALV1/IMC1 is the founding alveolin and a central structural component of the Toxoplasma gondii subpellicular network, forming 8–10 nm filaments on the cytoplasmic face of the IMC; it is loaded onto daughter buds during endodyogeny (with IMC4, but after IMC3/IMC6/IMC10), undergoes metacaspase-dependent proteolytic processing of its cysteine-containing C-terminus to transition from a detergent-labile to a detergent-resistant mature network, directly binds IMC4 (whose localization in daughter buds depends on IMC1) and the IMC-microtubule bridging protein IMT1, and is essential for cytoskeletal integrity, invasion, replication, egress, and establishment of chronic infection in vivo.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"ALV1 (TgIMC1) is the founding alveolin and a central structural component of the subpellicular network, a filamentous membrane skeleton of interwoven 8\\u201310 nm filaments associated with the cytoplasmic face of the Toxoplasma gondii pellicle [#0]. It is assembled onto daughter cytoskeletons during endodyogeny and multiple-daughter formation [#1], being loaded onto forming daughter buds together with IMC4 but later than IMC3, IMC6, and IMC10; a subregion of its alveolin domain is sufficient for IMC targeting and required for its function, and it binds IMC4 directly, with IMC1 loss mislocalizing IMC4 in daughter buds [#7]. Network maturation depends on proteolytic processing of the ALV1 C-terminus\\u2014requiring a single C-terminal cysteine\\u2014that converts the detergent-labile immature network into a detergent-resistant mature form [#2], a maturation step that requires the metacaspases MCA1 and MCA2 in RH [#4]. ALV1 also binds directly to IMT1, the protein bridging the IMC to subpellicular microtubules [#5]. ALV1 is expressed across tachyzoite, sporozoite, and other stages but absent from gametocytes [#3], and its disruption causes loss of cytoskeletal integrity with defects in invasion, replication, and egress, abolishing chronic infection in a type II (ME49) strain in vivo [#6]. Beyond these findings, nothing further about ALV1 is established in the available corpus.\"\n  ,\n  \"teleology\": [\n    {\n      \"year\": 2001,\n      \"claim\": \"Establishing that ALV1/TgIMC1 is a genuine structural component of the parasite membrane skeleton answered whether the pellicle-associated filamentous network had defined protein constituents.\",\n      \"evidence\": \"Electron microscopy of detergent-extracted tachyzoites with protein identification and sequence analysis\",\n      \"pmids\": [\"11420112\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Filament assembly mechanism not resolved\", \"Partner proteins within the network not yet identified\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Live imaging of an IMC1\\u2013YFP reporter answered when and how the network is built, showing it is assembled de novo during endodyogeny and multiple-daughter formation.\",\n      \"evidence\": \"Live-cell time-lapse fluorescence microscopy of an IMC1\\u2013YFP fusion in T. gondii\",\n      \"pmids\": [\"11854415\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Targeting determinants for daughter bud loading not defined\", \"Order relative to other IMC proteins unknown\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Identifying C-terminal proteolytic processing coupled to a detergent-labile-to-resistant transition answered how the immature network matures into a stable skeleton.\",\n      \"evidence\": \"Detergent fractionation plus site-directed mutagenesis of a C-terminal cysteine and immunoblotting\",\n      \"pmids\": [\"12177058\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Protease responsible not identified in this study\", \"Molecular basis of detergent resistance unresolved\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Mapping IMC1 expression across the life cycle answered whether its role is stage-specific, showing presence in tachyzoites and sporozoites but absence from gametocytes.\",\n      \"evidence\": \"Immunofluorescence across multiple life-cycle stages\",\n      \"pmids\": [\"29062899\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Functional consequence of gametocyte absence not tested\", \"Stage-specific regulation mechanism unknown\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Genetic loss of MCA1/MCA2 identified the enzymes required for IMC1 maturation, linking metacaspase activity to conversion of the network to its detergent-resistant form.\",\n      \"evidence\": \"MCA1/MCA2 double knockout in RH\\u0394ku80 with detergent fractionation and IFA\",\n      \"pmids\": [\"34384491\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct cleavage of IMC1 by metacaspases not demonstrated\", \"Whether processing is direct or indirect not resolved\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"RH\"}]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Direct binding of IMT1 to IMC1 answered how the alveolin network physically couples to subpellicular microtubules.\",\n      \"evidence\": \"Proximity labeling (BioID), co-immunoprecipitation/pulldown, and deletion analyses\",\n      \"pmids\": [\"40768331\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Binding interface on IMC1 not mapped\", \"Functional consequence of the IMC1\\u2013IMT1 interaction for microtubule anchoring not tested\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Knockout combined with domain mapping and interaction analysis established ALV1 as essential for cytoskeletal integrity and defined its daughter-bud loading order and its direct role in positioning IMC4.\",\n      \"evidence\": \"Gene knockout with morphology, invasion, replication, egress and mouse infection assays; fluorescent tagging with time-resolved IFA, deletion/mutagenesis, and IMC1\\u2013IMC4 pulldown\",\n      \"pmids\": [\"42113865\", \"40501855\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structure of the alveolin targeting subregion not determined\", \"How IMC1 nucleates or organizes the broader network hierarchy not resolved\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"RH\"}, {\"species\": \"Toxoplasma gondii\", \"strain\": \"ME49\"}]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The precise molecular mechanism by which ALV1 filaments assemble, mature into a detergent-resistant meshwork, and organize their partner alveolins into the intact subpellicular network remains open.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No structural model of the alveolin filament or its assembly\", \"Direct protease-substrate relationship for C-terminal processing not established\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [0, 6], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]},\n      {\"term_id\": \"GO:0005200\", \"supporting_discovery_ids\": [0], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]},\n      {\"term_id\": \"GO:0005515\", \"supporting_discovery_ids\": [5, 7], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0033113\", \"supporting_discovery_ids\": [0, 3], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]},\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [0, 6], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"GO:0061351\", \"supporting_discovery_ids\": [1, 7], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]},\n      {\"term_id\": \"GO:0044409\", \"supporting_discovery_ids\": [6], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"ME49\"}]}\n    ],\n    \"complexes\": [\"Subpellicular network / inner membrane complex alveolin meshwork\"],\n    \"partners\": [\"IMC4 (direct binding)\", \"IMT1 (direct binding, IMC-microtubule bridge)\", \"MCA1/MCA2 metacaspases (required for maturation)\"],\n    \"other_free_text\": [\n      \"Founding alveolin; predicted extended coiled-coil domains; homologous to articulins [#0]\",\n      \"C-terminal proteolytic processing requiring a single cysteine converts the detergent-labile network to detergent-resistant [#2]\",\n      \"Loaded onto daughter buds with IMC4 but after IMC3/IMC6/IMC10; alveolin subregion sufficient for IMC targeting [#7]\",\n      \"Loss mislocalizes IMC4 in forming daughter buds [#7]\",\n      \"Expressed in tachyzoites and sporozoites but absent from gametocytes [#3]\"\n    ]\n  },\n  \"strain_conflicts\": []\n}\n```","audit_flag":null,"strains":[{"strain":"ME49","gene_id":"TGME49_231640","name":"ALV1","product":"alveolin domain containing intermediate filament IMC1","uniprot_acc":"A0A125YFV7","chromosome":"TGME49_chrVIII","start_pos":1516113},{"strain":"GT1","gene_id":"TGGT1_231640","name":null,"product":"alveolin domain containing intermediate filament IMC1","uniprot_acc":null,"chromosome":"TGGT1_chrVIII","start_pos":1511977},{"strain":"RH88","gene_id":"TGRH88_075820","name":null,"product":"alveolin domain containing intermediate filament IMC1","uniprot_acc":null,"chromosome":"CM023089","start_pos":6622752},{"strain":"VEG","gene_id":"TGVEG_231640","name":null,"product":"alveolin domain containing intermediate filament IMC1","uniprot_acc":"A0A0F7V263","chromosome":"TGVEG_chrVIII","start_pos":1522790}]}