{"gene":"TG_226040","gene_symbol":"CAM3","run_date":"2026-09-08T11:06:44","timeline":{"discoveries":[{"year":2017,"finding":"CaM3 is essential for Toxoplasma gondii growth; auxin-induced degradation of CaM3 blocked parasite growth, and CaM3 was refractory to deletion by CRISPR/Cas9.","method":"CRISPR/Cas9-mediated gene editing and auxin-induced degron (AID) system with growth assays","journal":"PLoS pathogens","confidence":"High","confidence_rationale":"Tier 2 / Strong — two orthogonal loss-of-function approaches (CRISPR deletion and AID-mediated degradation) with clear growth phenotype in a single rigorous study","pmids":["28475612"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2017,"finding":"CaM3 localizes to the conoid at the apical tip of Toxoplasma gondii, overlapping with myosin H (MyoH).","method":"Super-resolution microscopy","journal":"PLoS pathogens","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — direct localization by super-resolution microscopy, single lab, co-localization only without functional rescue","pmids":["28475612"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2017,"finding":"CaM3 contributes to parasite motility, invasion, and egress from host cells, acting downstream of microneme and rhoptry secretion.","method":"AID-mediated conditional degradation with motility, invasion, and egress assays; secretion assays","journal":"PLoS pathogens","confidence":"High","confidence_rationale":"Tier 2 / Moderate — conditional knockdown with multiple orthogonal phenotypic readouts (motility, invasion, egress, secretion) in a single rigorous study","pmids":["28475612"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2017,"finding":"BirA-based proximity biotinylation with CaM3 labeled apical proteins including MyoH and its light chain MLC7, suggesting a physical interaction; disruption of MyoH led to degradation of CaM3, placing CaM3 functionally downstream of or in complex with MyoH.","method":"BirA proximity biotinylation and MyoH disruption with CaM3 stability assay","journal":"PLoS pathogens","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — proximity labeling (not reciprocal Co-IP) plus genetic epistasis showing CaM3 stability depends on MyoH, single lab","pmids":["28475612"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]}],"current_model":"CaM3 is an essential calmodulin-like protein that localizes to the Toxoplasma gondii conoid, where it acts downstream of microneme and rhoptry secretion to regulate parasite motility, invasion, and egress, likely by interacting with the conoid-associated motor myosin H (MyoH), as evidenced by proximity biotinylation and the observation that MyoH disruption causes CaM3 degradation."},"narrative":{"mechanistic_narrative":"CaM3 is an essential calmodulin-like protein of Toxoplasma gondii that operates at the conoid to drive the apical motility machinery underlying parasite invasion and egress [PMID:28475612]. It localizes to the conoid at the parasite's apical tip, overlapping with the conoid-associated motor myosin H (MyoH) [PMID:28475612]. Functionally, CaM3 acts downstream of microneme and rhoptry secretion to support gliding motility, host-cell invasion, and egress, since conditional depletion blocks these processes without impairing secretion [PMID:28475612]. Proximity biotinylation places CaM3 in the apical MyoH complex, labeling MyoH and its light chain MLC7, and disruption of MyoH triggers CaM3 degradation, indicating that CaM3 stability and function are coupled to the MyoH motor [PMID:28475612]. Beyond this conoid-localized, MyoH-associated role, no further biochemical or structural mechanism has been characterized in the available corpus.","teleology":[{"year":2017,"claim":"Establishing whether CaM3 is dispensable answered whether this calmodulin-like protein has a non-redundant role in the parasite lytic cycle.","evidence":"CRISPR/Cas9 deletion (refractory) and auxin-induced degron depletion with growth assays in T. gondii","pmids":["28475612"],"confidence":"High","gaps":["Does not identify which downstream molecular activity of CaM3 is essential","No structural or biochemical characterization of the protein"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2017,"claim":"Defining CaM3's subcellular position answered where in the parasite it acts, pinpointing it to the apical motility apparatus.","evidence":"Super-resolution microscopy showing conoid localization overlapping MyoH in T. gondii","pmids":["28475612"],"confidence":"Medium","gaps":["Co-localization only, no functional rescue at the conoid","Does not establish direct physical association from imaging alone"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2017,"claim":"Dissecting the step at which CaM3 acts answered whether it functions in secretion or in the downstream motor-driven events of the lytic cycle.","evidence":"AID conditional depletion with motility, invasion, egress, and secretion assays in T. gondii","pmids":["28475612"],"confidence":"High","gaps":["Does not resolve the molecular target CaM3 regulates during motility","Does not distinguish direct motor regulation from indirect effects"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2017,"claim":"Identifying CaM3's molecular neighbors answered how it is integrated into the motility machinery, linking it to the MyoH motor complex.","evidence":"BirA proximity biotinylation identifying MyoH and MLC7, plus MyoH disruption assessing CaM3 stability in T. gondii","pmids":["28475612"],"confidence":"Medium","gaps":["Proximity labeling, not reciprocal Co-IP, so direct binding is unproven","Mechanism by which MyoH loss destabilizes CaM3 is undefined","No demonstration that CaM3 modulates MyoH motor activity"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":null,"claim":"How CaM3 mechanistically regulates the MyoH motor and what Ca2+-dependent signals it transduces at the conoid remain unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No direct binding assay or structure of a CaM3-MyoH/MLC7 complex","No test of whether CaM3 confers Ca2+ sensitivity to motor function","No reciprocal Co-IP validating the interaction"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]}],"mechanism_profile":{"molecular_activity":[],"localization":[],"pathway":[],"complexes":["Apical MyoH motor complex (MyoH, MLC7, CaM3)"],"partners":["MYOH (MYOSIN H)","MLC7 (MYOSIN LIGHT CHAIN 7)"],"other_free_text":["CaM3 is essential; refractory to CRISPR deletion and its depletion blocks growth [#0].","CaM3 acts downstream of microneme and rhoptry secretion [#2].","CaM3 protein stability depends on MyoH; MyoH disruption causes CaM3 degradation [#3]."]},"strain_conflicts":[]},"prefetch_data":{},"corpus_meta":[{"pmid":"28475612","id":"PMC_28475612","title":"Calmodulin-like proteins localized to the conoid regulate motility and cell invasion by Toxoplasma gondii.","date":"2017","source":"PLoS pathogens","url":"https://pubmed.ncbi.nlm.nih.gov/28475612","citation_count":84,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":2883,"output_tokens":983,"usd":0.011697,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":5829,"output_tokens":1925,"usd":0.038635,"stage2_stop_reason":"end_turn"},"total_usd":0.050332,"stage1_batch_id":"msgbatch_01WokvBYTB6JLnYrpD4rXnyV","stage2_batch_id":"msgbatch_01M7hsQDvz6AqVC9DLVjQzzc","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2017,\n      \"finding\": \"CaM3 is essential for Toxoplasma gondii growth; auxin-induced degradation of CaM3 blocked parasite growth, and CaM3 was refractory to deletion by CRISPR/Cas9.\",\n      \"method\": \"CRISPR/Cas9-mediated gene editing and auxin-induced degron (AID) system with growth assays\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — two orthogonal loss-of-function approaches (CRISPR deletion and AID-mediated degradation) with clear growth phenotype in a single rigorous study\",\n      \"pmids\": [\"28475612\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"CaM3 localizes to the conoid at the apical tip of Toxoplasma gondii, overlapping with myosin H (MyoH).\",\n      \"method\": \"Super-resolution microscopy\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — direct localization by super-resolution microscopy, single lab, co-localization only without functional rescue\",\n      \"pmids\": [\"28475612\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"CaM3 contributes to parasite motility, invasion, and egress from host cells, acting downstream of microneme and rhoptry secretion.\",\n      \"method\": \"AID-mediated conditional degradation with motility, invasion, and egress assays; secretion assays\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — conditional knockdown with multiple orthogonal phenotypic readouts (motility, invasion, egress, secretion) in a single rigorous study\",\n      \"pmids\": [\"28475612\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"BirA-based proximity biotinylation with CaM3 labeled apical proteins including MyoH and its light chain MLC7, suggesting a physical interaction; disruption of MyoH led to degradation of CaM3, placing CaM3 functionally downstream of or in complex with MyoH.\",\n      \"method\": \"BirA proximity biotinylation and MyoH disruption with CaM3 stability assay\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — proximity labeling (not reciprocal Co-IP) plus genetic epistasis showing CaM3 stability depends on MyoH, single lab\",\n      \"pmids\": [\"28475612\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    }\n  ],\n  \"current_model\": \"CaM3 is an essential calmodulin-like protein that localizes to the Toxoplasma gondii conoid, where it acts downstream of microneme and rhoptry secretion to regulate parasite motility, invasion, and egress, likely by interacting with the conoid-associated motor myosin H (MyoH), as evidenced by proximity biotinylation and the observation that MyoH disruption causes CaM3 degradation.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"CaM3 is an essential calmodulin-like protein of Toxoplasma gondii that operates at the conoid to drive the apical motility machinery underlying parasite invasion and egress [#0, #2]. It localizes to the conoid at the parasite's apical tip, overlapping with the conoid-associated motor myosin H (MyoH) [#1]. Functionally, CaM3 acts downstream of microneme and rhoptry secretion to support gliding motility, host-cell invasion, and egress, since conditional depletion blocks these processes without impairing secretion [#2]. Proximity biotinylation places CaM3 in the apical MyoH complex, labeling MyoH and its light chain MLC7, and disruption of MyoH triggers CaM3 degradation, indicating that CaM3 stability and function are coupled to the MyoH motor [#3]. Beyond this conoid-localized, MyoH-associated role, no further biochemical or structural mechanism has been characterized in the available corpus.\",\n  \"teleology\": [\n    {\n      \"year\": 2017,\n      \"claim\": \"Establishing whether CaM3 is dispensable answered whether this calmodulin-like protein has a non-redundant role in the parasite lytic cycle.\",\n      \"evidence\": \"CRISPR/Cas9 deletion (refractory) and auxin-induced degron depletion with growth assays in T. gondii\",\n      \"pmids\": [\"28475612\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Does not identify which downstream molecular activity of CaM3 is essential\", \"No structural or biochemical characterization of the protein\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Defining CaM3's subcellular position answered where in the parasite it acts, pinpointing it to the apical motility apparatus.\",\n      \"evidence\": \"Super-resolution microscopy showing conoid localization overlapping MyoH in T. gondii\",\n      \"pmids\": [\"28475612\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Co-localization only, no functional rescue at the conoid\", \"Does not establish direct physical association from imaging alone\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Dissecting the step at which CaM3 acts answered whether it functions in secretion or in the downstream motor-driven events of the lytic cycle.\",\n      \"evidence\": \"AID conditional depletion with motility, invasion, egress, and secretion assays in T. gondii\",\n      \"pmids\": [\"28475612\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Does not resolve the molecular target CaM3 regulates during motility\", \"Does not distinguish direct motor regulation from indirect effects\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Identifying CaM3's molecular neighbors answered how it is integrated into the motility machinery, linking it to the MyoH motor complex.\",\n      \"evidence\": \"BirA proximity biotinylation identifying MyoH and MLC7, plus MyoH disruption assessing CaM3 stability in T. gondii\",\n      \"pmids\": [\"28475612\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Proximity labeling, not reciprocal Co-IP, so direct binding is unproven\", \"Mechanism by which MyoH loss destabilizes CaM3 is undefined\", \"No demonstration that CaM3 modulates MyoH motor activity\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How CaM3 mechanistically regulates the MyoH motor and what Ca2+-dependent signals it transduces at the conoid remain unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No direct binding assay or structure of a CaM3-MyoH/MLC7 complex\", \"No test of whether CaM3 confers Ca2+ sensitivity to motor function\", \"No reciprocal Co-IP validating the interaction\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [],\n    \"localization\": [\n      {\"term_id\": \"conoid (apical tip)\", \"supporting_discovery_ids\": [1], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"parasite motility, host-cell invasion, and egress\", \"supporting_discovery_ids\": [2], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]}\n    ],\n    \"complexes\": [\"Apical MyoH motor complex (MyoH, MLC7, CaM3)\"],\n    \"partners\": [\"MyoH (myosin H)\", \"MLC7 (myosin light chain 7)\"],\n    \"other_free_text\": [\n      \"CaM3 is essential; refractory to CRISPR deletion and its depletion blocks growth [#0].\",\n      \"CaM3 acts downstream of microneme and rhoptry secretion [#2].\",\n      \"CaM3 protein stability depends on MyoH; MyoH disruption causes CaM3 degradation [#3].\"\n    ]\n  },\n  \"strain_conflicts\": []\n}","audit_flag":null,"strains":[{"strain":"ME49","gene_id":"TGME49_226040","name":"CAM3","product":"calmodulin CAM3","uniprot_acc":"A0A125YWI5","chromosome":"TGME49_chrX","start_pos":1850471},{"strain":"GT1","gene_id":"TGGT1_226040","name":null,"product":"EF hand domain-containing protein","uniprot_acc":null,"chromosome":"TGGT1_chrX","start_pos":1759538},{"strain":"RH88","gene_id":"TGRH88_045710","name":null,"product":"EF hand domain-containing protein","uniprot_acc":null,"chromosome":"CM023091","start_pos":5718833},{"strain":"VEG","gene_id":"TGVEG_226040","name":null,"product":"EF hand domain-containing protein","uniprot_acc":"B9PML0","chromosome":"TGVEG_chrX","start_pos":1751288}]}