{"gene":"TG_249970","gene_symbol":"APH","run_date":"2026-09-08T12:58:25","timeline":{"discoveries":[{"year":2016,"finding":"APH (acylated pleckstrin homology domain-containing protein) localizes to the microneme surface and senses phosphatidic acid (PA) during microneme secretion; APH is necessary for microneme exocytosis, as its depletion impairs secretion.","method":"Genetic depletion (loss-of-function), localization experiments, lipid-sensing assays","journal":"Cell host & microbe","confidence":"High","confidence_rationale":"Tier 2 / Moderate — direct localization and functional knockout with defined cellular phenotype (microneme exocytosis defect), single lab but multiple orthogonal approaches","pmids":["26962945"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2016,"finding":"APH functions downstream of DAG-kinase-1 in the PA/DAG signaling balance: DAG-kinase-1 converts DAG to PA, and APH on the microneme surface senses the resulting PA to trigger exocytosis.","method":"Genetic epistasis (DAG-kinase-1 depletion combined with APH functional analysis), lipid second-messenger signaling assays","journal":"Cell host & microbe","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — pathway placement by genetic epistasis in single lab study; pathway position inferred from combinatorial depletion results","pmids":["26962945"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2018,"finding":"Crystal/structural analysis of TgAPH revealed the atomic basis of PA recognition: the PH domain inserts into PA-enriched membranes and clusters multiple phosphate head-groups at its bilayer-binding surface, defining the key determinants of PA sensing.","method":"Structural determination (atomic-resolution structure), phospholipid-membrane interaction assays, mutagenesis of PA-binding residues","journal":"Structure (London, England : 1993)","confidence":"High","confidence_rationale":"Tier 1 / Moderate — atomic structure with functional validation of PA-binding mechanism in a single rigorous study using structural and membrane-interaction methods","pmids":["29910186"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""},{"species":"Plasmodium falciparum","strain":""}]},{"year":2018,"finding":"TgAPH is acylated on the microneme surface and is recruited to PA-enriched membranes, with acylation contributing to its membrane association.","method":"Biochemical fractionation, localization experiments, lipid-binding assays","journal":"Structure (London, England : 1993)","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — direct localization and acylation characterization in a single lab study; abstract does not detail full mutagenesis of acylation sites","pmids":["29910186"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]}],"current_model":"TgAPH is an acylated PH domain-containing protein resident on the microneme surface that senses phosphatidic acid (PA) — generated by DAG-kinase-1 from DAG downstream of PLC activation — by inserting its PH domain into PA-enriched membranes and clustering phosphate head-groups, thereby triggering microneme exocytosis essential for parasite motility, invasion, and egress."},"narrative":{"mechanistic_narrative":"APH (acylated pleckstrin homology domain-containing protein) is a lipid-sensing effector required for regulated microneme exocytosis in apicomplexan parasites, a process essential for motility, invasion, and egress [PMID:26962945]. Resident on the microneme surface, APH senses phosphatidic acid (PA) generated by DAG-kinase-1 from diacylglycerol, placing APH downstream of this lipid second-messenger conversion so that the DAG/PA balance controls the timing of secretion [PMID:26962945]. Structural analysis defined the molecular basis of PA recognition: the PH domain inserts into PA-enriched bilayers and clusters multiple phosphate head-groups at its membrane-binding surface [PMID:29910186], while acylation contributes to APH membrane association and its recruitment to PA-enriched membranes [PMID:29910186].","teleology":[{"year":2016,"claim":"Established that APH is a microneme-surface lipid sensor whose PA-sensing function is necessary for regulated secretion, defining a molecular link between membrane lipid composition and exocytosis.","evidence":"Genetic depletion with localization and lipid-sensing assays in Toxoplasma gondii","pmids":["26962945"],"confidence":"High","gaps":["Does not resolve how PA sensing is mechanically coupled to the exocytic machinery","Does not identify downstream effectors of APH"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2016,"claim":"Placed APH downstream of DAG-kinase-1 in the DAG/PA signaling balance, showing that PA generated by DAG-kinase-1 is the ligand APH senses to trigger exocytosis.","evidence":"Genetic epistasis combining DAG-kinase-1 depletion with APH functional analysis in Toxoplasma gondii","pmids":["26962945"],"confidence":"Medium","gaps":["Pathway position inferred from combinatorial depletion rather than direct biochemical reconstitution","Upstream PLC activation and DAG dynamics not directly linked to APH in this evidence"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2018,"claim":"Resolved the atomic basis of PA recognition, showing the PH domain inserts into PA-enriched membranes and clusters phosphate head-groups, defining the structural determinants of lipid sensing.","evidence":"Atomic-resolution structure with phospholipid-membrane interaction assays and mutagenesis of PA-binding residues in Toxoplasma gondii and Plasmodium falciparum","pmids":["29910186"],"confidence":"High","gaps":["Does not establish how PA binding is transmitted to trigger vesicle fusion","In vivo consequences of specific PA-binding residue mutations on secretion not fully mapped"],"attribution":[{"species":"Toxoplasma gondii","strain":""},{"species":"Plasmodium falciparum","strain":""}]},{"year":2018,"claim":"Showed that acylation contributes to APH membrane association, adding a second membrane-targeting determinant alongside PA head-group binding.","evidence":"Biochemical fractionation, localization, and lipid-binding assays in Toxoplasma gondii","pmids":["29910186"],"confidence":"Medium","gaps":["Acylation sites not fully mutagenized to establish causality","Relative contribution of acylation versus PA binding to membrane targeting not quantified"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":null,"claim":"How PA sensing by APH is mechanically coupled to the microneme fusion machinery to drive exocytosis remains unresolved.","evidence":"","pmids":[],"confidence":"Low","gaps":["No identified downstream effector connecting APH to vesicle fusion","No reconstitution of PA-triggered secretion","Regulation of APH acylation state during signaling unknown"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]}],"mechanism_profile":{"molecular_activity":[],"localization":[],"pathway":[],"complexes":[],"partners":["DAG-KINASE-1 (FUNCTIONAL UPSTREAM, GENERATES PA LIGAND)"],"other_free_text":["APH is acylated on the microneme surface, a modification contributing to membrane association [#3].","PA recognition occurs by PH-domain insertion into PA-enriched membranes and clustering of phosphate head-groups [#2].","PA-sensing function shown in Toxoplasma gondii and Plasmodium falciparum [#2]."]},"strain_conflicts":[]},"prefetch_data":{},"corpus_meta":[{"pmid":"26962945","id":"PMC_26962945","title":"Phosphatidic Acid-Mediated Signaling Regulates Microneme Secretion in Toxoplasma.","date":"2016","source":"Cell host & microbe","url":"https://pubmed.ncbi.nlm.nih.gov/26962945","citation_count":121,"is_preprint":false},{"pmid":"29910186","id":"PMC_29910186","title":"Structural Basis of Phosphatidic Acid Sensing by APH in Apicomplexan Parasites.","date":"2018","source":"Structure (London, England : 1993)","url":"https://pubmed.ncbi.nlm.nih.gov/29910186","citation_count":17,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":3227,"output_tokens":1039,"usd":0.012633,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":5930,"output_tokens":2055,"usd":0.040512,"stage2_stop_reason":"end_turn"},"total_usd":0.053145,"stage1_batch_id":"msgbatch_01TEPNZ4MCzJ7xtsNwosiDRk","stage2_batch_id":"msgbatch_01FQShM7n1sfCr4pWTDShmki","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2016,\n      \"finding\": \"APH (acylated pleckstrin homology domain-containing protein) localizes to the microneme surface and senses phosphatidic acid (PA) during microneme secretion; APH is necessary for microneme exocytosis, as its depletion impairs secretion.\",\n      \"method\": \"Genetic depletion (loss-of-function), localization experiments, lipid-sensing assays\",\n      \"journal\": \"Cell host & microbe\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct localization and functional knockout with defined cellular phenotype (microneme exocytosis defect), single lab but multiple orthogonal approaches\",\n      \"pmids\": [\"26962945\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"APH functions downstream of DAG-kinase-1 in the PA/DAG signaling balance: DAG-kinase-1 converts DAG to PA, and APH on the microneme surface senses the resulting PA to trigger exocytosis.\",\n      \"method\": \"Genetic epistasis (DAG-kinase-1 depletion combined with APH functional analysis), lipid second-messenger signaling assays\",\n      \"journal\": \"Cell host & microbe\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — pathway placement by genetic epistasis in single lab study; pathway position inferred from combinatorial depletion results\",\n      \"pmids\": [\"26962945\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Crystal/structural analysis of TgAPH revealed the atomic basis of PA recognition: the PH domain inserts into PA-enriched membranes and clusters multiple phosphate head-groups at its bilayer-binding surface, defining the key determinants of PA sensing.\",\n      \"method\": \"Structural determination (atomic-resolution structure), phospholipid-membrane interaction assays, mutagenesis of PA-binding residues\",\n      \"journal\": \"Structure (London, England : 1993)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — atomic structure with functional validation of PA-binding mechanism in a single rigorous study using structural and membrane-interaction methods\",\n      \"pmids\": [\"29910186\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Toxoplasma gondii\", \"strain\": \"\"},\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"TgAPH is acylated on the microneme surface and is recruited to PA-enriched membranes, with acylation contributing to its membrane association.\",\n      \"method\": \"Biochemical fractionation, localization experiments, lipid-binding assays\",\n      \"journal\": \"Structure (London, England : 1993)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — direct localization and acylation characterization in a single lab study; abstract does not detail full mutagenesis of acylation sites\",\n      \"pmids\": [\"29910186\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}\n      ]\n    }\n  ],\n  \"current_model\": \"TgAPH is an acylated PH domain-containing protein resident on the microneme surface that senses phosphatidic acid (PA) — generated by DAG-kinase-1 from DAG downstream of PLC activation — by inserting its PH domain into PA-enriched membranes and clustering phosphate head-groups, thereby triggering microneme exocytosis essential for parasite motility, invasion, and egress.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"APH (acylated pleckstrin homology domain-containing protein) is a lipid-sensing effector required for regulated microneme exocytosis in apicomplexan parasites, a process essential for motility, invasion, and egress [#0]. Resident on the microneme surface, APH senses phosphatidic acid (PA) generated by DAG-kinase-1 from diacylglycerol, placing APH downstream of this lipid second-messenger conversion so that the DAG/PA balance controls the timing of secretion [#1]. Structural analysis defined the molecular basis of PA recognition: the PH domain inserts into PA-enriched bilayers and clusters multiple phosphate head-groups at its membrane-binding surface [#2], while acylation contributes to APH membrane association and its recruitment to PA-enriched membranes [#3].\",\n  \"teleology\": [\n    {\n      \"year\": 2016,\n      \"claim\": \"Established that APH is a microneme-surface lipid sensor whose PA-sensing function is necessary for regulated secretion, defining a molecular link between membrane lipid composition and exocytosis.\",\n      \"evidence\": \"Genetic depletion with localization and lipid-sensing assays in Toxoplasma gondii\",\n      \"pmids\": [\"26962945\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Does not resolve how PA sensing is mechanically coupled to the exocytic machinery\", \"Does not identify downstream effectors of APH\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Placed APH downstream of DAG-kinase-1 in the DAG/PA signaling balance, showing that PA generated by DAG-kinase-1 is the ligand APH senses to trigger exocytosis.\",\n      \"evidence\": \"Genetic epistasis combining DAG-kinase-1 depletion with APH functional analysis in Toxoplasma gondii\",\n      \"pmids\": [\"26962945\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Pathway position inferred from combinatorial depletion rather than direct biochemical reconstitution\", \"Upstream PLC activation and DAG dynamics not directly linked to APH in this evidence\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Resolved the atomic basis of PA recognition, showing the PH domain inserts into PA-enriched membranes and clusters phosphate head-groups, defining the structural determinants of lipid sensing.\",\n      \"evidence\": \"Atomic-resolution structure with phospholipid-membrane interaction assays and mutagenesis of PA-binding residues in Toxoplasma gondii and Plasmodium falciparum\",\n      \"pmids\": [\"29910186\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Does not establish how PA binding is transmitted to trigger vesicle fusion\", \"In vivo consequences of specific PA-binding residue mutations on secretion not fully mapped\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}, {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Showed that acylation contributes to APH membrane association, adding a second membrane-targeting determinant alongside PA head-group binding.\",\n      \"evidence\": \"Biochemical fractionation, localization, and lipid-binding assays in Toxoplasma gondii\",\n      \"pmids\": [\"29910186\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Acylation sites not fully mutagenized to establish causality\", \"Relative contribution of acylation versus PA binding to membrane targeting not quantified\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How PA sensing by APH is mechanically coupled to the microneme fusion machinery to drive exocytosis remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No identified downstream effector connecting APH to vesicle fusion\", \"No reconstitution of PA-triggered secretion\", \"Regulation of APH acylation state during signaling unknown\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0070300\", \"supporting_discovery_ids\": [0, 1, 2], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]},\n      {\"term_id\": \"GO:0008289\", \"supporting_discovery_ids\": [2, 3], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0020009\", \"supporting_discovery_ids\": [0, 3], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"GO:0017157\", \"supporting_discovery_ids\": [0, 1], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"DAG-kinase-1 (functional upstream, generates PA ligand)\"],\n    \"other_free_text\": [\n      \"APH is acylated on the microneme surface, a modification contributing to membrane association [#3].\",\n      \"PA recognition occurs by PH-domain insertion into PA-enriched membranes and clustering of phosphate head-groups [#2].\",\n      \"PA-sensing function shown in Toxoplasma gondii and Plasmodium falciparum [#2].\"\n    ]\n  },\n  \"strain_conflicts\": []\n}","audit_flag":null,"strains":[{"strain":"ME49","gene_id":"TGME49_249970","name":"APH","product":"acylated pleckstrin-homology domain-containing protein APH","uniprot_acc":"A0A125YY72","chromosome":"TGME49_chrXII","start_pos":4963719},{"strain":"GT1","gene_id":"TGGT1_249970","name":null,"product":"hypothetical protein","uniprot_acc":null,"chromosome":"TGGT1_chrXII","start_pos":4900104},{"strain":"RH88","gene_id":"TGRH88_063720","name":null,"product":"unspecified product","uniprot_acc":null,"chromosome":"CM023093","start_pos":5022070},{"strain":"VEG","gene_id":"TGVEG_249970","name":null,"product":"hypothetical protein","uniprot_acc":"B6KHK6","chromosome":"TGVEG_chrXII","start_pos":4939290}]}