{"gene":"TG_254000","gene_symbol":"GRA47","run_date":"2026-09-08T12:58:25","timeline":{"discoveries":[{"year":2024,"finding":"GRA47 physically interacts with GRA72, as demonstrated by immunoprecipitation experiments; GRA47 localizes to the parasitophorous vacuole membrane (PVM) and is required for normal PV morphology and PVM permeability to small molecules.","method":"Co-immunoprecipitation, genetic deletion (Δgra47), small molecule permeability assays, live imaging of vacuole morphology","journal":"mBio","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP plus genetic knockout with defined cellular phenotype, independently replicated in two separate publications (PMID:38380952 and PMID:38936501)","pmids":["38380952","38936501"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2024,"finding":"GRA47 is predicted by structural modelling to form a heptameric pore, with conserved histidine residues lining the pore channel; mutational analysis of these histidines abolished protein functionality (aberrant PV morphology, reduced permeability), and electrophysiology confirmed that loss of GRA47 alters membrane conductance, corroborating its pore-forming role.","method":"Computational structural prediction, site-directed mutagenesis of histidine residues, electrophysiology (membrane conductance assay), small molecule permeability assays","journal":"mBio","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — mutagenesis with functional readout plus independent electrophysiological validation in a single rigorous study with multiple orthogonal methods","pmids":["38380952"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2024,"finding":"Deletion of GRA47 causes 'bubble vacuole' morphology with reduced small molecule permeability across the PVM, reduced in vitro proliferation, and attenuated virulence in mice, phenocopying Δgra17 and Δgra72 parasites.","method":"Genetic knockout (Δgra47), in vitro plaque/proliferation assay, mouse virulence model, fluorescent small molecule permeability assay","journal":"mBio","confidence":"High","confidence_rationale":"Tier 2 / Strong — clean genetic knockout with multiple defined phenotypic readouts, independently replicated in two publications (PMID:38380952 and PMID:38936501)","pmids":["38380952","38936501"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2024,"finding":"Overexpression of GRA47 (or GRA72 or GRA17) in Δgra47, Δgra72, or Δgra17 strains partially rescued aberrant PV morphology, while expression of the Plasmodium falciparum PVM protein PfExp2 (an ortholog of GRA17/GRA23) fully rescued PV morphological defects in all three knockout strains, indicating these GRA proteins are not fully functionally redundant but regulate nutrient acquisition through different mechanisms.","method":"Genetic complementation (overexpression of GRA47, GRA72, GRA17, and PfExp2), PV morphology quantification","journal":"International journal for parasitology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic complementation with quantitative morphological readout, single lab, single publication","pmids":["38936501"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2024,"finding":"Loss of GRA72 in a Δgra23 background (Δgra23Δgra72 double knockout) significantly increased the proportion of aberrant PVs compared with Δgra23 alone, placing GRA72 (and by association, GRA47 through its interaction with GRA72) in the same functional pathway as GRA23 in PV morphology maintenance.","method":"Double gene deletion (Δgra23Δgra72), PV morphology quantification","journal":"International journal for parasitology","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — genetic epistasis via double knockout with morphological readout, single lab, single publication; GRA47 implicated indirectly through its interaction with GRA72","pmids":["38936501"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2023,"finding":"Preprint version of the mBio findings: GRA47 interacts with GRA72 (immunoprecipitation), forms a heptameric pore with pore-lining histidines critical for function (mutagenesis), and its deletion reduces PVM permeability and membrane conductance (electrophysiology), reducing parasite growth and virulence in mice.","method":"Immunoprecipitation, genetic deletion, structural prediction, histidine mutagenesis, electrophysiology, small molecule permeability assay, mouse virulence model","journal":"bioRxiv","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — multiple orthogonal methods in a single rigorous study; this is the preprint version of the peer-reviewed PMID:38380952","pmids":["38014337"],"is_preprint":true,"attribution":[{"species":"Toxoplasma gondii","strain":""}]}],"current_model":"GRA47 is a dense granule protein that localizes to the Toxoplasma gondii parasitophorous vacuole membrane (PVM), where it physically interacts with GRA72 and forms a heptameric pore structure; conserved pore-lining histidine residues are essential for its function, and GRA47 acts together with GRA72, GRA17, and GRA23 to regulate PVM permeability to small molecules, parasite proliferation, and virulence in mice."},"narrative":{"mechanistic_narrative":"GRA47 is a Toxoplasma gondii dense granule protein that localizes to the parasitophorous vacuole membrane (PVM), where it controls PVM permeability to small molecules and maintains normal PV morphology [PMID:38380952, PMID:38936501]. Structural modelling predicts that GRA47 assembles into a heptameric pore lined by conserved histidine residues; mutation of these histidines abolishes function and loss of GRA47 alters membrane conductance, establishing a pore-forming role at the PVM [PMID:38380952]. GRA47 physically interacts with GRA72 [PMID:38380952, PMID:38936501], and its deletion produces a 'bubble vacuole' morphology with reduced permeability, impaired in vitro proliferation, and attenuated virulence in mice, phenocopying Δgra17 and Δgra72 parasites [PMID:38380952, PMID:38936501]. Complementation and epistasis experiments place GRA47/GRA72 in a shared functional pathway with GRA17 and GRA23 in nutrient acquisition and PVM permeability, though the GRA proteins are not fully functionally redundant [PMID:38936501].","teleology":[{"year":2024,"claim":"Establishing that GRA47 is a PVM protein that physically partners with GRA72 and governs vacuole morphology and small-molecule permeability defined its cellular role in the host-parasite interface.","evidence":"Co-immunoprecipitation, Δgra47 knockout, permeability assays, and live imaging in T. gondii","pmids":["38380952","38936501"],"confidence":"High","gaps":["Molecular basis of the GRA47-GRA72 interaction (interface, stoichiometry) not resolved","Which specific small-molecule metabolites traverse the pore not identified"],"attribution":[]},{"year":2024,"claim":"Structural prediction combined with histidine mutagenesis and electrophysiology answered how GRA47 acts mechanistically, identifying it as a heptameric pore with conserved pore-lining histidines essential for conductance and permeability.","evidence":"Computational structural prediction, site-directed histidine mutagenesis with functional readout, and membrane conductance measurement in T. gondii","pmids":["38380952"],"confidence":"High","gaps":["Experimental high-resolution structure of the pore not determined","Whether GRA47 homo-oligomerizes or forms hetero-oligomers with GRA72 in the pore not established"],"attribution":[]},{"year":2024,"claim":"Deletion of GRA47 phenocopying Δgra17 and Δgra72 across proliferation and mouse virulence readouts connected pore function to parasite fitness and pathogenesis.","evidence":"Δgra47 knockout with plaque/proliferation assays and a mouse virulence model in T. gondii","pmids":["38380952","38936501"],"confidence":"High","gaps":["Causal chain from permeability defect to virulence attenuation not dissected","Host cell type dependence of the phenotype not explored"],"attribution":[]},{"year":2024,"claim":"Complementation with cognate GRAs and the Plasmodium ortholog PfExp2, plus Δgra23Δgra72 epistasis, positioned GRA47/GRA72 within a broader GRA17/GRA23 nutrient-acquisition pathway while showing the proteins are not fully redundant.","evidence":"Genetic complementation (GRA47/GRA72/GRA17/PfExp2 overexpression) and double-knockout epistasis with PV morphology quantification in T. gondii","pmids":["38936501"],"confidence":"Medium","gaps":["Distinct mechanisms distinguishing the GRA pores not defined","GRA47 placement in the GRA23 pathway inferred indirectly through GRA72"],"attribution":[]},{"year":null,"claim":"The identity of the substrates transported through the GRA47 pore and the precise architecture of the GRA47-GRA72 complex remain to be determined.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No experimental structure of the pore","Transported metabolites unidentified","Stoichiometry of GRA47-GRA72 assembly unknown"],"attribution":[]}],"mechanism_profile":{"molecular_activity":[],"localization":[],"pathway":[],"complexes":["GRA47-GRA72 complex"],"partners":["GRA72","GRA17","GRA23"],"other_free_text":["GRA47 is predicted to assemble into a heptameric pore with pore-lining histidine residues essential for function [#1].","Δgra47 produces a 'bubble vacuole' morphology and attenuates virulence in mice, phenocopying Δgra17 and Δgra72 [#2].","The Plasmodium falciparum PVM protein PfExp2 fully rescues PV morphology in Δgra47, Δgra72, and Δgra17 strains [#3]."]},"strain_conflicts":[]},"prefetch_data":{},"corpus_meta":[{"pmid":"38380952","id":"PMC_38380952","title":"Two Toxoplasma gondii putative pore-forming proteins, GRA47 and GRA72, influence small molecule permeability of the parasitophorous vacuole.","date":"2024","source":"mBio","url":"https://pubmed.ncbi.nlm.nih.gov/38380952","citation_count":24,"is_preprint":false},{"pmid":"38936501","id":"PMC_38936501","title":"GRA47 is important for the morphology and permeability of the parasitophorous vacuole in Toxoplasma gondii.","date":"2024","source":"International journal for parasitology","url":"https://pubmed.ncbi.nlm.nih.gov/38936501","citation_count":11,"is_preprint":false},{"pmid":"38014337","id":"PMC_38014337","title":"GRA47 and GRA72 are Toxoplasma gondii pore-forming proteins that influence small molecule permeability of the parasitophorous vacuole.","date":"2023","source":"bioRxiv : the preprint server for biology","url":"https://pubmed.ncbi.nlm.nih.gov/38014337","citation_count":1,"is_preprint":false},{"pmid":"42357722","id":"PMC_42357722","title":"Transcriptomic Profiling of GRA47 Deletion in Toxoplasma gondii Reveals Transcriptional Reprogramming of Stress Adaptation and Metabolic Compensation.","date":"2026","source":"Veterinary sciences","url":"https://pubmed.ncbi.nlm.nih.gov/42357722","citation_count":0,"is_preprint":false},{"pmid":"42253329","id":"PMC_42253329","title":"Protective Immunity Induced by DNA Vaccines Encoding TgGRA47 and TgGRA72 Against Toxoplasma gondii Infection in BALB/c Mice.","date":"2026","source":"Transboundary and emerging diseases","url":"https://pubmed.ncbi.nlm.nih.gov/42253329","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":5110,"output_tokens":1739,"usd":0.020708,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":6890,"output_tokens":2102,"usd":0.0435,"stage2_stop_reason":"end_turn"},"total_usd":0.064208,"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\": 2024,\n      \"finding\": \"GRA47 physically interacts with GRA72, as demonstrated by immunoprecipitation experiments; GRA47 localizes to the parasitophorous vacuole membrane (PVM) and is required for normal PV morphology and PVM permeability to small molecules.\",\n      \"method\": \"Co-immunoprecipitation, genetic deletion (Δgra47), small molecule permeability assays, live imaging of vacuole morphology\",\n      \"journal\": \"mBio\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP plus genetic knockout with defined cellular phenotype, independently replicated in two separate publications (PMID:38380952 and PMID:38936501)\",\n      \"pmids\": [\"38380952\", \"38936501\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"GRA47 is predicted by structural modelling to form a heptameric pore, with conserved histidine residues lining the pore channel; mutational analysis of these histidines abolished protein functionality (aberrant PV morphology, reduced permeability), and electrophysiology confirmed that loss of GRA47 alters membrane conductance, corroborating its pore-forming role.\",\n      \"method\": \"Computational structural prediction, site-directed mutagenesis of histidine residues, electrophysiology (membrane conductance assay), small molecule permeability assays\",\n      \"journal\": \"mBio\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — mutagenesis with functional readout plus independent electrophysiological validation in a single rigorous study with multiple orthogonal methods\",\n      \"pmids\": [\"38380952\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Deletion of GRA47 causes 'bubble vacuole' morphology with reduced small molecule permeability across the PVM, reduced in vitro proliferation, and attenuated virulence in mice, phenocopying Δgra17 and Δgra72 parasites.\",\n      \"method\": \"Genetic knockout (Δgra47), in vitro plaque/proliferation assay, mouse virulence model, fluorescent small molecule permeability assay\",\n      \"journal\": \"mBio\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — clean genetic knockout with multiple defined phenotypic readouts, independently replicated in two publications (PMID:38380952 and PMID:38936501)\",\n      \"pmids\": [\"38380952\", \"38936501\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Overexpression of GRA47 (or GRA72 or GRA17) in Δgra47, Δgra72, or Δgra17 strains partially rescued aberrant PV morphology, while expression of the Plasmodium falciparum PVM protein PfExp2 (an ortholog of GRA17/GRA23) fully rescued PV morphological defects in all three knockout strains, indicating these GRA proteins are not fully functionally redundant but regulate nutrient acquisition through different mechanisms.\",\n      \"method\": \"Genetic complementation (overexpression of GRA47, GRA72, GRA17, and PfExp2), PV morphology quantification\",\n      \"journal\": \"International journal for parasitology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic complementation with quantitative morphological readout, single lab, single publication\",\n      \"pmids\": [\"38936501\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Loss of GRA72 in a Δgra23 background (Δgra23Δgra72 double knockout) significantly increased the proportion of aberrant PVs compared with Δgra23 alone, placing GRA72 (and by association, GRA47 through its interaction with GRA72) in the same functional pathway as GRA23 in PV morphology maintenance.\",\n      \"method\": \"Double gene deletion (Δgra23Δgra72), PV morphology quantification\",\n      \"journal\": \"International journal for parasitology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — genetic epistasis via double knockout with morphological readout, single lab, single publication; GRA47 implicated indirectly through its interaction with GRA72\",\n      \"pmids\": [\"38936501\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Preprint version of the mBio findings: GRA47 interacts with GRA72 (immunoprecipitation), forms a heptameric pore with pore-lining histidines critical for function (mutagenesis), and its deletion reduces PVM permeability and membrane conductance (electrophysiology), reducing parasite growth and virulence in mice.\",\n      \"method\": \"Immunoprecipitation, genetic deletion, structural prediction, histidine mutagenesis, electrophysiology, small molecule permeability assay, mouse virulence model\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — multiple orthogonal methods in a single rigorous study; this is the preprint version of the peer-reviewed PMID:38380952\",\n      \"pmids\": [\"38014337\"],\n      \"is_preprint\": true,\n      \"attribution\": [\n        {\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}\n      ]\n    }\n  ],\n  \"current_model\": \"GRA47 is a dense granule protein that localizes to the Toxoplasma gondii parasitophorous vacuole membrane (PVM), where it physically interacts with GRA72 and forms a heptameric pore structure; conserved pore-lining histidine residues are essential for its function, and GRA47 acts together with GRA72, GRA17, and GRA23 to regulate PVM permeability to small molecules, parasite proliferation, and virulence in mice.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"GRA47 is a Toxoplasma gondii dense granule protein that localizes to the parasitophorous vacuole membrane (PVM), where it controls PVM permeability to small molecules and maintains normal PV morphology [#0, #2]. Structural modelling predicts that GRA47 assembles into a heptameric pore lined by conserved histidine residues; mutation of these histidines abolishes function and loss of GRA47 alters membrane conductance, establishing a pore-forming role at the PVM [#1]. GRA47 physically interacts with GRA72 [#0], and its deletion produces a 'bubble vacuole' morphology with reduced permeability, impaired in vitro proliferation, and attenuated virulence in mice, phenocopying \\u0394gra17 and \\u0394gra72 parasites [#2]. Complementation and epistasis experiments place GRA47/GRA72 in a shared functional pathway with GRA17 and GRA23 in nutrient acquisition and PVM permeability, though the GRA proteins are not fully functionally redundant [#3, #4].\",\n  \"teleology\": [\n    {\n      \"year\": 2024,\n      \"claim\": \"Establishing that GRA47 is a PVM protein that physically partners with GRA72 and governs vacuole morphology and small-molecule permeability defined its cellular role in the host-parasite interface.\",\n      \"evidence\": \"Co-immunoprecipitation, \\u0394gra47 knockout, permeability assays, and live imaging in T. gondii\",\n      \"pmids\": [\"38380952\", \"38936501\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular basis of the GRA47-GRA72 interaction (interface, stoichiometry) not resolved\", \"Which specific small-molecule metabolites traverse the pore not identified\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Structural prediction combined with histidine mutagenesis and electrophysiology answered how GRA47 acts mechanistically, identifying it as a heptameric pore with conserved pore-lining histidines essential for conductance and permeability.\",\n      \"evidence\": \"Computational structural prediction, site-directed histidine mutagenesis with functional readout, and membrane conductance measurement in T. gondii\",\n      \"pmids\": [\"38380952\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Experimental high-resolution structure of the pore not determined\", \"Whether GRA47 homo-oligomerizes or forms hetero-oligomers with GRA72 in the pore not established\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Deletion of GRA47 phenocopying \\u0394gra17 and \\u0394gra72 across proliferation and mouse virulence readouts connected pore function to parasite fitness and pathogenesis.\",\n      \"evidence\": \"\\u0394gra47 knockout with plaque/proliferation assays and a mouse virulence model in T. gondii\",\n      \"pmids\": [\"38380952\", \"38936501\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Causal chain from permeability defect to virulence attenuation not dissected\", \"Host cell type dependence of the phenotype not explored\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Complementation with cognate GRAs and the Plasmodium ortholog PfExp2, plus \\u0394gra23\\u0394gra72 epistasis, positioned GRA47/GRA72 within a broader GRA17/GRA23 nutrient-acquisition pathway while showing the proteins are not fully redundant.\",\n      \"evidence\": \"Genetic complementation (GRA47/GRA72/GRA17/PfExp2 overexpression) and double-knockout epistasis with PV morphology quantification in T. gondii\",\n      \"pmids\": [\"38936501\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Distinct mechanisms distinguishing the GRA pores not defined\", \"GRA47 placement in the GRA23 pathway inferred indirectly through GRA72\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The identity of the substrates transported through the GRA47 pore and the precise architecture of the GRA47-GRA72 complex remain to be determined.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No experimental structure of the pore\", \"Transported metabolites unidentified\", \"Stoichiometry of GRA47-GRA72 assembly unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0015267\", \"supporting_discovery_ids\": [1], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]},\n      {\"term_id\": \"GO:0022829\", \"supporting_discovery_ids\": [0, 1, 2], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0020005\", \"supporting_discovery_ids\": [0], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"GO:0015850\", \"supporting_discovery_ids\": [2, 3], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]}\n    ],\n    \"complexes\": [\"GRA47-GRA72 complex\"],\n    \"partners\": [\"GRA72\", \"GRA17\", \"GRA23\"],\n    \"other_free_text\": [\n      \"GRA47 is predicted to assemble into a heptameric pore with pore-lining histidine residues essential for function [#1].\",\n      \"\\u0394gra47 produces a 'bubble vacuole' morphology and attenuates virulence in mice, phenocopying \\u0394gra17 and \\u0394gra72 [#2].\",\n      \"The Plasmodium falciparum PVM protein PfExp2 fully rescues PV morphology in \\u0394gra47, \\u0394gra72, and \\u0394gra17 strains [#3].\"\n    ]\n  },\n  \"strain_conflicts\": []\n}","audit_flag":null,"strains":[{"strain":"ME49","gene_id":"TGME49_254000","name":"GRA47","product":"dense granule protein GRA47","uniprot_acc":"S8GIS3","chromosome":"TGME49_chrIII","start_pos":1391826},{"strain":"GT1","gene_id":"TGGT1_254000","name":null,"product":"hypothetical protein","uniprot_acc":null,"chromosome":"TGGT1_chrIII","start_pos":1373982},{"strain":"RH88","gene_id":"TGRH88_004350","name":null,"product":"unspecified product","uniprot_acc":null,"chromosome":"CM023084","start_pos":1212208},{"strain":"VEG","gene_id":"TGVEG_254000","name":null,"product":"hypothetical protein","uniprot_acc":"V4ZD55","chromosome":"TGVEG_chrIII","start_pos":1401083}]}