{"gene":"TG_270320","gene_symbol":"PPM3C","run_date":"2026-09-08T16:59:55+00:00","timeline":{"discoveries":[{"year":2020,"finding":"PPM3C (TGGT1_270320) was identified as a MYR1-interacting protein localized to the parasitophorous vacuole (PV) of Toxoplasma gondii, discovered via affinity purification of MYR1 followed by mass spectrometry.","method":"Affinity purification / co-immunoprecipitation with MYR1, mass spectrometry","journal":"mSphere","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — reciprocal Co-IP/affinity purification with MS identification, single lab, single method for this specific protein","pmids":["32075880"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":"TGGT1"}]},{"year":2020,"finding":"TgPPM3C is a secreted protein containing a PP2C-class serine/threonine phosphatase domain that localizes to the parasitophorous vacuole; parasites lacking TgPPM3C are defective specifically in the export of GRA16 and GRA28 effectors into the host cell, but not other MYR1-dependent effectors.","method":"Gene knockout (CRISPR/KO), effector export assays (GRA16, GRA28 localization), complementation","journal":"PLoS pathogens","confidence":"High","confidence_rationale":"Tier 2 / Moderate — clean KO with defined cellular phenotype (selective export defect), multiple effectors tested, in vivo and in vitro assays, single lab","pmids":["33370417"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2020,"finding":"Phosphoproteomic analysis of TgPPM3C-deleted parasites revealed altered phosphorylation of secreted vacuolar proteins including GRA16, GRA28, and MYR1, indicating TgPPM3C functions as a phosphatase that dephosphorylates these PV substrates.","method":"Phosphoproteomics (mass spectrometry) of KO parasite cultures","journal":"PLoS pathogens","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — phosphoproteomic profiling in KO background, single lab, no direct in vitro phosphatase reconstitution","pmids":["33370417"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2020,"finding":"Phosphomimetic mutation of two serine residues in GRA16 caused GRA16 export defects, establishing that dephosphorylation of GRA16 by TgPPM3C is a critical step required for its translocation across the PV membrane into the host cell.","method":"Site-directed mutagenesis (phosphomimetic serine substitutions in GRA16), effector export assay","journal":"PLoS pathogens","confidence":"High","confidence_rationale":"Tier 1 / Moderate — mutagenesis with direct functional readout (export assay), mechanistically links phosphorylation state to export, single lab","pmids":["33370417"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2020,"finding":"TgPPM3C-deficient parasites are avirulent during acute infection in mice and form fewer cysts in mouse brain during chronic infection, indicating TgPPM3C-mediated effector export is required for full parasite virulence in vivo.","method":"Mouse infection assay (acute virulence, chronic cyst burden) with KO parasites","journal":"PLoS pathogens","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — defined in vivo phenotype with KO, single lab, phenotype linked to molecular mechanism via export defect","pmids":["33370417"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]}],"current_model":"TgPPM3C is a PP2C-class serine/threonine phosphatase secreted into the Toxoplasma gondii parasitophorous vacuole, where it interacts with MYR1 and dephosphorylates specific substrates including GRA16 and GRA28; this dephosphorylation is required for selective export of these effectors across the vacuole membrane into the host cell, and loss of TgPPM3C causes avirulence in mice and reduced chronic cyst formation."},"narrative":{"mechanistic_narrative":"TgPPM3C is a secreted PP2C-class serine/threonine phosphatase of Toxoplasma gondii that acts within the parasitophorous vacuole (PV) to control selective export of parasite effectors into the host cell [PMID:33370417]. It was identified as a MYR1-interacting protein localized to the PV [PMID:32075880], and phosphoproteomics of knockout parasites shows altered phosphorylation of vacuolar substrates including GRA16, GRA28, and MYR1, consistent with TgPPM3C dephosphorylating these PV proteins [PMID:33370417]. Functionally, loss of TgPPM3C specifically blocks export of the GRA16 and GRA28 effectors across the PV membrane while sparing other MYR1-dependent effectors [PMID:33370417], and phosphomimetic substitution of two GRA16 serines reproduces the export defect, establishing that TgPPM3C-mediated dephosphorylation of GRA16 is a required step for its translocation into the host cell [PMID:33370417]. This effector export is required for pathogenesis: TgPPM3C-deficient parasites are avirulent in acute mouse infection and form fewer brain cysts during chronic infection [PMID:33370417]. Direct in vitro reconstitution of TgPPM3C phosphatase activity on its substrates has not been characterized in the available corpus.","teleology":[{"year":2020,"claim":"Establishing what protein partners MYR1 in the PV identified TgPPM3C, connecting a phosphatase to the known MYR1-dependent effector export machinery.","evidence":"Affinity purification of MYR1 with mass spectrometry in T. gondii TGGT1","pmids":["32075880"],"confidence":"Medium","gaps":["Single method (affinity purification/MS) without reciprocal orthogonal validation","Interaction stoichiometry and whether binding is direct not established","No functional consequence of the MYR1 interaction defined in this study"],"attribution":[{"species":"Toxoplasma gondii","strain":"TGGT1"}]},{"year":2020,"claim":"Knockout defined TgPPM3C as a secreted PP2C-domain phosphatase whose loss selectively blocks export of GRA16 and GRA28 but not other MYR1-dependent effectors, showing it acts as a substrate-selective export determinant.","evidence":"CRISPR knockout with complementation and effector export/localization assays in T. gondii","pmids":["33370417"],"confidence":"High","gaps":["Basis of substrate selectivity for GRA16/GRA28 over other effectors unresolved","Whether TgPPM3C phosphatase activity per se is required not directly tested by catalytic-dead mutant here"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2020,"claim":"Phosphoproteomics of the knockout revealed hyperphosphorylation of vacuolar substrates (GRA16, GRA28, MYR1), linking the export defect to a loss of dephosphorylation activity in the PV.","evidence":"Phosphoproteomic mass spectrometry of KO parasite cultures","pmids":["33370417"],"confidence":"Medium","gaps":["No in vitro phosphatase reconstitution demonstrating direct dephosphorylation of these substrates","Cannot distinguish direct substrates from indirect phosphorylation changes"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2020,"claim":"Phosphomimetic mutagenesis of two GRA16 serines recapitulated the export block, causally establishing that dephosphorylation of GRA16 is required for its translocation across the PV membrane.","evidence":"Site-directed phosphomimetic serine substitutions in GRA16 with export assay in T. gondii","pmids":["33370417"],"confidence":"High","gaps":["Whether the same phospho-switch mechanism governs GRA28 export not shown","Kinase counteracting TgPPM3C on these sites not identified"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2020,"claim":"In vivo infection linked the molecular export defect to pathogenesis, showing TgPPM3C is required for acute virulence and efficient chronic cyst formation.","evidence":"Mouse acute virulence and chronic brain cyst burden assays with KO parasites","pmids":["33370417"],"confidence":"Medium","gaps":["Which exported effector(s) drive the virulence phenotype not dissected","Host pathway downstream of GRA16/GRA28 export in vivo not mapped"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":null,"claim":"Direct biochemical demonstration of TgPPM3C catalytic activity on GRA16/GRA28 and the structural basis of its substrate selectivity remain open.","evidence":"No in vitro reconstitution or structural study in the available corpus","pmids":[],"confidence":"Medium","gaps":["No purified-enzyme phosphatase assay against defined substrates","No structure or catalytic-dead complementation defining the active site's role in export","Mechanism coupling dephosphorylation to membrane translocation unknown"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]}],"mechanism_profile":{"molecular_activity":[],"localization":[],"pathway":[],"complexes":[],"partners":["MYR1 (INTERACTOR; PV-LOCALIZED) [#0]","GRA16 (SUBSTRATE; DEPHOSPHORYLATION REQUIRED FOR EXPORT) [#2, #3]","GRA28 (SUBSTRATE; EXPORT-DEPENDENT ON TGPPM3C) [#1, #2]"],"other_free_text":["PP2C-class serine/threonine phosphatase domain [#1]","Loss of TgPPM3C causes avirulence in acute mouse infection and reduced chronic brain cyst formation [#4]","Export defect is selective for GRA16/GRA28 and spares other MYR1-dependent effectors [#1]"]},"strain_conflicts":[]},"prefetch_data":{},"corpus_meta":[{"pmid":"32075880","id":"PMC_32075880","title":"Coimmunoprecipitation with MYR1 Identifies Three Additional Proteins within the Toxoplasma gondii Parasitophorous Vacuole Required for Translocation of Dense Granule Effectors into Host Cells.","date":"2020","source":"mSphere","url":"https://pubmed.ncbi.nlm.nih.gov/32075880","citation_count":53,"is_preprint":false},{"pmid":"33370417","id":"PMC_33370417","title":"Toxoplasma gondii PPM3C, a secreted protein phosphatase, affects parasitophorous vacuole effector export.","date":"2020","source":"PLoS pathogens","url":"https://pubmed.ncbi.nlm.nih.gov/33370417","citation_count":20,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":3600,"output_tokens":1323,"usd":0.015322,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":6283,"output_tokens":2557,"usd":0.04767,"stage2_stop_reason":"end_turn"},"total_usd":0.0,"stage1_batch_id":"msgbatch_01XFXJD6iZecXyWn9uU5Qom9","stage2_batch_id":"msgbatch_013GToKgs4AKSCRE4yvueviH","note":"batch pricing = 50% of standard","skipped":true},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2020,\n      \"finding\": \"PPM3C (TGGT1_270320) was identified as a MYR1-interacting protein localized to the parasitophorous vacuole (PV) of Toxoplasma gondii, discovered via affinity purification of MYR1 followed by mass spectrometry.\",\n      \"method\": \"Affinity purification / co-immunoprecipitation with MYR1, mass spectrometry\",\n      \"journal\": \"mSphere\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — reciprocal Co-IP/affinity purification with MS identification, single lab, single method for this specific protein\",\n      \"pmids\": [\"32075880\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"TGGT1\"}]\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"TgPPM3C is a secreted protein containing a PP2C-class serine/threonine phosphatase domain that localizes to the parasitophorous vacuole; parasites lacking TgPPM3C are defective specifically in the export of GRA16 and GRA28 effectors into the host cell, but not other MYR1-dependent effectors.\",\n      \"method\": \"Gene knockout (CRISPR/KO), effector export assays (GRA16, GRA28 localization), complementation\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — clean KO with defined cellular phenotype (selective export defect), multiple effectors tested, in vivo and in vitro assays, single lab\",\n      \"pmids\": [\"33370417\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Phosphoproteomic analysis of TgPPM3C-deleted parasites revealed altered phosphorylation of secreted vacuolar proteins including GRA16, GRA28, and MYR1, indicating TgPPM3C functions as a phosphatase that dephosphorylates these PV substrates.\",\n      \"method\": \"Phosphoproteomics (mass spectrometry) of KO parasite cultures\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — phosphoproteomic profiling in KO background, single lab, no direct in vitro phosphatase reconstitution\",\n      \"pmids\": [\"33370417\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Phosphomimetic mutation of two serine residues in GRA16 caused GRA16 export defects, establishing that dephosphorylation of GRA16 by TgPPM3C is a critical step required for its translocation across the PV membrane into the host cell.\",\n      \"method\": \"Site-directed mutagenesis (phosphomimetic serine substitutions in GRA16), effector export assay\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — mutagenesis with direct functional readout (export assay), mechanistically links phosphorylation state to export, single lab\",\n      \"pmids\": [\"33370417\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"TgPPM3C-deficient parasites are avirulent during acute infection in mice and form fewer cysts in mouse brain during chronic infection, indicating TgPPM3C-mediated effector export is required for full parasite virulence in vivo.\",\n      \"method\": \"Mouse infection assay (acute virulence, chronic cyst burden) with KO parasites\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — defined in vivo phenotype with KO, single lab, phenotype linked to molecular mechanism via export defect\",\n      \"pmids\": [\"33370417\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    }\n  ],\n  \"current_model\": \"TgPPM3C is a PP2C-class serine/threonine phosphatase secreted into the Toxoplasma gondii parasitophorous vacuole, where it interacts with MYR1 and dephosphorylates specific substrates including GRA16 and GRA28; this dephosphorylation is required for selective export of these effectors across the vacuole membrane into the host cell, and loss of TgPPM3C causes avirulence in mice and reduced chronic cyst formation.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"TgPPM3C is a secreted PP2C-class serine/threonine phosphatase of Toxoplasma gondii that acts within the parasitophorous vacuole (PV) to control selective export of parasite effectors into the host cell [#1]. It was identified as a MYR1-interacting protein localized to the PV [#0], and phosphoproteomics of knockout parasites shows altered phosphorylation of vacuolar substrates including GRA16, GRA28, and MYR1, consistent with TgPPM3C dephosphorylating these PV proteins [#2]. Functionally, loss of TgPPM3C specifically blocks export of the GRA16 and GRA28 effectors across the PV membrane while sparing other MYR1-dependent effectors [#1], and phosphomimetic substitution of two GRA16 serines reproduces the export defect, establishing that TgPPM3C-mediated dephosphorylation of GRA16 is a required step for its translocation into the host cell [#3]. This effector export is required for pathogenesis: TgPPM3C-deficient parasites are avirulent in acute mouse infection and form fewer brain cysts during chronic infection [#4]. Direct in vitro reconstitution of TgPPM3C phosphatase activity on its substrates has not been characterized in the available corpus.\",\n  \"teleology\": [\n    {\n      \"year\": 2020,\n      \"claim\": \"Establishing what protein partners MYR1 in the PV identified TgPPM3C, connecting a phosphatase to the known MYR1-dependent effector export machinery.\",\n      \"evidence\": \"Affinity purification of MYR1 with mass spectrometry in T. gondii TGGT1\",\n      \"pmids\": [\"32075880\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Single method (affinity purification/MS) without reciprocal orthogonal validation\",\n        \"Interaction stoichiometry and whether binding is direct not established\",\n        \"No functional consequence of the MYR1 interaction defined in this study\"\n      ],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"TGGT1\"}]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Knockout defined TgPPM3C as a secreted PP2C-domain phosphatase whose loss selectively blocks export of GRA16 and GRA28 but not other MYR1-dependent effectors, showing it acts as a substrate-selective export determinant.\",\n      \"evidence\": \"CRISPR knockout with complementation and effector export/localization assays in T. gondii\",\n      \"pmids\": [\"33370417\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Basis of substrate selectivity for GRA16/GRA28 over other effectors unresolved\",\n        \"Whether TgPPM3C phosphatase activity per se is required not directly tested by catalytic-dead mutant here\"\n      ],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Phosphoproteomics of the knockout revealed hyperphosphorylation of vacuolar substrates (GRA16, GRA28, MYR1), linking the export defect to a loss of dephosphorylation activity in the PV.\",\n      \"evidence\": \"Phosphoproteomic mass spectrometry of KO parasite cultures\",\n      \"pmids\": [\"33370417\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"No in vitro phosphatase reconstitution demonstrating direct dephosphorylation of these substrates\",\n        \"Cannot distinguish direct substrates from indirect phosphorylation changes\"\n      ],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Phosphomimetic mutagenesis of two GRA16 serines recapitulated the export block, causally establishing that dephosphorylation of GRA16 is required for its translocation across the PV membrane.\",\n      \"evidence\": \"Site-directed phosphomimetic serine substitutions in GRA16 with export assay in T. gondii\",\n      \"pmids\": [\"33370417\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Whether the same phospho-switch mechanism governs GRA28 export not shown\",\n        \"Kinase counteracting TgPPM3C on these sites not identified\"\n      ],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"In vivo infection linked the molecular export defect to pathogenesis, showing TgPPM3C is required for acute virulence and efficient chronic cyst formation.\",\n      \"evidence\": \"Mouse acute virulence and chronic brain cyst burden assays with KO parasites\",\n      \"pmids\": [\"33370417\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Which exported effector(s) drive the virulence phenotype not dissected\",\n        \"Host pathway downstream of GRA16/GRA28 export in vivo not mapped\"\n      ],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"Direct biochemical demonstration of TgPPM3C catalytic activity on GRA16/GRA28 and the structural basis of its substrate selectivity remain open.\",\n      \"evidence\": \"No in vitro reconstitution or structural study in the available corpus\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"No purified-enzyme phosphatase assay against defined substrates\",\n        \"No structure or catalytic-dead complementation defining the active site's role in export\",\n        \"Mechanism coupling dephosphorylation to membrane translocation unknown\"\n      ],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0004722\", \"supporting_discovery_ids\": [1, 2, 3], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0020003\", \"supporting_discovery_ids\": [0, 1], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"TGGT1\"}]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"GO:0140321\", \"supporting_discovery_ids\": [1, 3], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\n      \"MYR1 (interactor; PV-localized) [#0]\",\n      \"GRA16 (substrate; dephosphorylation required for export) [#2, #3]\",\n      \"GRA28 (substrate; export-dependent on TgPPM3C) [#1, #2]\"\n    ],\n    \"other_free_text\": [\n      \"PP2C-class serine/threonine phosphatase domain [#1]\",\n      \"Loss of TgPPM3C causes avirulence in acute mouse infection and reduced chronic brain cyst formation [#4]\",\n      \"Export defect is selective for GRA16/GRA28 and spares other MYR1-dependent effectors [#1]\"\n    ]\n  },\n  \"strain_conflicts\": []\n}","audit_flag":null,"strains":[{"strain":"ME49","gene_id":"TGME49_270320","name":"PPM3C","product":"protein phosphatase PPM3C","uniprot_acc":"A0A125YZH2","chromosome":"TGME49_chrVIII","start_pos":5274034},{"strain":"GT1","gene_id":"TGGT1_270320","name":null,"product":"protein phosphatase 2C domain-containing protein","uniprot_acc":null,"chromosome":"TGGT1_chrVIII","start_pos":5274953},{"strain":"RH88","gene_id":"TGRH88_081050","name":null,"product":"protein phosphatase 2C domain-containing protein","uniprot_acc":null,"chromosome":"CM023089","start_pos":10395585},{"strain":"VEG","gene_id":"TGVEG_270320","name":null,"product":"protein phosphatase 2C domain-containing protein","uniprot_acc":"B9PHR8","chromosome":"TGVEG_chrVIII","start_pos":5290235}]}