{"gene":"PF3D7_0304200","run_date":"2026-09-08T16:26:55","timeline":{"discoveries":[{"year":2015,"finding":"EHD (PfEHD) is a membrane-bound protein that localizes to vesicles originating at the parasite plasma membrane; these vesicles migrate through the parasite cytosol and accumulate into a large multi-vesicular-like structure juxtaposed to but outside the food vacuole, which co-stains with the neutral lipid dye Nile Red, identifying it as a neutral lipid storage site.","method":"GFP-fusion live imaging, time-lapse microscopy, Nile Red co-staining, food-vacuole membrane co-staining","journal":"Biochimica et biophysica acta","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct live-cell imaging with GFP fusion and multiple co-localization readouts, single lab","pmids":["26284889"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2015,"finding":"EHD (PfEHD) co-purifies with endocytosis modulators as identified by proteomic analysis, supporting its role in the endocytic machinery of the parasite.","method":"Proteomic analysis (co-immunoprecipitation/pulldown followed by mass spectrometry)","journal":"Biochimica et biophysica acta","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single proteomics experiment, single lab, no reciprocal validation reported in abstract","pmids":["26284889"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2015,"finding":"Treatment with endocytosis inhibitors obstructed development of EHD (PfEHD)-labeled vesicles and blocked their targeting to the neutral lipid storage site near the food vacuole, placing EHD functionally in the endocytic vesicle biogenesis/trafficking pathway.","method":"Pharmacological inhibition of endocytosis combined with live GFP-fusion imaging","journal":"Biochimica et biophysica acta","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — functional inhibitor experiment with direct imaging readout, single lab, one study","pmids":["26284889"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2020,"finding":"EHD (PfEHD) co-immunoprecipitates with the AP-2μ subunit of the AP-2 adaptin complex in P. falciparum, identifying it as an AP-2-interacting protein and linking it to a clathrin-independent endosomal transport pathway.","method":"Co-immunoprecipitation (HA-tagged AP-2μ) followed by mass spectrometry; confirmed by reverse immunoprecipitation controls","journal":"mBio","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — co-IP/MS identification of EHD as AP-2μ interactor, replicated conceptually by two labs citing same finding, but mechanistic follow-up on EHD itself is limited","pmids":["32098816"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]}],"current_model":"EHD (PF3D7_0304200) is a membrane-bound NTPase that associates with vesicles originating at the parasite plasma membrane, traffics them to a neutral lipid storage site near the food vacuole, physically interacts with the AP-2 adaptin complex, and is functionally required for endocytic vesicle biogenesis and lipid mobilization during the intra-erythrocytic stages of P. falciparum."},"narrative":{"mechanistic_narrative":"EHD (PF3D7_0304200) is a membrane-bound protein that functions in endocytic vesicle biogenesis and lipid mobilization during the intra-erythrocytic development of Plasmodium falciparum [PMID:26284889]. It localizes to vesicles that originate at the parasite plasma membrane and migrate through the cytosol to accumulate in a large multi-vesicular-like structure juxtaposed to the food vacuole; this structure co-stains with Nile Red, marking it as a neutral lipid storage site [PMID:26284889]. Pharmacological inhibition of endocytosis blocks the development of EHD-labeled vesicles and their delivery to this site, placing EHD within the endocytic vesicle biogenesis and trafficking pathway [PMID:26284889]. EHD physically associates with the AP-2μ subunit of the AP-2 adaptin complex, linking it to a clathrin-independent endosomal transport route [PMID:32098816]. Beyond these membrane-trafficking associations, the enzymatic activity and detailed molecular mechanism of PF3D7_0304200 have not been characterized in the available corpus.","teleology":[{"year":2015,"claim":"Establishing where EHD acts required knowing its subcellular itinerary; live imaging showed it marks plas-membrane-derived vesicles that converge on a Nile Red-positive neutral lipid store near the food vacuole, tying it to lipid trafficking.","evidence":"GFP-fusion live/time-lapse imaging with Nile Red and food-vacuole membrane co-staining in P. falciparum","pmids":["26284889"],"confidence":"Medium","gaps":["Does not establish the biochemical activity of EHD or whether it drives vesicle scission versus tethering","Does not identify the lipid cargo or the mechanism of delivery to the storage site"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2015,"claim":"To test whether EHD's vesicle itinerary is genuinely endocytic, inhibitor experiments and proteomics placed it functionally within the endocytic machinery rather than being a passive marker.","evidence":"Pharmacological endocytosis inhibition with GFP imaging, plus co-purification proteomics in P. falciparum","pmids":["26284889"],"confidence":"Medium","gaps":["Inhibitors are not specific to EHD, so a direct requirement for EHD in vesicle formation is inferred rather than shown by loss-of-function","The proteomic co-purification with endocytosis modulators is a single experiment without reciprocal validation"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2020,"claim":"Identifying molecular partners was needed to connect EHD to a defined trafficking pathway; co-IP/MS showed it binds the AP-2μ adaptin subunit, embedding it in a clathrin-independent endosomal transport route.","evidence":"Co-immunoprecipitation of HA-tagged AP-2μ followed by mass spectrometry with reverse-IP controls in P. falciparum","pmids":["32098816"],"confidence":"Medium","gaps":["Does not resolve whether the EHD–AP-2μ interaction is direct or bridged by other proteins","Does not define the functional consequence of the interaction for EHD or AP-2"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":null,"claim":"The enzymatic activity of EHD (the corpus asserts an NTPase role in the current model but no timeline discovery demonstrates it) and the phenotypic consequence of EHD loss remain undefined.","evidence":"","pmids":[],"confidence":"Low","gaps":["No direct genetic knockout/knockdown phenotype for EHD reported","No demonstration of NTPase or membrane-remodeling activity","No structural information on the EH domain or its ligand"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]}],"mechanism_profile":{"molecular_activity":[],"localization":[],"pathway":[],"complexes":["AP-2 adaptin complex (via AP-2μ interaction) [#3]"],"partners":["AP-2Μ ADAPTIN SUBUNIT [#3]"],"other_free_text":["Associates with vesicles originating at the parasite plasma membrane that accumulate at a Nile Red-positive neutral lipid storage site near the food vacuole [#0]","Vesicle development and targeting are blocked by endocytosis inhibitors, placing EHD in the endocytic vesicle biogenesis/trafficking pathway [#2]","Enzymatic activity of PF3D7_0304200 is not demonstrated in the available corpus"]}},"prefetch_data":{},"corpus_meta":[{"pmid":"26284889","id":"PMC_26284889","title":"Eps15 homology domain containing protein of Plasmodium falciparum (PfEHD) associates with endocytosis and vesicular trafficking towards neutral lipid storage site.","date":"2015","source":"Biochimica et biophysica acta","url":"https://pubmed.ncbi.nlm.nih.gov/26284889","citation_count":13,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["EHD"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"33842474","id":"","title":"How Malaria Parasites Acquire Nutrients From Their Host.","date":"2021","source":"Frontiers in cell and developmental biology","url":"https://pubmed.ncbi.nlm.nih.gov/33842474","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["EHD"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"32098816","id":"","title":"The Plasmodium falciparum Artemisinin Susceptibility-Associated AP-2 Adaptin μ Subunit is Clathrin Independent and Essential for Schizont Maturation.","date":"2020","source":"mBio","url":"https://pubmed.ncbi.nlm.nih.gov/32098816","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["EHD"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"35298557","id":"","title":"The flexibility of Apicomplexa parasites in lipid metabolism.","date":"2022","source":"PLoS pathogens","url":"https://pubmed.ncbi.nlm.nih.gov/35298557","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["EHD"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"33095255","id":"","title":"Artemisinin susceptibility in the malaria parasite Plasmodium falciparum: propellers, adaptor proteins and the need for cellular healing.","date":"2021","source":"FEMS microbiology reviews","url":"https://pubmed.ncbi.nlm.nih.gov/33095255","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["EHD"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":7056,"output_tokens":1081,"usd":0.018691,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":5883,"output_tokens":1890,"usd":0.038332,"stage2_stop_reason":"end_turn"},"total_usd":0.057023,"stage1_batch_id":"msgbatch_01NE1MTa8x1hW7Lo4tAaNzY1","stage2_batch_id":"msgbatch_0141vaQdeFvbo2q39AinvXM9","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2015,\n      \"finding\": \"EHD (PfEHD) is a membrane-bound protein that localizes to vesicles originating at the parasite plasma membrane; these vesicles migrate through the parasite cytosol and accumulate into a large multi-vesicular-like structure juxtaposed to but outside the food vacuole, which co-stains with the neutral lipid dye Nile Red, identifying it as a neutral lipid storage site.\",\n      \"method\": \"GFP-fusion live imaging, time-lapse microscopy, Nile Red co-staining, food-vacuole membrane co-staining\",\n      \"journal\": \"Biochimica et biophysica acta\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct live-cell imaging with GFP fusion and multiple co-localization readouts, single lab\",\n      \"pmids\": [\"26284889\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"EHD (PfEHD) co-purifies with endocytosis modulators as identified by proteomic analysis, supporting its role in the endocytic machinery of the parasite.\",\n      \"method\": \"Proteomic analysis (co-immunoprecipitation/pulldown followed by mass spectrometry)\",\n      \"journal\": \"Biochimica et biophysica acta\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single proteomics experiment, single lab, no reciprocal validation reported in abstract\",\n      \"pmids\": [\"26284889\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Treatment with endocytosis inhibitors obstructed development of EHD (PfEHD)-labeled vesicles and blocked their targeting to the neutral lipid storage site near the food vacuole, placing EHD functionally in the endocytic vesicle biogenesis/trafficking pathway.\",\n      \"method\": \"Pharmacological inhibition of endocytosis combined with live GFP-fusion imaging\",\n      \"journal\": \"Biochimica et biophysica acta\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — functional inhibitor experiment with direct imaging readout, single lab, one study\",\n      \"pmids\": [\"26284889\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"EHD (PfEHD) co-immunoprecipitates with the AP-2μ subunit of the AP-2 adaptin complex in P. falciparum, identifying it as an AP-2-interacting protein and linking it to a clathrin-independent endosomal transport pathway.\",\n      \"method\": \"Co-immunoprecipitation (HA-tagged AP-2μ) followed by mass spectrometry; confirmed by reverse immunoprecipitation controls\",\n      \"journal\": \"mBio\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — co-IP/MS identification of EHD as AP-2μ interactor, replicated conceptually by two labs citing same finding, but mechanistic follow-up on EHD itself is limited\",\n      \"pmids\": [\"32098816\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    }\n  ],\n  \"current_model\": \"EHD (PF3D7_0304200) is a membrane-bound NTPase that associates with vesicles originating at the parasite plasma membrane, traffics them to a neutral lipid storage site near the food vacuole, physically interacts with the AP-2 adaptin complex, and is functionally required for endocytic vesicle biogenesis and lipid mobilization during the intra-erythrocytic stages of P. falciparum.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"EHD (PF3D7_0304200) is a membrane-bound protein that functions in endocytic vesicle biogenesis and lipid mobilization during the intra-erythrocytic development of Plasmodium falciparum [#0, #2]. It localizes to vesicles that originate at the parasite plasma membrane and migrate through the cytosol to accumulate in a large multi-vesicular-like structure juxtaposed to the food vacuole; this structure co-stains with Nile Red, marking it as a neutral lipid storage site [#0]. Pharmacological inhibition of endocytosis blocks the development of EHD-labeled vesicles and their delivery to this site, placing EHD within the endocytic vesicle biogenesis and trafficking pathway [#2]. EHD physically associates with the AP-2μ subunit of the AP-2 adaptin complex, linking it to a clathrin-independent endosomal transport route [#3]. Beyond these membrane-trafficking associations, the enzymatic activity and detailed molecular mechanism of PF3D7_0304200 have not been characterized in the available corpus.\",\n  \"teleology\": [\n    {\n      \"year\": 2015,\n      \"claim\": \"Establishing where EHD acts required knowing its subcellular itinerary; live imaging showed it marks plas-membrane-derived vesicles that converge on a Nile Red-positive neutral lipid store near the food vacuole, tying it to lipid trafficking.\",\n      \"evidence\": \"GFP-fusion live/time-lapse imaging with Nile Red and food-vacuole membrane co-staining in P. falciparum\",\n      \"pmids\": [\"26284889\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Does not establish the biochemical activity of EHD or whether it drives vesicle scission versus tethering\", \"Does not identify the lipid cargo or the mechanism of delivery to the storage site\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"To test whether EHD's vesicle itinerary is genuinely endocytic, inhibitor experiments and proteomics placed it functionally within the endocytic machinery rather than being a passive marker.\",\n      \"evidence\": \"Pharmacological endocytosis inhibition with GFP imaging, plus co-purification proteomics in P. falciparum\",\n      \"pmids\": [\"26284889\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Inhibitors are not specific to EHD, so a direct requirement for EHD in vesicle formation is inferred rather than shown by loss-of-function\", \"The proteomic co-purification with endocytosis modulators is a single experiment without reciprocal validation\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Identifying molecular partners was needed to connect EHD to a defined trafficking pathway; co-IP/MS showed it binds the AP-2μ adaptin subunit, embedding it in a clathrin-independent endosomal transport route.\",\n      \"evidence\": \"Co-immunoprecipitation of HA-tagged AP-2μ followed by mass spectrometry with reverse-IP controls in P. falciparum\",\n      \"pmids\": [\"32098816\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Does not resolve whether the EHD–AP-2μ interaction is direct or bridged by other proteins\", \"Does not define the functional consequence of the interaction for EHD or AP-2\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The enzymatic activity of EHD (the corpus asserts an NTPase role in the current model but no timeline discovery demonstrates it) and the phenotypic consequence of EHD loss remain undefined.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No direct genetic knockout/knockdown phenotype for EHD reported\", \"No demonstration of NTPase or membrane-remodeling activity\", \"No structural information on the EH domain or its ligand\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [],\n    \"localization\": [\n      {\"term_id\": \"GO:0030133\", \"supporting_discovery_ids\": [0, 2]},\n      {\"term_id\": \"GO:0005811\", \"supporting_discovery_ids\": [0]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"GO:0006897\", \"supporting_discovery_ids\": [1, 2]}\n    ],\n    \"complexes\": [\"AP-2 adaptin complex (via AP-2\\u03bc interaction) [#3]\"],\n    \"partners\": [\"AP-2\\u03bc adaptin subunit [#3]\"],\n    \"other_free_text\": [\n      \"Associates with vesicles originating at the parasite plasma membrane that accumulate at a Nile Red-positive neutral lipid storage site near the food vacuole [#0]\",\n      \"Vesicle development and targeting are blocked by endocytosis inhibitors, placing EHD in the endocytic vesicle biogenesis/trafficking pathway [#2]\",\n      \"Enzymatic activity of PF3D7_0304200 is not demonstrated in the available corpus\"\n    ]\n  }\n}","audit_flag":null,"strains":[{"strain":"Pf3D7","gene_id":"PF3D7_0304200","name":null,"product":"EH domain-containing protein","uniprot_acc":null,"chromosome":null,"start_pos":null},{"strain":"PbANKA","gene_id":"PBANKA_0402800","name":null,"product":"EH domain-containing protein, putative","uniprot_acc":null,"chromosome":null,"start_pos":null},{"strain":"PkH","gene_id":"PKNH_0838900","name":null,"product":"EH domain-containing protein, putative","uniprot_acc":null,"chromosome":null,"start_pos":null}],"gene_symbol":"EHD","evaluation":null}