{"gene":"PF3D7_1025000","run_date":"2026-09-08T19:44:17","timeline":{"discoveries":[{"year":2015,"finding":"EPS15 (PfEHD / PF3D7_1025000) is a membrane-bound protein that associates with a vesicular network in P. falciparum. GFP-tagging showed vesicles originating at the parasite plasma membrane, migrating through the cytosol, and culminating in a large multi-vesicular-like structure juxtaposed to but outside the food vacuole. This structure co-stained with the neutral-lipid dye Nile Red, identifying it as a neutral lipid storage site. Proteomic analysis identified endocytosis modulators as EPS15-associated proteins. Treatment with endocytosis inhibitors blocked vesicle development and their targeting to this lipid storage site, demonstrating a role in endocytic vesicle generation at the parasite plasma membrane.","method":"GFP-fusion live-cell imaging (time-lapse microscopy), Nile Red neutral-lipid staining, co-staining with food vacuole membrane marker, proteomic (mass spectrometry) identification of associated proteins, pharmacological inhibition of endocytosis","journal":"Biochimica et biophysica acta","confidence":"Medium","confidence_rationale":"Tier 2/3 / Moderate — live-cell imaging with functional consequence (vesicle formation blocked by inhibitors), proteomic interaction data, single lab, multiple orthogonal methods","pmids":["26284889"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2020,"finding":"EPS15 (PF3D7_1025000) was identified as a component of the K13 compartment in P. falciparum by dimerization-induced proximity biotinylation (DiQ-BioID). EPS15 co-localised with K13 at a clathrin-independent, AP-2-labelled endocytic compartment (cytostome-associated). Reciprocal DiQ-BioID using EPS15 as bait also identified K13, confirming the interaction. Conditional inactivation of EPS15 (along with UBP1 and AP-2μ) reduced hemoglobin uptake to the digestive vacuole at both ring and trophozoite stages and conferred ring-stage artemisinin tolerance, demonstrating that EPS15 is required for host-cell-cytosol endocytosis throughout asexual blood stages.","method":"DiQ-BioID proximity biotinylation (reciprocal), live-cell fluorescence imaging/co-localization, conditional protein inactivation (knock-sideways or analogous), fluorescent dextran uptake assay, ring-stage survival assay (RSA) for artemisinin susceptibility","journal":"PLoS pathogens (cited as Birnbaum et al. 2020 throughout corpus)","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal proximity-biotinylation, conditional loss-of-function with defined cellular phenotype (reduced hemoglobin uptake + ART resistance), independently cited and replicated across multiple subsequent labs","pmids":["38039338","33842474","32310999","34765267","33095255"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2023,"finding":"EPS15 (PF3D7_1025000) was detected (at low abundance) in GFP-trap pulldowns of clathrin light chain (PfCLC) and AP-1 γ subunit in P. falciparum, overlapping with the K13 interactome. This suggests EPS15 may be loosely or indirectly associated with the clathrin-AP-1 complex, though it is not a core component of it.","method":"GFP-trap pulldown followed by mass spectrometry (interactome analysis)","journal":"Traffic (Copenhagen, Denmark)","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single co-IP/pulldown experiment, EPS15 detected at low abundance, authors themselves note the association is loose or indirect","pmids":["41451970"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2024,"finding":"Testing 135 mutations in K13 compartment proteins (including multiple mutations in EPS15/PF3D7_1025000) introduced into 3D7 parasites via selection-linked integration showed that none of the EPS15 mutations tested conferred artemisinin resistance in the ring-stage survival assay, indicating that the specific field-derived variants of EPS15 tested do not independently reduce ART susceptibility.","method":"CRISPR/selection-linked integration of recodonized multi-mutant gene, ring-stage survival assay (RSA) with dihydroartemisinin","journal":"mBio","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — isogenic parasite engineering with RSA phenotyping; NEGATIVE result that is mechanistically informative; single lab","pmids":["38700363"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":"3D7"}]}],"current_model":"PF3D7_1025000 (EPS15) is a membrane-associated endocytic protein in P. falciparum that localises to a clathrin-independent, AP-2-labelled cytostome-proximal compartment (the K13 compartment) where it physically interacts with K13; conditional inactivation of EPS15 reduces host-cell-cytosol (hemoglobin) uptake at both ring and trophozoite stages and confers ring-stage artemisinin tolerance, while GFP-based imaging shows it also participates in vesicle trafficking toward a neutral-lipid storage site near the food vacuole."},"narrative":{"mechanistic_narrative":"PF3D7_1025000 (EPS15) is a membrane-associated endocytic protein of Plasmodium falciparum that mediates uptake of host-cell cytosol (hemoglobin) at the parasite plasma membrane throughout the asexual blood stages [PMID:26284889, PMID:38039338, PMID:33842474, PMID:32310999, PMID:34765267, PMID:33095255]. Live-cell imaging of GFP-tagged EPS15 traces vesicles that originate at the parasite plasma membrane, migrate through the cytosol, and terminate at a Nile Red-positive neutral-lipid storage structure juxtaposed to the food vacuole; pharmacological inhibition of endocytosis blocks both vesicle development and their delivery to this site, establishing EPS15 in endocytic vesicle generation [PMID:26284889]. EPS15 is a component of the clathrin-independent, AP-2-labelled cytostome-proximal 'K13 compartment', where reciprocal proximity biotinylation shows it physically associates with K13, and its conditional inactivation reduces hemoglobin uptake to the digestive vacuole at both ring and trophozoite stages while conferring ring-stage artemisinin tolerance [PMID:38039338, PMID:33842474, PMID:32310999, PMID:34765267, PMID:33095255]. Beyond this endocytic/host-cytosol-uptake role, no further biochemical mechanism (e.g. defined enzymatic activity or direct structural role) for EPS15 has been characterized in the available corpus.","teleology":[{"year":2015,"claim":"It was unknown whether PF3D7_1025000 participated in membrane trafficking; GFP imaging established it as a membrane-bound protein driving endocytic vesicles from the parasite plasma membrane to a neutral-lipid storage site near the food vacuole.","evidence":"GFP-fusion time-lapse imaging, Nile Red staining, food-vacuole marker co-staining, associated-protein proteomics, and pharmacological endocytosis inhibition in P. falciparum","pmids":["26284889"],"confidence":"Medium","gaps":["Molecular partners driving vesicle scission or transport not defined beyond generic 'endocytosis modulators'","Direct biochemical activity of the protein not established","Relationship of the lipid-storage endpoint to hemoglobin catabolism not resolved"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2020,"claim":"The identity of the endocytic machinery underlying host-cell-cytosol uptake and artemisinin response was unclear; reciprocal proximity biotinylation and conditional inactivation placed EPS15 in the AP-2-labelled K13 compartment as a K13 interactor required for hemoglobin uptake and ring-stage artemisinin susceptibility.","evidence":"Reciprocal DiQ-BioID, co-localization imaging, conditional inactivation, dextran uptake assay, and ring-stage survival assay in P. falciparum","pmids":["38039338","33842474","32310999","34765267","33095255"],"confidence":"High","gaps":["Whether the EPS15–K13 interaction is direct or bridged by other compartment proteins is not resolved","Structural basis of EPS15 recruitment to the cytostome-proximal compartment unknown","Mechanism linking reduced endocytosis to artemisinin tolerance not defined at the molecular level"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2023,"claim":"It was unclear whether EPS15 associates with the clathrin/AP-1 trafficking machinery; GFP-trap interactome pulldowns detected EPS15 at low abundance with PfCLC and AP-1, indicating at most a loose or indirect link rather than core membership.","evidence":"GFP-trap pulldown with mass spectrometry of PfCLC and AP-1 γ subunit in P. falciparum","pmids":["41451970"],"confidence":"Low","gaps":["Single low-abundance pulldown without reciprocal or functional validation of the association","Cannot distinguish direct interaction from co-compartmentalization","Functional significance of any clathrin/AP-1 link untested"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2024,"claim":"Because EPS15 loss confers artemisinin tolerance, it was important to test whether field-derived EPS15 sequence variants drive resistance; engineered isogenic parasites showed none of the EPS15 mutations tested independently reduced artemisinin susceptibility.","evidence":"CRISPR/selection-linked integration of recodonized multi-mutant EPS15 into 3D7 and ring-stage survival assay with dihydroartemisinin","pmids":["38700363"],"confidence":"Medium","gaps":["Only specific tested variants assessed; other mutations or combinations not excluded","Does not address whether EPS15 expression-level changes affect resistance","Single lab, single genetic background (3D7)"],"attribution":[{"species":"Plasmodium falciparum","strain":"3D7"}]},{"year":null,"claim":"The direct biochemical activity of EPS15, its precise molecular role within the K13/AP-2 compartment, and the mechanistic link between its endocytic function and artemisinin tolerance remain undefined.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No defined enzymatic or structural function for the protein","Whether EPS15–K13 binding is direct is unresolved","Mechanism connecting reduced endocytosis to drug tolerance not established"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]}],"mechanism_profile":{"molecular_activity":[],"localization":[],"pathway":[],"complexes":["K13 compartment (AP-2-labelled, clathrin-independent cytostome-proximal endocytic compartment)"],"partners":["K13 (KELCH13)","AP-2 (AP-2Μ)","UBP1","PFCLC (CLATHRIN LIGHT CHAIN) — LOW-ABUNDANCE/INDIRECT","AP-1 Γ SUBUNIT — LOW-ABUNDANCE/INDIRECT"],"other_free_text":["Conditional inactivation reduces hemoglobin (host-cell-cytosol) uptake to the digestive vacuole at ring and trophozoite stages and confers ring-stage artemisinin tolerance [#1]","GFP-tagged protein traffics vesicles from the parasite plasma membrane to a Nile Red-positive neutral-lipid storage structure near the food vacuole [#0]","Tested field-derived EPS15 mutations did not independently confer artemisinin resistance in 3D7 [#3]"]}},"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":["EPS15","Eps15"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"10404267","id":"PMC_10404267","title":"Expression and characterisation of a Plasmodium falciparum protein containing domains homologous to sarcalumenin and a tyrosine kinase substrate, eps15.","date":"1999","source":"International journal for parasitology","url":"https://pubmed.ncbi.nlm.nih.gov/10404267","citation_count":4,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["EPS15","Eps15"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"42266499","id":"","title":"Hemoglobin Digestion Genes Are Conserved in Lizard-Infective Plasmodium Species With Different Host Cellular Niches.","date":"2026","source":"Ecology and evolution","url":"https://pubmed.ncbi.nlm.nih.gov/42266499","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["EPS15"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"42001110","id":"","title":"Targeting protein-protein interactions in Plasmodium: from asexual replication to sexual development.","date":"2026","source":"Parasites & vectors","url":"https://pubmed.ncbi.nlm.nih.gov/42001110","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["EPS15"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"41662857","id":"","title":"Parasite clearance in patients with Plasmodium vivax monoinfection treated with artesunate in Cambodia: an observational secondary analysis of trial data.","date":"2026","source":"The Lancet. 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microbiology","url":"https://pubmed.ncbi.nlm.nih.gov/35096663","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["EPS15"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"34090067","id":"","title":"Plasmodium falciparum resistance to ACTs: Emergence, mechanisms, and outlook.","date":"2021","source":"International journal for parasitology. Drugs and drug resistance","url":"https://pubmed.ncbi.nlm.nih.gov/34090067","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["EPS15"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"34102588","id":"","title":"Integration of population and functional genomics to understand mechanisms of artemisinin resistance in Plasmodium falciparum.","date":"2021","source":"International journal for parasitology. Drugs and drug resistance","url":"https://pubmed.ncbi.nlm.nih.gov/34102588","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["EPS15"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"35938722","id":"","title":"The Plasmodium falciparum Nuclear Protein Phosphatase NIF4 Is Required for Efficient Merozoite Invasion and Regulates Artemisinin Sensitivity.","date":"2022","source":"mBio","url":"https://pubmed.ncbi.nlm.nih.gov/35938722","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["EPS15"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"34606334","id":"","title":"Mutation in the Plasmodium falciparum BTB/POZ Domain of K13 Protein Confers Artemisinin Resistance.","date":"2021","source":"Antimicrobial agents and chemotherapy","url":"https://pubmed.ncbi.nlm.nih.gov/34606334","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["EPS15"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"37061511","id":"","title":"Stable endocytic structures navigate the complex pellicle of apicomplexan parasites.","date":"2023","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/37061511","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["EPS15"],"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":["EPS15"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"32310999","id":"","title":"Insights into the intracellular localization, protein associations and artemisinin resistance properties of Plasmodium falciparum K13.","date":"2020","source":"PLoS pathogens","url":"https://pubmed.ncbi.nlm.nih.gov/32310999","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["EPS15"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"34765267","id":"","title":"Protecting future antimalarials from the trap of resistance: Lessons from artemisinin-based combination therapy (ACT) failures.","date":"2020","source":"Journal of pharmaceutical analysis","url":"https://pubmed.ncbi.nlm.nih.gov/34765267","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["EPS15"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":30720,"output_tokens":1580,"usd":0.05793,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":6591,"output_tokens":2725,"usd":0.05054,"stage2_stop_reason":"end_turn"},"total_usd":0.10847,"stage1_batch_id":"msgbatch_01SRp5hyRc82zNjddxyCCXGk","stage2_batch_id":"msgbatch_01K3BTmBazRo3pRrMvyaPMTY","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2015,\n      \"finding\": \"EPS15 (PfEHD / PF3D7_1025000) is a membrane-bound protein that associates with a vesicular network in P. falciparum. GFP-tagging showed vesicles originating at the parasite plasma membrane, migrating through the cytosol, and culminating in a large multi-vesicular-like structure juxtaposed to but outside the food vacuole. This structure co-stained with the neutral-lipid dye Nile Red, identifying it as a neutral lipid storage site. Proteomic analysis identified endocytosis modulators as EPS15-associated proteins. Treatment with endocytosis inhibitors blocked vesicle development and their targeting to this lipid storage site, demonstrating a role in endocytic vesicle generation at the parasite plasma membrane.\",\n      \"method\": \"GFP-fusion live-cell imaging (time-lapse microscopy), Nile Red neutral-lipid staining, co-staining with food vacuole membrane marker, proteomic (mass spectrometry) identification of associated proteins, pharmacological inhibition of endocytosis\",\n      \"journal\": \"Biochimica et biophysica acta\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2/3 / Moderate — live-cell imaging with functional consequence (vesicle formation blocked by inhibitors), proteomic interaction data, single lab, multiple orthogonal methods\",\n      \"pmids\": [\"26284889\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"EPS15 (PF3D7_1025000) was identified as a component of the K13 compartment in P. falciparum by dimerization-induced proximity biotinylation (DiQ-BioID). EPS15 co-localised with K13 at a clathrin-independent, AP-2-labelled endocytic compartment (cytostome-associated). Reciprocal DiQ-BioID using EPS15 as bait also identified K13, confirming the interaction. Conditional inactivation of EPS15 (along with UBP1 and AP-2μ) reduced hemoglobin uptake to the digestive vacuole at both ring and trophozoite stages and conferred ring-stage artemisinin tolerance, demonstrating that EPS15 is required for host-cell-cytosol endocytosis throughout asexual blood stages.\",\n      \"method\": \"DiQ-BioID proximity biotinylation (reciprocal), live-cell fluorescence imaging/co-localization, conditional protein inactivation (knock-sideways or analogous), fluorescent dextran uptake assay, ring-stage survival assay (RSA) for artemisinin susceptibility\",\n      \"journal\": \"PLoS pathogens (cited as Birnbaum et al. 2020 throughout corpus)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal proximity-biotinylation, conditional loss-of-function with defined cellular phenotype (reduced hemoglobin uptake + ART resistance), independently cited and replicated across multiple subsequent labs\",\n      \"pmids\": [\"38039338\", \"33842474\", \"32310999\", \"34765267\", \"33095255\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"EPS15 (PF3D7_1025000) was detected (at low abundance) in GFP-trap pulldowns of clathrin light chain (PfCLC) and AP-1 γ subunit in P. falciparum, overlapping with the K13 interactome. This suggests EPS15 may be loosely or indirectly associated with the clathrin-AP-1 complex, though it is not a core component of it.\",\n      \"method\": \"GFP-trap pulldown followed by mass spectrometry (interactome analysis)\",\n      \"journal\": \"Traffic (Copenhagen, Denmark)\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single co-IP/pulldown experiment, EPS15 detected at low abundance, authors themselves note the association is loose or indirect\",\n      \"pmids\": [\"41451970\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Testing 135 mutations in K13 compartment proteins (including multiple mutations in EPS15/PF3D7_1025000) introduced into 3D7 parasites via selection-linked integration showed that none of the EPS15 mutations tested conferred artemisinin resistance in the ring-stage survival assay, indicating that the specific field-derived variants of EPS15 tested do not independently reduce ART susceptibility.\",\n      \"method\": \"CRISPR/selection-linked integration of recodonized multi-mutant gene, ring-stage survival assay (RSA) with dihydroartemisinin\",\n      \"journal\": \"mBio\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — isogenic parasite engineering with RSA phenotyping; NEGATIVE result that is mechanistically informative; single lab\",\n      \"pmids\": [\"38700363\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"3D7\"}]\n    }\n  ],\n  \"current_model\": \"PF3D7_1025000 (EPS15) is a membrane-associated endocytic protein in P. falciparum that localises to a clathrin-independent, AP-2-labelled cytostome-proximal compartment (the K13 compartment) where it physically interacts with K13; conditional inactivation of EPS15 reduces host-cell-cytosol (hemoglobin) uptake at both ring and trophozoite stages and confers ring-stage artemisinin tolerance, while GFP-based imaging shows it also participates in vesicle trafficking toward a neutral-lipid storage site near the food vacuole.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"PF3D7_1025000 (EPS15) is a membrane-associated endocytic protein of Plasmodium falciparum that mediates uptake of host-cell cytosol (hemoglobin) at the parasite plasma membrane throughout the asexual blood stages [#0, #1]. Live-cell imaging of GFP-tagged EPS15 traces vesicles that originate at the parasite plasma membrane, migrate through the cytosol, and terminate at a Nile Red-positive neutral-lipid storage structure juxtaposed to the food vacuole; pharmacological inhibition of endocytosis blocks both vesicle development and their delivery to this site, establishing EPS15 in endocytic vesicle generation [#0]. EPS15 is a component of the clathrin-independent, AP-2-labelled cytostome-proximal 'K13 compartment', where reciprocal proximity biotinylation shows it physically associates with K13, and its conditional inactivation reduces hemoglobin uptake to the digestive vacuole at both ring and trophozoite stages while conferring ring-stage artemisinin tolerance [#1]. Beyond this endocytic/host-cytosol-uptake role, no further biochemical mechanism (e.g. defined enzymatic activity or direct structural role) for EPS15 has been characterized in the available corpus.\",\n  \"teleology\": [\n    {\n      \"year\": 2015,\n      \"claim\": \"It was unknown whether PF3D7_1025000 participated in membrane trafficking; GFP imaging established it as a membrane-bound protein driving endocytic vesicles from the parasite plasma membrane to a neutral-lipid storage site near the food vacuole.\",\n      \"evidence\": \"GFP-fusion time-lapse imaging, Nile Red staining, food-vacuole marker co-staining, associated-protein proteomics, and pharmacological endocytosis inhibition in P. falciparum\",\n      \"pmids\": [\"26284889\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Molecular partners driving vesicle scission or transport not defined beyond generic 'endocytosis modulators'\",\n        \"Direct biochemical activity of the protein not established\",\n        \"Relationship of the lipid-storage endpoint to hemoglobin catabolism not resolved\"\n      ],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"The identity of the endocytic machinery underlying host-cell-cytosol uptake and artemisinin response was unclear; reciprocal proximity biotinylation and conditional inactivation placed EPS15 in the AP-2-labelled K13 compartment as a K13 interactor required for hemoglobin uptake and ring-stage artemisinin susceptibility.\",\n      \"evidence\": \"Reciprocal DiQ-BioID, co-localization imaging, conditional inactivation, dextran uptake assay, and ring-stage survival assay in P. falciparum\",\n      \"pmids\": [\"38039338\", \"33842474\", \"32310999\", \"34765267\", \"33095255\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Whether the EPS15–K13 interaction is direct or bridged by other compartment proteins is not resolved\",\n        \"Structural basis of EPS15 recruitment to the cytostome-proximal compartment unknown\",\n        \"Mechanism linking reduced endocytosis to artemisinin tolerance not defined at the molecular level\"\n      ],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"It was unclear whether EPS15 associates with the clathrin/AP-1 trafficking machinery; GFP-trap interactome pulldowns detected EPS15 at low abundance with PfCLC and AP-1, indicating at most a loose or indirect link rather than core membership.\",\n      \"evidence\": \"GFP-trap pulldown with mass spectrometry of PfCLC and AP-1 γ subunit in P. falciparum\",\n      \"pmids\": [\"41451970\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\n        \"Single low-abundance pulldown without reciprocal or functional validation of the association\",\n        \"Cannot distinguish direct interaction from co-compartmentalization\",\n        \"Functional significance of any clathrin/AP-1 link untested\"\n      ],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Because EPS15 loss confers artemisinin tolerance, it was important to test whether field-derived EPS15 sequence variants drive resistance; engineered isogenic parasites showed none of the EPS15 mutations tested independently reduced artemisinin susceptibility.\",\n      \"evidence\": \"CRISPR/selection-linked integration of recodonized multi-mutant EPS15 into 3D7 and ring-stage survival assay with dihydroartemisinin\",\n      \"pmids\": [\"38700363\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Only specific tested variants assessed; other mutations or combinations not excluded\",\n        \"Does not address whether EPS15 expression-level changes affect resistance\",\n        \"Single lab, single genetic background (3D7)\"\n      ],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"3D7\"}]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The direct biochemical activity of EPS15, its precise molecular role within the K13/AP-2 compartment, and the mechanistic link between its endocytic function and artemisinin tolerance remain undefined.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"No defined enzymatic or structural function for the protein\",\n        \"Whether EPS15–K13 binding is direct is unresolved\",\n        \"Mechanism connecting reduced endocytosis to drug tolerance not established\"\n      ],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0030276\", \"supporting_discovery_ids\": [0, 1]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [0, 1]},\n      {\"term_id\": \"GO:0030665\", \"supporting_discovery_ids\": [1]},\n      {\"term_id\": \"GO:0005811\", \"supporting_discovery_ids\": [0]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"GO:0006897\", \"supporting_discovery_ids\": [0, 1]},\n      {\"term_id\": \"GO:0072583\", \"supporting_discovery_ids\": [1]}\n    ],\n    \"complexes\": [\n      \"K13 compartment (AP-2-labelled, clathrin-independent cytostome-proximal endocytic compartment)\"\n    ],\n    \"partners\": [\n      \"K13 (Kelch13)\",\n      \"AP-2 (AP-2\\u03bc)\",\n      \"UBP1\",\n      \"PfCLC (clathrin light chain) — low-abundance/indirect\",\n      \"AP-1 \\u03b3 subunit — low-abundance/indirect\"\n    ],\n    \"other_free_text\": [\n      \"Conditional inactivation reduces hemoglobin (host-cell-cytosol) uptake to the digestive vacuole at ring and trophozoite stages and confers ring-stage artemisinin tolerance [#1]\",\n      \"GFP-tagged protein traffics vesicles from the parasite plasma membrane to a Nile Red-positive neutral-lipid storage structure near the food vacuole [#0]\",\n      \"Tested field-derived EPS15 mutations did not independently confer artemisinin resistance in 3D7 [#3]\"\n    ]\n  }\n}","audit_flag":null,"strains":[{"strain":"Pf3D7","gene_id":"PF3D7_1025000","name":null,"product":"Eps15-like protein","uniprot_acc":null,"chromosome":null,"start_pos":null},{"strain":"PbANKA","gene_id":"PBANKA_0509200","name":null,"product":"Eps15-like protein, putative","uniprot_acc":null,"chromosome":null,"start_pos":null},{"strain":"PkH","gene_id":"PKNH_0609500","name":null,"product":"Eps15-like protein, putative","uniprot_acc":null,"chromosome":null,"start_pos":null}],"gene_symbol":"EPS15","evaluation":null}