{"gene":"PF3D7_1468700","run_date":"2026-09-08T14:19:07","timeline":{"discoveries":[{"year":2009,"finding":"P. falciparum eIF4A (PfeIF4A, annotated as PfH45) physically interacts with PfeIF4G as a component of the eIF4F translation initiation complex; PfeIF4G was shown to contain binding sites for both PfeIF4A and PfeIF4E and to interact with PfPABP, establishing PfeIF4A's role within the canonical eIF4F scaffold.","method":"Biochemical isolation, functional characterization, and demonstration of protein–protein interaction (PfeIF4G interaction with PfeIF4A, PfeIF4E, and PfPABP)","journal":"Parasitology international","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — single-lab pulldown/interaction data with functional annotation; interactions demonstrated but no reciprocal Co-IP or structural validation reported in abstract","pmids":["19751846"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2010,"finding":"P. falciparum eIF4A (PfeIF4A) and eIF4E (PfeIF4E) co-localize and are predominantly found in the cytoplasm of intraerythrocytic-stage parasites; co-treatment with dsRNA targeting both PfeIF4A and PfeIF4E caused ~67% growth inhibition, greater than either alone, confirming PfeIF4A's essential role in parasite proliferation.","method":"Immunofluorescence co-localization; dsRNA knockdown (RNAi-like) of PfeIF4A and PfeIF4E individually and in combination with parasite growth assay","journal":"Communicative & integrative biology","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — direct localization experiment and loss-of-function growth phenotype, single lab, two orthogonal methods (IF + dsRNA inhibition)","pmids":["21331256"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2018,"finding":"The rocaglate CR-1-31B perturbs the association of P. falciparum eIF4A (PfeIF4A) with RNA, blocks protein synthesis initiation, and shows potent antiplasmodial activity in vitro against P. falciparum (including drug-resistant isolates) and in vivo in P. berghei and P. chabaudi mouse models.","method":"RNA-binding perturbation assay with rocaglate CR-1-31B; ex vivo P. falciparum replication assay; in vivo mouse infection models (P. berghei cerebral malaria, P. chabaudi blood-stage)","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct biochemical demonstration of eIF4A:RNA disruption plus in vivo validation across two Plasmodium species; single lab","pmids":["29463745"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""},{"species":"Plasmodium berghei","strain":""},{"species":"Plasmodium chabaudi","strain":""}]},{"year":2023,"finding":"In vitro thermal shift assays with recombinantly expressed P. falciparum eIF4A confirmed sensitivity to the rocaglate silvestrol, consistent with in silico docking of the eIF4A:RNA:rocaglate complex predicting low binding energies; this validates the six critical amino acid residues in eIF4A as determinants of rocaglate binding.","method":"In vitro thermal shift assay with recombinant PfeIF4A; molecular docking of eIF4A:RNA:rocaglate complexes; in vitro antiplasmodial testing with silvestrol","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — biochemical thermal shift assay plus molecular docking, validated by in vitro testing; single lab, two complementary methods","pmids":["37291191"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2008,"finding":"P. falciparum eIF4A is represented by multiple protein isoforms in schizont-stage parasites, detected as distinct spots by 2D-DIGE and identified by mass spectrometry, with different isoforms showing distinct temporal abundance profiles during intraerythrocytic development, suggesting post-translational modifications generate functionally distinct eIF4A forms.","method":"Two-dimensional differential gel electrophoresis (2D-DIGE) with mass spectrometry identification; quantitative time-course proteomic profiling of synchronized parasites","journal":"Genome biology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — mass spectrometry identification of isoforms is consistent with PTMs but specific modifications not characterized; single study, indirect inference of PTM from spot pattern","pmids":["19091060"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2021,"finding":"P. falciparum eIF4A was detected in proteomic analysis of proteins enriched on a 5' cap affinity matrix from asexual-stage parasite lysates, consistent with its membership in the eIF4F cap-associated complex, though with low reproducibility.","method":"5' cap affinity matrix pulldown followed by proteomic (LC-MS/MS) analysis of P. falciparum asexual stage lysates","journal":"Molecular and biochemical parasitology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single affinity-enrichment proteomics experiment with low reproducibility for eIF4A; no functional validation reported","pmids":["34890716"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2012,"finding":"Purified recombinant P. falciparum eIF4A (PfeIF4A) exhibits bipolar DNA helicase activity with a preference for the 3'-to-5' direction (2-fold higher rate than 5'-to-3'), is Mg2+-dependent, optimally active at 37°C, and has basal ATP turnover that doubles in the presence of single- or double-stranded DNA; the enzyme does not unwind blunt-ended duplexes.","method":"Helicase unwinding assays with purified recombinant codon-optimized PfeIF4A; ATPase activity measurement; substrate specificity tests with various DNA constructs","journal":"Protein expression and purification","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — in vitro enzymatic characterization of purified recombinant protein with multiple substrate and condition tests; single lab","pmids":["22750398"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]}],"current_model":"P. falciparum eIF4A (PF3D7_1468700) is a cytoplasmic DEAD-box RNA helicase that forms part of the eIF4F translation initiation complex by interacting with eIF4G and co-localizing with eIF4E; it exhibits ATP-dependent bipolar nucleic acid unwinding activity (with a preference for the 3'-to-5' direction), is subject to post-translational modifications generating multiple isoforms during intraerythrocytic development, is essential for parasite growth, and is directly targeted by rocaglate compounds (e.g., CR-1-31B, silvestrol) that disrupt its RNA association and block translation initiation."},"narrative":{"mechanistic_narrative":"PF3D7_1468700 (eIF4A) is a cytoplasmic DEAD-box ATP-dependent RNA/DNA helicase that serves as the helicase component of the P. falciparum eIF4F cap-binding translation initiation complex [PMID:19751846, PMID:21331256]. It is incorporated into eIF4F through direct interaction with eIF4G, which independently binds eIF4E and PABP to assemble the initiation scaffold, and eIF4A co-localizes with eIF4E in the cytoplasm of intraerythrocytic-stage parasites [PMID:19751846, PMID:21331256]. Purified recombinant eIF4A displays Mg2+-dependent, bipolar nucleic acid unwinding activity with a ~2-fold preference for the 3'-to-5' direction and basal ATP turnover that is stimulated by single- or double-stranded nucleic acid but does not unwind blunt-ended duplexes [PMID:22750398]. eIF4A is required for parasite proliferation: combined dsRNA knockdown of eIF4A and eIF4E inhibits growth more than either alone [PMID:21331256]. The protein is a validated antimalarial target of rocaglate compounds (CR-1-31B, silvestrol), which perturb its association with RNA and block translation initiation, with defined active in vitro and in vivo antiplasmodial activity across Plasmodium species [PMID:29463745, PMID:37291191]. Beyond its role in cap-dependent initiation and rocaglate targeting, no further mechanistic detail — including the specific post-translational modifications inferred to generate multiple developmental isoforms — has been characterized in the available corpus [PMID:19091060, PMID:34890716].","teleology":[{"year":2008,"claim":"Before functional dissection, it was unknown whether eIF4A exists as a single form or is developmentally modulated; proteomic profiling showed multiple isoforms with distinct temporal abundance, raising the possibility of PTM-generated functional diversity during the erythrocytic cycle.","evidence":"2D-DIGE with mass spectrometry identification and quantitative time-course profiling of synchronized schizont-stage parasites","pmids":["19091060"],"confidence":"Low","gaps":["Specific post-translational modifications not identified","Functional consequence of distinct isoforms not tested","Isoform assignment inferred from spot pattern rather than direct modification mapping"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2009,"claim":"It was unclear how eIF4A integrated into the parasite translation machinery; biochemical interaction data placed it within the canonical eIF4F complex via eIF4G, which also binds eIF4E and PABP.","evidence":"Biochemical isolation and protein-protein interaction mapping of eIF4G binding sites","pmids":["19751846"],"confidence":"Medium","gaps":["No reciprocal Co-IP or structural validation of the eIF4A:eIF4G interface","Stoichiometry and assembly kinetics of eIF4F not determined"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2010,"claim":"The subcellular site of action and essentiality of eIF4A were unresolved; co-localization with eIF4E and a combinatorial knockdown growth defect established it as a cytoplasmic, proliferation-essential factor.","evidence":"Immunofluorescence co-localization and dsRNA knockdown of eIF4A and eIF4E with growth assays","pmids":["21331256"],"confidence":"Medium","gaps":["dsRNA specificity/off-target effects not fully controlled","Single-gene essentiality via genetic knockout not established","Contribution of eIF4A vs eIF4E to the combined phenotype not separable"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2012,"claim":"The intrinsic enzymatic properties of eIF4A were undefined; in vitro assays on purified recombinant protein demonstrated Mg2+-dependent bipolar unwinding with 3'-to-5' preference and nucleic-acid-stimulated ATPase activity.","evidence":"Helicase unwinding, ATPase, and substrate-specificity assays with purified recombinant codon-optimized eIF4A","pmids":["22750398"],"confidence":"Medium","gaps":["Assays performed on DNA substrates rather than mRNA in a translation context","No structural basis for directional preference","Physiological substrate not identified"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2018,"claim":"Whether eIF4A was a druggable antimalarial target was untested; the rocaglate CR-1-31B was shown to disrupt eIF4A:RNA association and block initiation with potent in vitro and in vivo antiplasmodial activity.","evidence":"RNA-binding perturbation assay, ex vivo P. falciparum replication, and in vivo P. berghei and P. chabaudi mouse models","pmids":["29463745"],"confidence":"Medium","gaps":["Direct binding to eIF4A vs an eIF4A-containing complex not fully separated in vivo","Selectivity over host eIF4A not quantified","Resistance mechanisms not mapped"],"attribution":[{"species":"Plasmodium falciparum","strain":""},{"species":"Plasmodium berghei","strain":""},{"species":"Plasmodium chabaudi","strain":""}]},{"year":2021,"claim":"Whether eIF4A associates with the mRNA 5' cap complex biochemically was addressed; cap affinity proteomics detected eIF4A, consistent with eIF4F membership but with low reproducibility.","evidence":"5' cap affinity matrix pulldown with LC-MS/MS of asexual-stage lysates","pmids":["34890716"],"confidence":"Low","gaps":["Low reproducibility of eIF4A detection","No functional validation of cap association","Indirect vs direct cap-complex recruitment not resolved"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2023,"claim":"The molecular determinants of rocaglate sensitivity were unconfirmed; thermal shift and docking validated silvestrol engagement and six critical eIF4A residues as binding determinants.","evidence":"In vitro thermal shift assay with recombinant eIF4A, molecular docking of eIF4A:RNA:rocaglate complexes, and in vitro antiplasmodial testing","pmids":["37291191"],"confidence":"Medium","gaps":["Predicted binding residues not validated by mutagenesis","No experimental co-crystal structure of the eIF4A:RNA:rocaglate complex","Docking energies are computational predictions"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":null,"claim":"The specific post-translational modifications generating eIF4A isoforms and the physiological mRNA targets whose translation depends on eIF4A helicase activity remain unresolved.","evidence":"","pmids":[],"confidence":"Low","gaps":["No modification mapping of the observed isoforms","No genome-wide identification of eIF4A-dependent transcripts","No high-resolution structure of parasite eIF4A alone or in eIF4F"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]}],"mechanism_profile":{"molecular_activity":[],"localization":[],"pathway":[],"complexes":["eIF4F cap-binding translation initiation complex"],"partners":["EIF4G (PFEIF4G)","EIF4E (PFEIF4E)"],"other_free_text":["Essential for P. falciparum intraerythrocytic proliferation (combined eIF4A/eIF4E dsRNA knockdown gives ~67% growth inhibition) [#1]","Validated antimalarial drug target of rocaglates CR-1-31B and silvestrol, which disrupt eIF4A:RNA association [#2, #3]","Six eIF4A residues predicted as rocaglate-binding determinants by docking and thermal shift [#3]","Represented by multiple protein isoforms across intraerythrocytic development, inferred to arise from post-translational modification [#4]"]}},"prefetch_data":{},"corpus_meta":[{"pmid":"19091060","id":"PMC_19091060","title":"Quantitative protein expression profiling reveals extensive post-transcriptional regulation and post-translational modifications in schizont-stage malaria parasites.","date":"2008","source":"Genome biology","url":"https://pubmed.ncbi.nlm.nih.gov/19091060","citation_count":99,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["eIF4A"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"17822710","id":"PMC_17822710","title":"Bipolar, Dual Plasmodium falciparum helicase 45 expressed in the intraerythrocytic developmental cycle is required for parasite growth.","date":"2007","source":"Journal of molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/17822710","citation_count":47,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["eIF4A"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"29463745","id":"PMC_29463745","title":"Rocaglates as dual-targeting agents for experimental cerebral malaria.","date":"2018","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/29463745","citation_count":30,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["eIF4A","eukaryotic initiation factor 4A","eukaryotic initiation factor 4a","PfeIF4A"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"19641745","id":"PMC_19641745","title":"Identification and bioinformatics characterization of translation initiation complex eIF4F components and poly(A)-binding protein from Plasmodium falciparum.","date":"2009","source":"Communicative & integrative biology","url":"https://pubmed.ncbi.nlm.nih.gov/19641745","citation_count":19,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["eIF4A"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"12745261","id":"PMC_12745261","title":"Replication fork-stimulated eIF-4A from Plasmodium cynomolgi unwinds DNA in the 3' to 5' direction and is inhibited by DNA-interacting compounds.","date":"2003","source":"Archives of biochemistry and biophysics","url":"https://pubmed.ncbi.nlm.nih.gov/12745261","citation_count":17,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["eIF4A"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"19751846","id":"PMC_19751846","title":"Isolation and functional characterization of eIF4F components and poly(A)-binding protein from Plasmodium falciparum.","date":"2009","source":"Parasitology international","url":"https://pubmed.ncbi.nlm.nih.gov/19751846","citation_count":16,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["eIF4A","PfeIF4A"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"12387853","id":"PMC_12387853","title":"Isolation and characterization of an eIF-4A homologue from Plasmodium cynomolgi.","date":"2002","source":"Molecular and biochemical parasitology","url":"https://pubmed.ncbi.nlm.nih.gov/12387853","citation_count":14,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["eIF4A"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"21331256","id":"PMC_21331256","title":"PfeIF4E and PfeIF4A colocalize and their double-stranded RNA inhibits Plasmodium falciparum proliferation.","date":"2010","source":"Communicative & integrative biology","url":"https://pubmed.ncbi.nlm.nih.gov/21331256","citation_count":13,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["eIF4A","eukaryotic initiation factor 4A","eukaryotic initiation factor 4a","PfeIF4A"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"18571876","id":"PMC_18571876","title":"Characterization of replication fork and phosphorylation stimulated Plasmodium falciparum helicase 45.","date":"2008","source":"Gene","url":"https://pubmed.ncbi.nlm.nih.gov/18571876","citation_count":13,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["curated_ref"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"37291191","id":"PMC_37291191","title":"Broad anti-pathogen potential of DEAD box RNA helicase eIF4A-targeting rocaglates.","date":"2023","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/37291191","citation_count":12,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["eIF4A","eukaryotic initiation factor 4A","eukaryotic initiation factor 4a"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"34890716","id":"PMC_34890716","title":"Identification of mRNA 5' cap-associated proteins in the human malaria parasite Plasmodium falciparum.","date":"2021","source":"Molecular and biochemical parasitology","url":"https://pubmed.ncbi.nlm.nih.gov/34890716","citation_count":2,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["eIF4A"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"22750398","id":"PMC_22750398","title":"Enhanced purification and characterization of the PfeIF4A (PfH45) helicase from Plasmodium falciparum using a codon-optimised clone.","date":"2012","source":"Protein expression and purification","url":"https://pubmed.ncbi.nlm.nih.gov/22750398","citation_count":2,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["PfeIF4A"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"38018538","id":"PMC_38018538","title":"Architecture, domain organization, and functional signatures of trypanosomatid keIF4A1s and Plasmodium peIF4A1s suggest conserved functions.","date":"2023","source":"Journal of biosciences","url":"https://pubmed.ncbi.nlm.nih.gov/38018538","citation_count":1,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["eIF4A"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"42615421","id":"PMC_42615421","title":"Poly(A)-binding proteins in trypanosomatids and Plasmodium.","date":"2026","source":"Journal of biosciences","url":"https://pubmed.ncbi.nlm.nih.gov/42615421","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["eIF4A"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":8525,"output_tokens":2166,"usd":0.029032,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":7483,"output_tokens":3437,"usd":0.06167,"stage2_stop_reason":"end_turn"},"total_usd":0.090702,"stage1_batch_id":"msgbatch_011BPjN2BL33F8cJXD1r7V7T","stage2_batch_id":"msgbatch_01Qwg6vP7qxkpL6Gsc7tajsb","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2009,\n      \"finding\": \"P. falciparum eIF4A (PfeIF4A, annotated as PfH45) physically interacts with PfeIF4G as a component of the eIF4F translation initiation complex; PfeIF4G was shown to contain binding sites for both PfeIF4A and PfeIF4E and to interact with PfPABP, establishing PfeIF4A's role within the canonical eIF4F scaffold.\",\n      \"method\": \"Biochemical isolation, functional characterization, and demonstration of protein–protein interaction (PfeIF4G interaction with PfeIF4A, PfeIF4E, and PfPABP)\",\n      \"journal\": \"Parasitology international\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — single-lab pulldown/interaction data with functional annotation; interactions demonstrated but no reciprocal Co-IP or structural validation reported in abstract\",\n      \"pmids\": [\"19751846\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"P. falciparum eIF4A (PfeIF4A) and eIF4E (PfeIF4E) co-localize and are predominantly found in the cytoplasm of intraerythrocytic-stage parasites; co-treatment with dsRNA targeting both PfeIF4A and PfeIF4E caused ~67% growth inhibition, greater than either alone, confirming PfeIF4A's essential role in parasite proliferation.\",\n      \"method\": \"Immunofluorescence co-localization; dsRNA knockdown (RNAi-like) of PfeIF4A and PfeIF4E individually and in combination with parasite growth assay\",\n      \"journal\": \"Communicative & integrative biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — direct localization experiment and loss-of-function growth phenotype, single lab, two orthogonal methods (IF + dsRNA inhibition)\",\n      \"pmids\": [\"21331256\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"The rocaglate CR-1-31B perturbs the association of P. falciparum eIF4A (PfeIF4A) with RNA, blocks protein synthesis initiation, and shows potent antiplasmodial activity in vitro against P. falciparum (including drug-resistant isolates) and in vivo in P. berghei and P. chabaudi mouse models.\",\n      \"method\": \"RNA-binding perturbation assay with rocaglate CR-1-31B; ex vivo P. falciparum replication assay; in vivo mouse infection models (P. berghei cerebral malaria, P. chabaudi blood-stage)\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct biochemical demonstration of eIF4A:RNA disruption plus in vivo validation across two Plasmodium species; single lab\",\n      \"pmids\": [\"29463745\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"},\n        {\"species\": \"Plasmodium berghei\", \"strain\": \"\"},\n        {\"species\": \"Plasmodium chabaudi\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"In vitro thermal shift assays with recombinantly expressed P. falciparum eIF4A confirmed sensitivity to the rocaglate silvestrol, consistent with in silico docking of the eIF4A:RNA:rocaglate complex predicting low binding energies; this validates the six critical amino acid residues in eIF4A as determinants of rocaglate binding.\",\n      \"method\": \"In vitro thermal shift assay with recombinant PfeIF4A; molecular docking of eIF4A:RNA:rocaglate complexes; in vitro antiplasmodial testing with silvestrol\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — biochemical thermal shift assay plus molecular docking, validated by in vitro testing; single lab, two complementary methods\",\n      \"pmids\": [\"37291191\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"P. falciparum eIF4A is represented by multiple protein isoforms in schizont-stage parasites, detected as distinct spots by 2D-DIGE and identified by mass spectrometry, with different isoforms showing distinct temporal abundance profiles during intraerythrocytic development, suggesting post-translational modifications generate functionally distinct eIF4A forms.\",\n      \"method\": \"Two-dimensional differential gel electrophoresis (2D-DIGE) with mass spectrometry identification; quantitative time-course proteomic profiling of synchronized parasites\",\n      \"journal\": \"Genome biology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — mass spectrometry identification of isoforms is consistent with PTMs but specific modifications not characterized; single study, indirect inference of PTM from spot pattern\",\n      \"pmids\": [\"19091060\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"P. falciparum eIF4A was detected in proteomic analysis of proteins enriched on a 5' cap affinity matrix from asexual-stage parasite lysates, consistent with its membership in the eIF4F cap-associated complex, though with low reproducibility.\",\n      \"method\": \"5' cap affinity matrix pulldown followed by proteomic (LC-MS/MS) analysis of P. falciparum asexual stage lysates\",\n      \"journal\": \"Molecular and biochemical parasitology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single affinity-enrichment proteomics experiment with low reproducibility for eIF4A; no functional validation reported\",\n      \"pmids\": [\"34890716\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Purified recombinant P. falciparum eIF4A (PfeIF4A) exhibits bipolar DNA helicase activity with a preference for the 3'-to-5' direction (2-fold higher rate than 5'-to-3'), is Mg2+-dependent, optimally active at 37°C, and has basal ATP turnover that doubles in the presence of single- or double-stranded DNA; the enzyme does not unwind blunt-ended duplexes.\",\n      \"method\": \"Helicase unwinding assays with purified recombinant codon-optimized PfeIF4A; ATPase activity measurement; substrate specificity tests with various DNA constructs\",\n      \"journal\": \"Protein expression and purification\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro enzymatic characterization of purified recombinant protein with multiple substrate and condition tests; single lab\",\n      \"pmids\": [\"22750398\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    }\n  ],\n  \"current_model\": \"P. falciparum eIF4A (PF3D7_1468700) is a cytoplasmic DEAD-box RNA helicase that forms part of the eIF4F translation initiation complex by interacting with eIF4G and co-localizing with eIF4E; it exhibits ATP-dependent bipolar nucleic acid unwinding activity (with a preference for the 3'-to-5' direction), is subject to post-translational modifications generating multiple isoforms during intraerythrocytic development, is essential for parasite growth, and is directly targeted by rocaglate compounds (e.g., CR-1-31B, silvestrol) that disrupt its RNA association and block translation initiation.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"PF3D7_1468700 (eIF4A) is a cytoplasmic DEAD-box ATP-dependent RNA/DNA helicase that serves as the helicase component of the P. falciparum eIF4F cap-binding translation initiation complex [#0, #1]. It is incorporated into eIF4F through direct interaction with eIF4G, which independently binds eIF4E and PABP to assemble the initiation scaffold, and eIF4A co-localizes with eIF4E in the cytoplasm of intraerythrocytic-stage parasites [#0, #1]. Purified recombinant eIF4A displays Mg2+-dependent, bipolar nucleic acid unwinding activity with a ~2-fold preference for the 3'-to-5' direction and basal ATP turnover that is stimulated by single- or double-stranded nucleic acid but does not unwind blunt-ended duplexes [#6]. eIF4A is required for parasite proliferation: combined dsRNA knockdown of eIF4A and eIF4E inhibits growth more than either alone [#1]. The protein is a validated antimalarial target of rocaglate compounds (CR-1-31B, silvestrol), which perturb its association with RNA and block translation initiation, with defined active in vitro and in vivo antiplasmodial activity across Plasmodium species [#2, #3]. Beyond its role in cap-dependent initiation and rocaglate targeting, no further mechanistic detail — including the specific post-translational modifications inferred to generate multiple developmental isoforms — has been characterized in the available corpus [#4, #5].\",\n  \"teleology\": [\n    {\n      \"year\": 2008,\n      \"claim\": \"Before functional dissection, it was unknown whether eIF4A exists as a single form or is developmentally modulated; proteomic profiling showed multiple isoforms with distinct temporal abundance, raising the possibility of PTM-generated functional diversity during the erythrocytic cycle.\",\n      \"evidence\": \"2D-DIGE with mass spectrometry identification and quantitative time-course profiling of synchronized schizont-stage parasites\",\n      \"pmids\": [\"19091060\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Specific post-translational modifications not identified\", \"Functional consequence of distinct isoforms not tested\", \"Isoform assignment inferred from spot pattern rather than direct modification mapping\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"It was unclear how eIF4A integrated into the parasite translation machinery; biochemical interaction data placed it within the canonical eIF4F complex via eIF4G, which also binds eIF4E and PABP.\",\n      \"evidence\": \"Biochemical isolation and protein-protein interaction mapping of eIF4G binding sites\",\n      \"pmids\": [\"19751846\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No reciprocal Co-IP or structural validation of the eIF4A:eIF4G interface\", \"Stoichiometry and assembly kinetics of eIF4F not determined\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"The subcellular site of action and essentiality of eIF4A were unresolved; co-localization with eIF4E and a combinatorial knockdown growth defect established it as a cytoplasmic, proliferation-essential factor.\",\n      \"evidence\": \"Immunofluorescence co-localization and dsRNA knockdown of eIF4A and eIF4E with growth assays\",\n      \"pmids\": [\"21331256\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"dsRNA specificity/off-target effects not fully controlled\", \"Single-gene essentiality via genetic knockout not established\", \"Contribution of eIF4A vs eIF4E to the combined phenotype not separable\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"The intrinsic enzymatic properties of eIF4A were undefined; in vitro assays on purified recombinant protein demonstrated Mg2+-dependent bipolar unwinding with 3'-to-5' preference and nucleic-acid-stimulated ATPase activity.\",\n      \"evidence\": \"Helicase unwinding, ATPase, and substrate-specificity assays with purified recombinant codon-optimized eIF4A\",\n      \"pmids\": [\"22750398\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Assays performed on DNA substrates rather than mRNA in a translation context\", \"No structural basis for directional preference\", \"Physiological substrate not identified\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Whether eIF4A was a druggable antimalarial target was untested; the rocaglate CR-1-31B was shown to disrupt eIF4A:RNA association and block initiation with potent in vitro and in vivo antiplasmodial activity.\",\n      \"evidence\": \"RNA-binding perturbation assay, ex vivo P. falciparum replication, and in vivo P. berghei and P. chabaudi mouse models\",\n      \"pmids\": [\"29463745\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct binding to eIF4A vs an eIF4A-containing complex not fully separated in vivo\", \"Selectivity over host eIF4A not quantified\", \"Resistance mechanisms not mapped\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}, {\"species\": \"Plasmodium berghei\", \"strain\": \"\"}, {\"species\": \"Plasmodium chabaudi\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Whether eIF4A associates with the mRNA 5' cap complex biochemically was addressed; cap affinity proteomics detected eIF4A, consistent with eIF4F membership but with low reproducibility.\",\n      \"evidence\": \"5' cap affinity matrix pulldown with LC-MS/MS of asexual-stage lysates\",\n      \"pmids\": [\"34890716\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Low reproducibility of eIF4A detection\", \"No functional validation of cap association\", \"Indirect vs direct cap-complex recruitment not resolved\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"The molecular determinants of rocaglate sensitivity were unconfirmed; thermal shift and docking validated silvestrol engagement and six critical eIF4A residues as binding determinants.\",\n      \"evidence\": \"In vitro thermal shift assay with recombinant eIF4A, molecular docking of eIF4A:RNA:rocaglate complexes, and in vitro antiplasmodial testing\",\n      \"pmids\": [\"37291191\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Predicted binding residues not validated by mutagenesis\", \"No experimental co-crystal structure of the eIF4A:RNA:rocaglate complex\", \"Docking energies are computational predictions\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The specific post-translational modifications generating eIF4A isoforms and the physiological mRNA targets whose translation depends on eIF4A helicase activity remain unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No modification mapping of the observed isoforms\", \"No genome-wide identification of eIF4A-dependent transcripts\", \"No high-resolution structure of parasite eIF4A alone or in eIF4F\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0003724\", \"supporting_discovery_ids\": [6]},\n      {\"term_id\": \"GO:0004386\", \"supporting_discovery_ids\": [6]},\n      {\"term_id\": \"GO:0016887\", \"supporting_discovery_ids\": [6]},\n      {\"term_id\": \"GO:0003723\", \"supporting_discovery_ids\": [2, 5]},\n      {\"term_id\": \"GO:0008135\", \"supporting_discovery_ids\": [0, 1]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005737\", \"supporting_discovery_ids\": [1]},\n      {\"term_id\": \"GO:0016281\", \"supporting_discovery_ids\": [0, 5]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"GO:0006413\", \"supporting_discovery_ids\": [0, 2]},\n      {\"term_id\": \"GO:0002181\", \"supporting_discovery_ids\": [1]}\n    ],\n    \"complexes\": [\n      \"eIF4F cap-binding translation initiation complex\"\n    ],\n    \"partners\": [\n      \"eIF4G (PfeIF4G)\",\n      \"eIF4E (PfeIF4E)\"\n    ],\n    \"other_free_text\": [\n      \"Essential for P. falciparum intraerythrocytic proliferation (combined eIF4A/eIF4E dsRNA knockdown gives ~67% growth inhibition) [#1]\",\n      \"Validated antimalarial drug target of rocaglates CR-1-31B and silvestrol, which disrupt eIF4A:RNA association [#2, #3]\",\n      \"Six eIF4A residues predicted as rocaglate-binding determinants by docking and thermal shift [#3]\",\n      \"Represented by multiple protein isoforms across intraerythrocytic development, inferred to arise from post-translational modification [#4]\"\n    ]\n  }\n}","audit_flag":null,"strains":[{"strain":"Pf3D7","gene_id":"PF3D7_1468700","name":null,"product":"eukaryotic initiation factor 4A","uniprot_acc":null,"chromosome":null,"start_pos":null},{"strain":"PbANKA","gene_id":"PBANKA_1331900","name":null,"product":"eukaryotic initiation factor 4a, putative","uniprot_acc":null,"chromosome":null,"start_pos":null},{"strain":"PkH","gene_id":"PKNH_1212500","name":null,"product":"eukaryotic initiation factor 4a, putative","uniprot_acc":null,"chromosome":null,"start_pos":null}],"gene_symbol":"eIF4A","evaluation":null}