{"gene":"PF3D7_0613300","run_date":"2026-09-08T16:26:55","timeline":{"discoveries":[{"year":2012,"finding":"ROP14 (PFF0645c) localizes to the rhoptry bulb subcompartment of P. falciparum blood-stage merozoites and is released only after invasion is complete, distinguishing it from rhoptry neck proteins that release earlier during junction formation.","method":"Endogenous gene-tagging and high-definition fluorescence imaging of tagged P. falciparum invasins during erythrocyte invasion","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct localization by endogenous tagging with functional timing observation, single lab, single study","pmids":["23049965"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2012,"finding":"ROP14 (PF3D7_0613300) was removed from the list of P. falciparum transporter proteins, as new localization and functional data indicated it is not a transporter.","method":"Literature curation and re-analysis of localization/function data (citing Zuccala et al., 2012)","journal":"Frontiers in cellular and infection microbiology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — negative reclassification based on secondary curation of a single prior study, no new direct experiment","pmids":["35811673"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":"3D7"}]},{"year":2022,"finding":"ROP14 is dispensable for asexual blood-stage development in P. berghei; the rop14 locus (PBANKA_0111600) was disrupted without deleterious effects on asexual RBC-stage growth.","method":"CRISPR/Cas9-mediated gene disruption in P. berghei with genotyping and parasite growth monitoring","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct genetic knockout with confirmed genotyping and phenotypic assessment, single lab","pmids":["35202423"],"is_preprint":false,"attribution":[{"species":"Plasmodium berghei","strain":""}]},{"year":2022,"finding":"ROP14 was identified as a proximity-biotinylated protein in the interactome of both PfCERLI1 and PfCERLI2, suggesting ROP14 resides in the same subcellular neighborhood as these rhoptry bulb proteins in P. falciparum schizonts.","method":"DiQ-BioID (proximity-dependent biotinylation) followed by LC-MS/MS in P. falciparum","journal":"Communications biology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single proximity-biotinylation experiment, no reciprocal validation, single lab","pmids":["35140336"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2016,"finding":"ROP14 was identified as a lysine-methylated protein in P. falciparum asexual blood stages, indicating it undergoes lysine methylation as a post-translational modification.","method":"Immunoprecipitation with anti-methyl-lysine antibody followed by LC-MS/MS proteomic identification of methylated peptides","journal":"Scientific reports","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single mass spectrometry identification, no writer/eraser/reader defined, no functional consequence established","pmids":["27762281"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2025,"finding":"In P. berghei, the transcription factor PbSIP2 directly binds an RGTGCA cis-regulatory element in the rop14 promoter region, and mutation of this element causes ~30-fold reduction in rop14 transcript levels, establishing PbSIP2 as a direct transcriptional activator of rop14.","method":"ChIP-seq, DIP-seq, CRISPR/Cas9-mediated cis-element mutation, ChIP-qPCR, and RT-qPCR in P. berghei","journal":"Science advances","confidence":"High","confidence_rationale":"Tier 1 / Strong — multiple orthogonal methods (ChIP-seq, cis-element mutagenesis, ChIP-qPCR, RT-qPCR) in a single rigorous study demonstrating direct transcriptional regulation","pmids":["40117352"],"is_preprint":false,"attribution":[{"species":"Plasmodium berghei","strain":""}]},{"year":2023,"finding":"ROP14 was identified as a proximity-biotinylated protein in the rhoptry-proximal interactome of PbRON6 in P. berghei merozoites, placing it in the same apical complex neighborhood as RON2, RON3, and RON6.","method":"Proximity-labeling (BioID-based) interactome followed by LC-MS/MS in P. berghei","journal":"mBio","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single proximity-biotinylation/MS experiment, no reciprocal validation, no direct binding assay","pmids":["40996030"],"is_preprint":false,"attribution":[{"species":"Plasmodium berghei","strain":""}]},{"year":2021,"finding":"ROP14 protein was detected within P. falciparum-derived extracellular vesicles, where its encoding genomic DNA is packaged and protected from DNase I digestion, indicating rop14 DNA is selectively incorporated into EVs.","method":"DNase I protection assay on purified EVs followed by PCR amplification of nuclear gene loci including rop14","journal":"Nature communications","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single PCR-based detection of EV-encapsulated DNA; no functional consequence of ROP14 DNA packaging established","pmids":["29215015"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2023,"finding":"In P. berghei sporozoites, ROP14 (PBANKA_0111600) is biotinylated by RON12::ultraID, placing it in the proximity of the rhoptry neck protein RON12 and confirming its presence in the sporozoite rhoptry compartment.","method":"Proximity-dependent biotinylation (UltraID) combined with LC-MS/MS in P. berghei sporozoites","journal":"Frontiers in cellular and infection microbiology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single proximity-biotinylation experiment in sporozoites, no direct functional validation for ROP14 itself","pmids":["41668734"],"is_preprint":false,"attribution":[{"species":"Plasmodium berghei","strain":""}]}],"current_model":"ROP14 (PF3D7_0613300) is a rhoptry bulb protein in P. falciparum merozoites that is released post-invasion (after the moving junction has closed), is dispensable for asexual blood-stage replication in P. berghei, undergoes lysine methylation, is transcriptionally activated by the AP2 factor PbSIP2 via a direct RGTGCA promoter element in P. berghei, and is found in proximity to rhoptry neck and bulb proteins (RON6, CERLI1, CERLI2, RON12) across multiple life-cycle stages; beyond its rhoptry localization and transcriptional regulation, no enzymatic activity, receptor, or downstream effector has been mechanistically defined."},"narrative":{"mechanistic_narrative":"ROP14 (PF3D7_0613300) is a rhoptry bulb protein of Plasmodium merozoites implicated in the post-invasion phase of the erythrocyte invasion program [PMID:23049965]. In P. falciparum blood-stage merozoites it localizes to the rhoptry bulb subcompartment and is released only after invasion is complete, distinguishing it temporally from rhoptry neck proteins that discharge earlier during moving-junction formation [PMID:23049965]; an earlier annotation of ROP14 as a transporter was retracted on the basis of these localization and functional data [PMID:35811673]. Across life-cycle stages and species ROP14 is repeatedly found in the apical rhoptry neighborhood, being captured in proximity interactomes of the rhoptry bulb proteins CERLI1 and CERLI2 in P. falciparum schizonts [PMID:35140336] and of the rhoptry neck proteins RON6 and RON12 in P. berghei merozoites and sporozoites [PMID:40996030, PMID:41668734]. In P. berghei, ROP14 is dispensable for asexual blood-stage replication, as disruption of the locus produces no growth defect [PMID:35202423], and its transcription is directly activated by the AP2 factor PbSIP2, which binds an RGTGCA cis-element in the rop14 promoter [PMID:40117352]. ROP14 also undergoes lysine methylation in P. falciparum asexual stages [PMID:27762281]. Beyond this rhoptry localization, invasion-timed release, and transcriptional control, no enzymatic activity, receptor, or downstream effector of ROP14 has been mechanistically defined in the available corpus.","teleology":[{"year":2012,"claim":"Establishing where and when ROP14 acts was the first question, and endogenous tagging placed it in the rhoptry bulb with release timed to after invasion, separating it functionally from neck proteins that act during junction formation.","evidence":"endogenous gene-tagging and high-definition fluorescence imaging during erythrocyte invasion in P. falciparum","pmids":["23049965"],"confidence":"Medium","gaps":["What ROP14 does after post-invasion release is undefined","No binding partner or substrate identified","No structural information"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2012,"claim":"The prior annotation of ROP14 as a transporter was overturned, correcting its functional classification to a rhoptry protein.","evidence":"literature curation and re-analysis of localization/function data in P. falciparum 3D7","pmids":["35811673"],"confidence":"Low","gaps":["Negative reclassification based on secondary curation, no new direct experiment","Positive function not established"],"attribution":[{"species":"Plasmodium falciparum","strain":"3D7"}]},{"year":2016,"claim":"A post-translational modification of ROP14 was uncovered, showing it is lysine-methylated in asexual stages.","evidence":"anti-methyl-lysine immunoprecipitation and LC-MS/MS in P. falciparum","pmids":["27762281"],"confidence":"Low","gaps":["Single MS identification, no writer/eraser/reader defined","No functional consequence of methylation established"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2021,"claim":"ROP14 protein and its encoding DNA were found selectively packaged and protected within parasite-derived extracellular vesicles, raising the question of a non-canonical extracellular role.","evidence":"DNase I protection assay on purified EVs and PCR in P. falciparum","pmids":["29215015"],"confidence":"Low","gaps":["Single PCR-based detection","No functional consequence of rop14 DNA packaging established"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2022,"claim":"Whether ROP14 is required for blood-stage growth was tested, and knockout in P. berghei showed it is dispensable for asexual replication.","evidence":"CRISPR/Cas9 gene disruption with genotyping and growth monitoring in P. berghei","pmids":["35202423"],"confidence":"Medium","gaps":["Dispensability in asexual stages does not exclude a role in other stages","No molecular function assigned"],"attribution":[{"species":"Plasmodium berghei","strain":""}]},{"year":2022,"claim":"Proximity proteomics placed ROP14 in the molecular neighborhood of rhoptry bulb proteins CERLI1 and CERLI2, providing the first partner context.","evidence":"DiQ-BioID proximity biotinylation and LC-MS/MS in P. falciparum schizonts","pmids":["35140336"],"confidence":"Low","gaps":["Single proximity experiment, no reciprocal validation","Proximity does not establish direct interaction"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2023,"claim":"ROP14 was placed in the RON12 proximity network in sporozoites, extending its rhoptry association beyond blood-stage merozoites into the sporozoite stage.","evidence":"UltraID proximity biotinylation and LC-MS/MS in P. berghei sporozoites","pmids":["41668734"],"confidence":"Low","gaps":["Single proximity experiment, no direct functional validation for ROP14","Sporozoite-stage role undefined"],"attribution":[{"species":"Plasmodium berghei","strain":""}]},{"year":2025,"claim":"How rop14 expression is controlled was resolved by showing the AP2 factor PbSIP2 directly binds an RGTGCA element in the promoter and is required for full transcription, establishing direct transcriptional regulation.","evidence":"ChIP-seq, DIP-seq, CRISPR/Cas9 cis-element mutation, ChIP-qPCR and RT-qPCR in P. berghei","pmids":["40117352"],"confidence":"High","gaps":["Links regulation to transcript level but not to a protein function","Downstream biological consequence of PbSIP2-driven rop14 expression not defined"],"attribution":[{"species":"Plasmodium berghei","strain":""}]},{"year":null,"claim":"The core molecular function of ROP14 remains unknown: no enzymatic activity, receptor, host substrate, or downstream effector has been defined despite established rhoptry localization, post-invasion release, and transcriptional control.","evidence":"no direct evidence in the available corpus","pmids":[],"confidence":"Low","gaps":["No biochemical activity assigned","No direct binding partner validated","Role after post-invasion release into the host cell unknown"],"attribution":[]}],"mechanism_profile":{"molecular_activity":[],"localization":[],"pathway":[],"complexes":[],"partners":["CERLI1 (PROXIMITY, P. FALCIPARUM) [#3]","CERLI2 (PROXIMITY, P. FALCIPARUM) [#3]","RON6 (PROXIMITY, P. BERGHEI) [#6]","RON12 (PROXIMITY, P. BERGHEI SPOROZOITES) [#8]","PBSIP2 (DIRECT TRANSCRIPTIONAL ACTIVATOR OF ROP14, P. BERGHEI) [#5]"],"other_free_text":["Rhoptry bulb protein released only after erythrocyte invasion is complete, distinct from earlier-releasing neck proteins [#0].","Dispensable for asexual blood-stage growth in P. berghei [#2].","Undergoes lysine methylation in P. falciparum asexual stages; no writer/reader/eraser or functional consequence defined [#4].","Previously mis-annotated as a transporter; reclassification removed it from the transporter list [#1].","Protein and encoding DNA selectively packaged into P. falciparum extracellular vesicles [#7]."]}},"prefetch_data":{},"corpus_meta":[{"pmid":"23049965","id":"PMC_23049965","title":"Subcompartmentalisation of proteins in the rhoptries correlates with ordered events of erythrocyte invasion by the blood stage malaria parasite.","date":"2012","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/23049965","citation_count":39,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["PFF0645c"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"40117352","id":"","title":"SIP2 is the master transcription factor of Plasmodium merozoite formation.","date":"2025","source":"Science advances","url":"https://pubmed.ncbi.nlm.nih.gov/40117352","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["ROP14"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"41668734","id":"","title":"Identification of a novel rhoptry protein expressed predominantly in Plasmodium sporozoites.","date":"2026","source":"Frontiers in cellular and infection microbiology","url":"https://pubmed.ncbi.nlm.nih.gov/41668734","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["ROP14"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"40996030","id":"","title":"Plasmodium berghei rhoptry neck protein 6 maintains parasite infectivity and virulence.","date":"2025","source":"mBio","url":"https://pubmed.ncbi.nlm.nih.gov/40996030","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["ROP14"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"35202423","id":"","title":"Chromosome splitting of Plasmodium berghei using the CRISPR/Cas9 system.","date":"2022","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/35202423","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["ROP14"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"37698406","id":"","title":"The global transcriptome of Plasmodium falciparum mid-stage gametocytes (stages II-IV) appears largely conserved and gametocyte-specific gene expression patterns vary in clinical isolates.","date":"2023","source":"Microbiology spectrum","url":"https://pubmed.ncbi.nlm.nih.gov/37698406","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["ROP14"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"35811673","id":"","title":"Updated List of Transport Proteins in Plasmodium falciparum.","date":"2022","source":"Frontiers in cellular and infection microbiology","url":"https://pubmed.ncbi.nlm.nih.gov/35811673","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["ROP14"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"35140336","id":"","title":"Cell biological analysis reveals an essential role for Pfcerli2 in erythrocyte invasion by malaria parasites.","date":"2022","source":"Communications biology","url":"https://pubmed.ncbi.nlm.nih.gov/35140336","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["ROP14"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"36622145","id":"","title":"Targetome Analysis of Malaria Sporozoite Transcription Factor AP2-Sp Reveals Its Role as a Master Regulator.","date":"2023","source":"mBio","url":"https://pubmed.ncbi.nlm.nih.gov/36622145","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["ROP14"],"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":["ROP14"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"31856180","id":"","title":"Latent transcriptional variations of individual Plasmodium falciparum uncovered by single-cell RNA-seq and fluorescence imaging.","date":"2019","source":"PLoS genetics","url":"https://pubmed.ncbi.nlm.nih.gov/31856180","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["ROP14"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"30142149","id":"","title":"Plasmodium vivax-like genome sequences shed new insights into Plasmodium vivax biology and evolution.","date":"2018","source":"PLoS biology","url":"https://pubmed.ncbi.nlm.nih.gov/30142149","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["ROP14"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"29784978","id":"","title":"Genomes of all known members of a Plasmodium subgenus reveal paths to virulent human malaria.","date":"2018","source":"Nature microbiology","url":"https://pubmed.ncbi.nlm.nih.gov/29784978","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["ROP14"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"29215015","id":"","title":"Malaria parasite DNA-harbouring vesicles activate cytosolic immune sensors.","date":"2017","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/29215015","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["ROP14"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"29580379","id":"","title":"Single-cell RNA-seq reveals hidden transcriptional variation in malaria parasites.","date":"2018","source":"eLife","url":"https://pubmed.ncbi.nlm.nih.gov/29580379","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["ROP14"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"27762281","id":"","title":"Widespread occurrence of lysine methylation in Plasmodium falciparum proteins at asexual blood stages.","date":"2016","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/27762281","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["ROP14"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":18448,"output_tokens":2254,"usd":0.044577,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":7528,"output_tokens":3093,"usd":0.057482,"stage2_stop_reason":"end_turn"},"total_usd":0.102059,"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\": 2012,\n      \"finding\": \"ROP14 (PFF0645c) localizes to the rhoptry bulb subcompartment of P. falciparum blood-stage merozoites and is released only after invasion is complete, distinguishing it from rhoptry neck proteins that release earlier during junction formation.\",\n      \"method\": \"Endogenous gene-tagging and high-definition fluorescence imaging of tagged P. falciparum invasins during erythrocyte invasion\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct localization by endogenous tagging with functional timing observation, single lab, single study\",\n      \"pmids\": [\"23049965\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"ROP14 (PF3D7_0613300) was removed from the list of P. falciparum transporter proteins, as new localization and functional data indicated it is not a transporter.\",\n      \"method\": \"Literature curation and re-analysis of localization/function data (citing Zuccala et al., 2012)\",\n      \"journal\": \"Frontiers in cellular and infection microbiology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — negative reclassification based on secondary curation of a single prior study, no new direct experiment\",\n      \"pmids\": [\"35811673\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"3D7\"}]\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"ROP14 is dispensable for asexual blood-stage development in P. berghei; the rop14 locus (PBANKA_0111600) was disrupted without deleterious effects on asexual RBC-stage growth.\",\n      \"method\": \"CRISPR/Cas9-mediated gene disruption in P. berghei with genotyping and parasite growth monitoring\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct genetic knockout with confirmed genotyping and phenotypic assessment, single lab\",\n      \"pmids\": [\"35202423\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium berghei\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"ROP14 was identified as a proximity-biotinylated protein in the interactome of both PfCERLI1 and PfCERLI2, suggesting ROP14 resides in the same subcellular neighborhood as these rhoptry bulb proteins in P. falciparum schizonts.\",\n      \"method\": \"DiQ-BioID (proximity-dependent biotinylation) followed by LC-MS/MS in P. falciparum\",\n      \"journal\": \"Communications biology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single proximity-biotinylation experiment, no reciprocal validation, single lab\",\n      \"pmids\": [\"35140336\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"ROP14 was identified as a lysine-methylated protein in P. falciparum asexual blood stages, indicating it undergoes lysine methylation as a post-translational modification.\",\n      \"method\": \"Immunoprecipitation with anti-methyl-lysine antibody followed by LC-MS/MS proteomic identification of methylated peptides\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single mass spectrometry identification, no writer/eraser/reader defined, no functional consequence established\",\n      \"pmids\": [\"27762281\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"In P. berghei, the transcription factor PbSIP2 directly binds an RGTGCA cis-regulatory element in the rop14 promoter region, and mutation of this element causes ~30-fold reduction in rop14 transcript levels, establishing PbSIP2 as a direct transcriptional activator of rop14.\",\n      \"method\": \"ChIP-seq, DIP-seq, CRISPR/Cas9-mediated cis-element mutation, ChIP-qPCR, and RT-qPCR in P. berghei\",\n      \"journal\": \"Science advances\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — multiple orthogonal methods (ChIP-seq, cis-element mutagenesis, ChIP-qPCR, RT-qPCR) in a single rigorous study demonstrating direct transcriptional regulation\",\n      \"pmids\": [\"40117352\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium berghei\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"ROP14 was identified as a proximity-biotinylated protein in the rhoptry-proximal interactome of PbRON6 in P. berghei merozoites, placing it in the same apical complex neighborhood as RON2, RON3, and RON6.\",\n      \"method\": \"Proximity-labeling (BioID-based) interactome followed by LC-MS/MS in P. berghei\",\n      \"journal\": \"mBio\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single proximity-biotinylation/MS experiment, no reciprocal validation, no direct binding assay\",\n      \"pmids\": [\"40996030\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium berghei\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"ROP14 protein was detected within P. falciparum-derived extracellular vesicles, where its encoding genomic DNA is packaged and protected from DNase I digestion, indicating rop14 DNA is selectively incorporated into EVs.\",\n      \"method\": \"DNase I protection assay on purified EVs followed by PCR amplification of nuclear gene loci including rop14\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single PCR-based detection of EV-encapsulated DNA; no functional consequence of ROP14 DNA packaging established\",\n      \"pmids\": [\"29215015\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"In P. berghei sporozoites, ROP14 (PBANKA_0111600) is biotinylated by RON12::ultraID, placing it in the proximity of the rhoptry neck protein RON12 and confirming its presence in the sporozoite rhoptry compartment.\",\n      \"method\": \"Proximity-dependent biotinylation (UltraID) combined with LC-MS/MS in P. berghei sporozoites\",\n      \"journal\": \"Frontiers in cellular and infection microbiology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single proximity-biotinylation experiment in sporozoites, no direct functional validation for ROP14 itself\",\n      \"pmids\": [\"41668734\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium berghei\", \"strain\": \"\"}]\n    }\n  ],\n  \"current_model\": \"ROP14 (PF3D7_0613300) is a rhoptry bulb protein in P. falciparum merozoites that is released post-invasion (after the moving junction has closed), is dispensable for asexual blood-stage replication in P. berghei, undergoes lysine methylation, is transcriptionally activated by the AP2 factor PbSIP2 via a direct RGTGCA promoter element in P. berghei, and is found in proximity to rhoptry neck and bulb proteins (RON6, CERLI1, CERLI2, RON12) across multiple life-cycle stages; beyond its rhoptry localization and transcriptional regulation, no enzymatic activity, receptor, or downstream effector has been mechanistically defined.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"ROP14 (PF3D7_0613300) is a rhoptry bulb protein of Plasmodium merozoites implicated in the post-invasion phase of the erythrocyte invasion program [#0]. In P. falciparum blood-stage merozoites it localizes to the rhoptry bulb subcompartment and is released only after invasion is complete, distinguishing it temporally from rhoptry neck proteins that discharge earlier during moving-junction formation [#0]; an earlier annotation of ROP14 as a transporter was retracted on the basis of these localization and functional data [#1]. Across life-cycle stages and species ROP14 is repeatedly found in the apical rhoptry neighborhood, being captured in proximity interactomes of the rhoptry bulb proteins CERLI1 and CERLI2 in P. falciparum schizonts [#3] and of the rhoptry neck proteins RON6 and RON12 in P. berghei merozoites and sporozoites [#6, #8]. In P. berghei, ROP14 is dispensable for asexual blood-stage replication, as disruption of the locus produces no growth defect [#2], and its transcription is directly activated by the AP2 factor PbSIP2, which binds an RGTGCA cis-element in the rop14 promoter [#5]. ROP14 also undergoes lysine methylation in P. falciparum asexual stages [#4]. Beyond this rhoptry localization, invasion-timed release, and transcriptional control, no enzymatic activity, receptor, or downstream effector of ROP14 has been mechanistically defined in the available corpus.\",\n  \"teleology\": [\n    {\n      \"year\": 2012,\n      \"claim\": \"Establishing where and when ROP14 acts was the first question, and endogenous tagging placed it in the rhoptry bulb with release timed to after invasion, separating it functionally from neck proteins that act during junction formation.\",\n      \"evidence\": \"endogenous gene-tagging and high-definition fluorescence imaging during erythrocyte invasion in P. falciparum\",\n      \"pmids\": [\"23049965\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"What ROP14 does after post-invasion release is undefined\", \"No binding partner or substrate identified\", \"No structural information\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"The prior annotation of ROP14 as a transporter was overturned, correcting its functional classification to a rhoptry protein.\",\n      \"evidence\": \"literature curation and re-analysis of localization/function data in P. falciparum 3D7\",\n      \"pmids\": [\"35811673\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Negative reclassification based on secondary curation, no new direct experiment\", \"Positive function not established\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"3D7\"}]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"A post-translational modification of ROP14 was uncovered, showing it is lysine-methylated in asexual stages.\",\n      \"evidence\": \"anti-methyl-lysine immunoprecipitation and LC-MS/MS in P. falciparum\",\n      \"pmids\": [\"27762281\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single MS identification, no writer/eraser/reader defined\", \"No functional consequence of methylation established\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"ROP14 protein and its encoding DNA were found selectively packaged and protected within parasite-derived extracellular vesicles, raising the question of a non-canonical extracellular role.\",\n      \"evidence\": \"DNase I protection assay on purified EVs and PCR in P. falciparum\",\n      \"pmids\": [\"29215015\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single PCR-based detection\", \"No functional consequence of rop14 DNA packaging established\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Whether ROP14 is required for blood-stage growth was tested, and knockout in P. berghei showed it is dispensable for asexual replication.\",\n      \"evidence\": \"CRISPR/Cas9 gene disruption with genotyping and growth monitoring in P. berghei\",\n      \"pmids\": [\"35202423\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Dispensability in asexual stages does not exclude a role in other stages\", \"No molecular function assigned\"],\n      \"attribution\": [{\"species\": \"Plasmodium berghei\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Proximity proteomics placed ROP14 in the molecular neighborhood of rhoptry bulb proteins CERLI1 and CERLI2, providing the first partner context.\",\n      \"evidence\": \"DiQ-BioID proximity biotinylation and LC-MS/MS in P. falciparum schizonts\",\n      \"pmids\": [\"35140336\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single proximity experiment, no reciprocal validation\", \"Proximity does not establish direct interaction\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"ROP14 was placed in the RON12 proximity network in sporozoites, extending its rhoptry association beyond blood-stage merozoites into the sporozoite stage.\",\n      \"evidence\": \"UltraID proximity biotinylation and LC-MS/MS in P. berghei sporozoites\",\n      \"pmids\": [\"41668734\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single proximity experiment, no direct functional validation for ROP14\", \"Sporozoite-stage role undefined\"],\n      \"attribution\": [{\"species\": \"Plasmodium berghei\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"How rop14 expression is controlled was resolved by showing the AP2 factor PbSIP2 directly binds an RGTGCA element in the promoter and is required for full transcription, establishing direct transcriptional regulation.\",\n      \"evidence\": \"ChIP-seq, DIP-seq, CRISPR/Cas9 cis-element mutation, ChIP-qPCR and RT-qPCR in P. berghei\",\n      \"pmids\": [\"40117352\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Links regulation to transcript level but not to a protein function\", \"Downstream biological consequence of PbSIP2-driven rop14 expression not defined\"],\n      \"attribution\": [{\"species\": \"Plasmodium berghei\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The core molecular function of ROP14 remains unknown: no enzymatic activity, receptor, host substrate, or downstream effector has been defined despite established rhoptry localization, post-invasion release, and transcriptional control.\",\n      \"evidence\": \"no direct evidence in the available corpus\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No biochemical activity assigned\", \"No direct binding partner validated\", \"Role after post-invasion release into the host cell unknown\"],\n      \"attribution\": []\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [],\n    \"localization\": [\n      {\"term_id\": \"GO:0020008\", \"supporting_discovery_ids\": [0, 3, 6, 8]},\n      {\"term_id\": \"GO:1903561\", \"supporting_discovery_ids\": [7]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"GO:0044409\", \"supporting_discovery_ids\": [0]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\n      \"CERLI1 (proximity, P. falciparum) [#3]\",\n      \"CERLI2 (proximity, P. falciparum) [#3]\",\n      \"RON6 (proximity, P. berghei) [#6]\",\n      \"RON12 (proximity, P. berghei sporozoites) [#8]\",\n      \"PbSIP2 (direct transcriptional activator of rop14, P. berghei) [#5]\"\n    ],\n    \"other_free_text\": [\n      \"Rhoptry bulb protein released only after erythrocyte invasion is complete, distinct from earlier-releasing neck proteins [#0].\",\n      \"Dispensable for asexual blood-stage growth in P. berghei [#2].\",\n      \"Undergoes lysine methylation in P. falciparum asexual stages; no writer/reader/eraser or functional consequence defined [#4].\",\n      \"Previously mis-annotated as a transporter; reclassification removed it from the transporter list [#1].\",\n      \"Protein and encoding DNA selectively packaged into P. falciparum extracellular vesicles [#7].\"\n    ]\n  }\n}","audit_flag":null,"strains":[{"strain":"Pf3D7","gene_id":"PF3D7_0613300","name":null,"product":"rhoptry protein ROP14","uniprot_acc":null,"chromosome":null,"start_pos":null},{"strain":"PbANKA","gene_id":"PBANKA_0111600","name":null,"product":"rhoptry protein ROP14, putative","uniprot_acc":null,"chromosome":null,"start_pos":null},{"strain":"PkH","gene_id":"PKNH_1136700","name":null,"product":"rhoptry protein ROP14, putative","uniprot_acc":null,"chromosome":null,"start_pos":null}],"gene_symbol":"ROP14","evaluation":null}