{"gene":"PF3D7_0629800","run_date":"2026-09-08T12:51:26","timeline":{"discoveries":[{"year":2024,"finding":"PfCullin-2 (PF3D7_0629800) forms a functional human CRL4-like complex (PfCRL4) comprising PfRbx1, cleavage and polyadenylation specificity factor subunit_A, and WD40 repeat proteins, as established by immunoprecipitation, sequence analysis, and in vitro ubiquitination assay.","method":"Immunoprecipitation, sequence analysis, in vitro ubiquitination assay","journal":"PLoS pathogens","confidence":"High","confidence_rationale":"Tier 2 / Moderate — reciprocal Co-IP plus functional ubiquitination assay in a single rigorous study, multiple orthogonal methods confirming complex assembly and activity","pmids":["38416790"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2024,"finding":"PfCullin-2 is predominantly expressed in erythrocytic trophozoite and schizont stages, with nucleocytoplasmic localization and chromatin association, as determined by tagged-parasite imaging and fractionation.","method":"Tagged parasite expression, subcellular localization (fluorescence imaging/fractionation)","journal":"PLoS pathogens","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — direct localization experiment with chromatin association noted, single lab, single paper","pmids":["38416790"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2024,"finding":"Knock-down of PfCullin-2 at the protein level significantly decreased asexual and sexual erythrocytic stage development, caused poorly delimited merozoites and internal membrane structures in schizonts, and reduced the number of nuclei per parasite, establishing a critical role for PfCRL4 in cell division and membrane integrity.","method":"Conditional protein knock-down, morphological analysis (microscopy), quantification of nuclei per parasite","journal":"PLoS pathogens","confidence":"High","confidence_rationale":"Tier 2 / Moderate — clean loss-of-function with defined cellular phenotypes (growth, morphology, nuclei count) and proteomic pathway analysis, multiple readouts in one study","pmids":["38416790"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2024,"finding":"PfCullin-2 depletion significantly altered protein levels in multiple pathways, including protein translation and folding, lipid biosynthesis and transport, DNA replication, and protein degradation, as detected by proteomics following knock-down.","method":"Quantitative proteomics after conditional PfCullin-2 knock-down","journal":"PLoS pathogens","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — mass spectrometry proteomics in a single lab study; functional pathway links are inferred from protein-level changes, not directly validated","pmids":["38416790"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2020,"finding":"PfCullin-2 (and a second putative cullin) was identified as a physiological substrate of PfNEDD8 by immunoprecipitation of tagged wild-type PfNEDD8 followed by mass spectrometry, establishing that PfCullin-2 undergoes neddylation in P. falciparum.","method":"Immunoprecipitation followed by mass spectrometry of PfNEDD8-expressing parasites; heterologous complementation in S. cerevisiae rub1Δ and ubc12Δ strains","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — IP-MS identification of cullin substrate plus cross-species functional validation in yeast, single lab","pmids":["33214620"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]}],"current_model":"PF3D7_0629800 (PfCullin-2) is a scaffold subunit of a functional CRL4-like E3 ubiquitin ligase complex (PfCRL4) that also contains PfRbx1, cleavage and polyadenylation specificity factor subunit_A, and WD40 repeat proteins; it is neddylated by PfNEDD8, localizes to the nucleus/cytoplasm and chromatin in trophozoites and schizonts, and is essential for asexual and sexual erythrocytic development, being specifically required for nuclear division (schizogony) and maintenance of merozoite membrane integrity."},"narrative":{"mechanistic_narrative":"PF3D7_0629800 (PfCullin-2) is the scaffold subunit of a functional CRL4-like cullin-RING E3 ubiquitin ligase (PfCRL4) in Plasmodium falciparum, assembling with PfRbx1, a cleavage and polyadenylation specificity factor subunit, and WD40 repeat proteins into a complex with demonstrated in vitro ubiquitination activity [PMID:38416790]. Like canonical cullins, PfCullin-2 is regulated by neddylation: it was recovered as a physiological substrate of PfNEDD8 in P. falciparum [PMID:33214620]. The protein is expressed predominantly in erythrocytic trophozoite and schizont stages and shows nucleocytoplasmic localization with chromatin association [PMID:38416790]. Functionally, PfCullin-2/PfCRL4 is required for asexual and sexual erythrocytic development; its depletion reduces the number of nuclei per parasite and produces poorly delimited merozoites and aberrant internal membranes in schizonts, implicating the complex in nuclear division during schizogony and in merozoite membrane integrity [PMID:38416790]. Proteomic profiling after knock-down shows altered levels of proteins in translation and folding, lipid biosynthesis and transport, DNA replication, and protein degradation pathways [PMID:38416790], though the direct ubiquitination substrates of PfCRL4 have not been identified in the available corpus.","teleology":[{"year":2020,"claim":"Establishing whether PfCullin-2 is post-translationally regulated addressed whether Plasmodium cullins use the conserved neddylation switch that activates cullin-RING ligases.","evidence":"IP-MS of tagged wild-type PfNEDD8 in P. falciparum with heterologous complementation in S. cerevisiae rub1Δ/ubc12Δ strains","pmids":["33214620"],"confidence":"Medium","gaps":["Does not define the sites or functional consequence of neddylation on PfCullin-2 activity","Does not distinguish PfCullin-2 from the second putative cullin also recovered"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2024,"claim":"Identifying the composition and catalytic activity of the PfCullin-2 complex answered whether it forms a bona fide E3 ligase and defined its subunit architecture.","evidence":"Immunoprecipitation, sequence analysis, and in vitro ubiquitination assay in P. falciparum","pmids":["38416790"],"confidence":"High","gaps":["Physiological ubiquitination substrates of PfCRL4 not identified","Role of the CPSF subunit and WD40 proteins as substrate receptors not resolved"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2024,"claim":"Localization at trophozoite and schizont stages placed PfCullin-2 activity in the nucleus/cytoplasm and on chromatin, linking the complex to nuclear processes during erythrocytic development.","evidence":"Tagged-parasite fluorescence imaging and subcellular fractionation in P. falciparum","pmids":["38416790"],"confidence":"Medium","gaps":["Chromatin association not mapped to specific genomic loci or substrates","Single lab, single study"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2024,"claim":"Conditional depletion defined the essential cellular role of PfCullin-2, showing it is required for nuclear division and merozoite membrane integrity during schizogony.","evidence":"Conditional protein knock-down with morphological analysis and nuclei-per-parasite quantification in P. falciparum","pmids":["38416790"],"confidence":"High","gaps":["Mechanistic link between ligase activity and the nuclear-division/membrane defects not established","Sexual-stage requirement described but not mechanistically dissected"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2024,"claim":"Proteomics after knock-down provided the first systems-level view of pathways downstream of PfCullin-2, but did not establish which changes are direct.","evidence":"Quantitative proteomics after conditional PfCullin-2 knock-down in P. falciparum","pmids":["38416790"],"confidence":"Medium","gaps":["Cannot distinguish direct ubiquitination targets from downstream secondary effects","No target validated as a CRL4 substrate"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":null,"claim":"The direct ubiquitination substrates of PfCRL4 and how substrate selection is coupled to nuclear division and merozoite membrane biogenesis remain unknown.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No physiological substrate identified","Mechanism connecting E3 activity to schizogony phenotypes unresolved","No structural model of the PfCRL4 assembly"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]}],"mechanism_profile":{"molecular_activity":[],"localization":[],"pathway":[],"complexes":["PfCRL4 (CRL4-like cullin-RING E3 ubiquitin ligase: PfCullin-2, PfRbx1, cleavage and polyadenylation specificity factor subunit, WD40 repeat proteins)"],"partners":["PFRBX1","CLEAVAGE AND POLYADENYLATION SPECIFICITY FACTOR SUBUNIT_A","WD40 REPEAT PROTEINS","PFNEDD8"],"other_free_text":["PfCullin-2 knock-down alters protein levels in translation/folding, lipid biosynthesis and transport, DNA replication, and protein degradation pathways [#3].","Required for both asexual and sexual erythrocytic-stage development [#2]."]}},"prefetch_data":{},"corpus_meta":[{"pmid":"38416790","id":"PMC_38416790","title":"Plasmodium falciparum contains functional SCF and CRL4 ubiquitin E3 ligases, and CRL4 is critical for cell division and membrane integrity.","date":"2024","source":"PLoS pathogens","url":"https://pubmed.ncbi.nlm.nih.gov/38416790","citation_count":12,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["cullin-2"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"33214620","id":"PMC_33214620","title":"Characterization of Plasmodium falciparum NEDD8 and identification of cullins as its substrates.","date":"2020","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/33214620","citation_count":11,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["curated_ref"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":4175,"output_tokens":1366,"usd":0.016508,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":6236,"output_tokens":2607,"usd":0.048177,"stage2_stop_reason":"end_turn"},"total_usd":0.064685,"stage1_batch_id":"msgbatch_01DisAaTypwmC8QwdMBuePHo","stage2_batch_id":"msgbatch_0184zxZTqxiEfdYYfPMfkqjb","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2024,\n      \"finding\": \"PfCullin-2 (PF3D7_0629800) forms a functional human CRL4-like complex (PfCRL4) comprising PfRbx1, cleavage and polyadenylation specificity factor subunit_A, and WD40 repeat proteins, as established by immunoprecipitation, sequence analysis, and in vitro ubiquitination assay.\",\n      \"method\": \"Immunoprecipitation, sequence analysis, in vitro ubiquitination assay\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal Co-IP plus functional ubiquitination assay in a single rigorous study, multiple orthogonal methods confirming complex assembly and activity\",\n      \"pmids\": [\"38416790\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"PfCullin-2 is predominantly expressed in erythrocytic trophozoite and schizont stages, with nucleocytoplasmic localization and chromatin association, as determined by tagged-parasite imaging and fractionation.\",\n      \"method\": \"Tagged parasite expression, subcellular localization (fluorescence imaging/fractionation)\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — direct localization experiment with chromatin association noted, single lab, single paper\",\n      \"pmids\": [\"38416790\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Knock-down of PfCullin-2 at the protein level significantly decreased asexual and sexual erythrocytic stage development, caused poorly delimited merozoites and internal membrane structures in schizonts, and reduced the number of nuclei per parasite, establishing a critical role for PfCRL4 in cell division and membrane integrity.\",\n      \"method\": \"Conditional protein knock-down, morphological analysis (microscopy), quantification of nuclei per parasite\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — clean loss-of-function with defined cellular phenotypes (growth, morphology, nuclei count) and proteomic pathway analysis, multiple readouts in one study\",\n      \"pmids\": [\"38416790\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"PfCullin-2 depletion significantly altered protein levels in multiple pathways, including protein translation and folding, lipid biosynthesis and transport, DNA replication, and protein degradation, as detected by proteomics following knock-down.\",\n      \"method\": \"Quantitative proteomics after conditional PfCullin-2 knock-down\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — mass spectrometry proteomics in a single lab study; functional pathway links are inferred from protein-level changes, not directly validated\",\n      \"pmids\": [\"38416790\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"PfCullin-2 (and a second putative cullin) was identified as a physiological substrate of PfNEDD8 by immunoprecipitation of tagged wild-type PfNEDD8 followed by mass spectrometry, establishing that PfCullin-2 undergoes neddylation in P. falciparum.\",\n      \"method\": \"Immunoprecipitation followed by mass spectrometry of PfNEDD8-expressing parasites; heterologous complementation in S. cerevisiae rub1Δ and ubc12Δ strains\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — IP-MS identification of cullin substrate plus cross-species functional validation in yeast, single lab\",\n      \"pmids\": [\"33214620\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    }\n  ],\n  \"current_model\": \"PF3D7_0629800 (PfCullin-2) is a scaffold subunit of a functional CRL4-like E3 ubiquitin ligase complex (PfCRL4) that also contains PfRbx1, cleavage and polyadenylation specificity factor subunit_A, and WD40 repeat proteins; it is neddylated by PfNEDD8, localizes to the nucleus/cytoplasm and chromatin in trophozoites and schizonts, and is essential for asexual and sexual erythrocytic development, being specifically required for nuclear division (schizogony) and maintenance of merozoite membrane integrity.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"PF3D7_0629800 (PfCullin-2) is the scaffold subunit of a functional CRL4-like cullin-RING E3 ubiquitin ligase (PfCRL4) in Plasmodium falciparum, assembling with PfRbx1, a cleavage and polyadenylation specificity factor subunit, and WD40 repeat proteins into a complex with demonstrated in vitro ubiquitination activity [#0]. Like canonical cullins, PfCullin-2 is regulated by neddylation: it was recovered as a physiological substrate of PfNEDD8 in P. falciparum [#4]. The protein is expressed predominantly in erythrocytic trophozoite and schizont stages and shows nucleocytoplasmic localization with chromatin association [#1]. Functionally, PfCullin-2/PfCRL4 is required for asexual and sexual erythrocytic development; its depletion reduces the number of nuclei per parasite and produces poorly delimited merozoites and aberrant internal membranes in schizonts, implicating the complex in nuclear division during schizogony and in merozoite membrane integrity [#2]. Proteomic profiling after knock-down shows altered levels of proteins in translation and folding, lipid biosynthesis and transport, DNA replication, and protein degradation pathways [#3], though the direct ubiquitination substrates of PfCRL4 have not been identified in the available corpus.\"\n  ,\n  \"teleology\": [\n    {\n      \"year\": 2020,\n      \"claim\": \"Establishing whether PfCullin-2 is post-translationally regulated addressed whether Plasmodium cullins use the conserved neddylation switch that activates cullin-RING ligases.\",\n      \"evidence\": \"IP-MS of tagged wild-type PfNEDD8 in P. falciparum with heterologous complementation in S. cerevisiae rub1\\u0394/ubc12\\u0394 strains\",\n      \"pmids\": [\"33214620\"],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Does not define the sites or functional consequence of neddylation on PfCullin-2 activity\", \"Does not distinguish PfCullin-2 from the second putative cullin also recovered\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Identifying the composition and catalytic activity of the PfCullin-2 complex answered whether it forms a bona fide E3 ligase and defined its subunit architecture.\",\n      \"evidence\": \"Immunoprecipitation, sequence analysis, and in vitro ubiquitination assay in P. falciparum\",\n      \"pmids\": [\"38416790\"],\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Physiological ubiquitination substrates of PfCRL4 not identified\", \"Role of the CPSF subunit and WD40 proteins as substrate receptors not resolved\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Localization at trophozoite and schizont stages placed PfCullin-2 activity in the nucleus/cytoplasm and on chromatin, linking the complex to nuclear processes during erythrocytic development.\",\n      \"evidence\": \"Tagged-parasite fluorescence imaging and subcellular fractionation in P. falciparum\",\n      \"pmids\": [\"38416790\"],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Chromatin association not mapped to specific genomic loci or substrates\", \"Single lab, single study\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Conditional depletion defined the essential cellular role of PfCullin-2, showing it is required for nuclear division and merozoite membrane integrity during schizogony.\",\n      \"evidence\": \"Conditional protein knock-down with morphological analysis and nuclei-per-parasite quantification in P. falciparum\",\n      \"pmids\": [\"38416790\"],\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Mechanistic link between ligase activity and the nuclear-division/membrane defects not established\", \"Sexual-stage requirement described but not mechanistically dissected\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Proteomics after knock-down provided the first systems-level view of pathways downstream of PfCullin-2, but did not establish which changes are direct.\",\n      \"evidence\": \"Quantitative proteomics after conditional PfCullin-2 knock-down in P. falciparum\",\n      \"pmids\": [\"38416790\"],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Cannot distinguish direct ubiquitination targets from downstream secondary effects\", \"No target validated as a CRL4 substrate\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The direct ubiquitination substrates of PfCRL4 and how substrate selection is coupled to nuclear division and merozoite membrane biogenesis remain unknown.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"No physiological substrate identified\", \"Mechanism connecting E3 activity to schizogony phenotypes unresolved\", \"No structural model of the PfCRL4 assembly\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0004842\", \"supporting_discovery_ids\": [0]},\n      {\"term_id\": \"GO:0031625\", \"supporting_discovery_ids\": [0]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [1]},\n      {\"term_id\": \"GO:0005737\", \"supporting_discovery_ids\": [1]},\n      {\"term_id\": \"GO:0000785\", \"supporting_discovery_ids\": [1]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"GO:0016567\", \"supporting_discovery_ids\": [0]},\n      {\"term_id\": \"GO:0045116\", \"supporting_discovery_ids\": [4]},\n      {\"term_id\": \"GO:0007067\", \"supporting_discovery_ids\": [2]}\n    ],\n    \"complexes\": [\n      \"PfCRL4 (CRL4-like cullin-RING E3 ubiquitin ligase: PfCullin-2, PfRbx1, cleavage and polyadenylation specificity factor subunit, WD40 repeat proteins)\"\n    ],\n    \"partners\": [\n      \"PfRbx1\",\n      \"cleavage and polyadenylation specificity factor subunit_A\",\n      \"WD40 repeat proteins\",\n      \"PfNEDD8\"\n    ],\n    \"other_free_text\": [\n      \"PfCullin-2 knock-down alters protein levels in translation/folding, lipid biosynthesis and transport, DNA replication, and protein degradation pathways [#3].\",\n      \"Required for both asexual and sexual erythrocytic-stage development [#2].\"\n    ]\n  }\n}\n```","audit_flag":{"gene":"PF3D7_0629800","tier":"IDENTITY_CHECK","verdict":"identity unverified","subtype":"names_other_cards","uniprot_band":"","rules_fired":"R12","issue":"No cited paper names this card; 2 of 2 cited papers name other catalogue genes (PF3D7_0319100, PF3D7_1367000, PF3D7_1245300). The summary may describe one of those genes."},"strains":[{"strain":"Pf3D7","gene_id":"PF3D7_0629800","name":null,"product":"cullin-2","uniprot_acc":null,"chromosome":null,"start_pos":null},{"strain":"PbANKA","gene_id":"PBANKA_1128600","name":null,"product":"cullin-like protein, putative","uniprot_acc":null,"chromosome":null,"start_pos":null},{"strain":"PkH","gene_id":"PKNH_1119700","name":null,"product":"cullin-like protein, putative","uniprot_acc":null,"chromosome":null,"start_pos":null}],"gene_symbol":"","evaluation":null}