{"gene":"PF3D7_0319100","run_date":"2026-09-08T19:44:17","timeline":{"discoveries":[{"year":2024,"finding":"PfRBX1 (PF3D7_0319100) is a component of a functional P. falciparum SCF complex (PfSCF), comprising PfCullin-1, PfRbx1, PfSkp1, PfFBXO1, and calcyclin binding protein, as confirmed by immunoprecipitation, in vitro protein-protein interaction assay, and in vitro ubiquitination assay.","method":"Immunoprecipitation, in vitro protein-protein interaction assay, in vitro ubiquitination assay","journal":"PLoS pathogens","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — reciprocal co-IP plus in vitro ubiquitination assay plus direct protein-protein interaction assay, single lab but multiple orthogonal methods","pmids":["38416790"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2024,"finding":"PfRBX1 (PF3D7_0319100) is a component of a functional P. falciparum CRL4-like complex (PfCRL4), comprising PfCullin-2, PfRbx1, cleavage and polyadenylation specificity factor subunit_A, and WD40 repeat proteins, as confirmed 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 1–2 / Moderate — co-IP plus in vitro ubiquitination assay plus sequence analysis, single lab but multiple orthogonal methods","pmids":["38416790"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2024,"finding":"In P. falciparum asexual blood stages, a cullin-4 scaffold (PfCRL4-related complex) interacts with the RING protein RBX1 (PF3D7_0319100), the adaptor protein DDB1, and a set of putative WD40-repeat receptor proteins including WIG1, as identified by co-immunoprecipitation and proteomic analysis.","method":"Co-immunoprecipitation, mass spectrometry/proteomics","journal":"mBio","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — co-IP with proteomic readout, single lab, two complementary methods confirming complex membership","pmids":["39207167"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2025,"finding":"PfRBX1 (PF3D7_0319100) co-precipitates with the novel P. falciparum E2 ubiquitin-conjugating enzyme PF3D7_0811400, and the N-terminus of PF3D7_0811400 is critical for mediating this interaction, as shown by co-immunoprecipitation with myc-tagged PfRbx1 and FLAG-tagged wild-type versus N-terminal truncation mutant of PF3D7_0811400.","method":"Co-immunoprecipitation, N-terminal truncation mutagenesis, immunoblotting","journal":"PLoS pathogens","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP with domain mutagenesis, single lab, two orthogonal approaches (pulldown + truncation mutant)","pmids":["40249735"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2023,"finding":"In P. berghei, RBX1 co-immunoprecipitates with the SCF complex components FBXO1-HA and FBXL2-HA, along with SKP1, CUL1, polyubiquitin, and Nedd8, establishing RBX1 as a component of the P. berghei SCFFBXO1 complex.","method":"Immunoprecipitation followed by mass spectrometry","journal":"Nature communications","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — reciprocal co-IP with mass spectrometry readout, single lab","pmids":["36898988"],"is_preprint":false,"attribution":[{"species":"Plasmodium berghei","strain":""}]},{"year":2025,"finding":"PfRBX1 (PF3D7_0319100) is a component of both a PfCullin1-linked CRL complex required for inner-membrane biogenesis and DNA replication, and a PfCullin4-linked CRL complex supporting DNA replication, as established by tagging and conditional deletion alongside interactome analysis.","method":"Endogenous tagging (HA-tag), conditional deletion (DiCre/LoxP), immunoprecipitation, proteomics","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — conditional KO with defined phenotypic readout plus co-IP/proteomics, single lab, multiple orthogonal methods","pmids":["40592953"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":"3D7"}]}],"current_model":"PfRBX1 (PF3D7_0319100) is a RING-domain protein that functions as the catalytic subunit shared between at least two distinct Cullin-RING Ligase (CRL) complexes in P. falciparum: a PfSCF complex (PfCullin-1/PfRbx1/PfSkp1/PfFBXO1) and a CRL4-like complex (PfCullin-2 or PfCullin-4/PfRbx1/DDB1/WD40-repeat receptors), both of which are enzymatically active for ubiquitination; RBX1 additionally interacts directly with a novel parasite-specific E2 conjugating enzyme (PF3D7_0811400), and orthologous RBX1 participation in the SCFFBXO1 complex has been confirmed in P. berghei, where the complex regulates cell division, gamete egress, and motile-form integrity."},"narrative":{"mechanistic_narrative":"RBX1 (PF3D7_0319100) is the shared RING-domain catalytic subunit of Cullin-RING ligase (CRL) complexes that drive protein ubiquitination in Plasmodium [PMID:38416790]. In P. falciparum it assembles into at least two distinct enzymatically active CRLs: a PfSCF complex comprising PfCullin-1, PfRbx1, PfSkp1, PfFBXO1 and calcyclin-binding protein [PMID:38416790], and a CRL4-like complex built on a cullin-4/cullin-2 scaffold with the DDB1 adaptor and WD40-repeat substrate receptors including WIG1 [PMID:38416790, PMID:39207167]. Both complexes are competent for in vitro ubiquitination, with RBX1 supplying the RING activity common to each [PMID:38416790]. RBX1 also binds directly to a novel parasite-specific E2 ubiquitin-conjugating enzyme, PF3D7_0811400, an interaction dependent on the E2's N-terminus [PMID:40249735]. Functionally, the PfCullin-1-linked CRL is required for inner-membrane biogenesis and DNA replication, while the PfCullin-4-linked CRL supports DNA replication [PMID:40592953]. In P. berghei, the orthologous RBX1 is confirmed as a component of the SCF-FBXO1 complex alongside SKP1, CUL1, FBXL2, polyubiquitin and Nedd8 [PMID:36898988]. Beyond complex membership and these replication/membrane phenotypes, the specific substrates ubiquitinated by RBX1-containing CRLs have not been characterized in the available corpus.","teleology":[{"year":2023,"claim":"Establishing whether a Plasmodium RBX1 participates in a canonical SCF architecture was needed to place parasite CRL machinery in the ubiquitin system; showing RBX1 co-purifies with SCF components in P. berghei anchored it as a bona fide SCF subunit.","evidence":"Immunoprecipitation followed by mass spectrometry of FBXO1-HA/FBXL2-HA in P. berghei","pmids":["36898988"],"confidence":"Medium","gaps":["Single-lab co-IP without RBX1 loss-of-function to test its individual requirement","Does not identify substrates of the P. berghei SCF-FBXO1 complex","P. falciparum orthology inferred, not directly tested here"],"attribution":[{"species":"Plasmodium berghei","strain":""}]},{"year":2024,"claim":"It was unknown whether P. falciparum RBX1 forms functional multi-complex CRLs; demonstrating its incorporation into both an enzymatically active PfSCF and a CRL4-like complex established RBX1 as a shared catalytic RING subunit of distinct parasite ligases.","evidence":"Reciprocal immunoprecipitation, in vitro protein-protein interaction and in vitro ubiquitination assays in P. falciparum","pmids":["38416790"],"confidence":"High","gaps":["Substrates ubiquitinated by either complex not defined","Contribution of RBX1 alone versus the assembled complex to activity not dissected","Structural basis of RBX1 partitioning between cullin scaffolds unresolved"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2024,"claim":"Defining the receptor architecture of the P. falciparum CRL4-like complex clarified how substrate specificity might be achieved; identifying RBX1 with a cullin-4 scaffold, DDB1, and WD40-repeat receptors including WIG1 mapped the adaptor/receptor layer around the RING subunit.","evidence":"Co-immunoprecipitation with mass spectrometry/proteomics in P. falciparum asexual blood stages","pmids":["39207167"],"confidence":"Medium","gaps":["Direct substrates recruited by the WD40 receptors not identified","Which receptors act via RBX1-dependent ubiquitination not established"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2025,"claim":"Connecting RBX1 to an E2 enzyme was needed to define the catalytic partner delivering ubiquitin; showing direct interaction with the parasite-specific E2 PF3D7_0811400 via its N-terminus identified the conjugating enzyme working with RBX1.","evidence":"Co-immunoprecipitation of myc-PfRbx1 with FLAG-tagged wild-type versus N-terminal truncation PF3D7_0811400, immunoblotting","pmids":["40249735"],"confidence":"Medium","gaps":["Single-lab co-IP without reciprocal in vitro reconstitution of E2-RBX1 ubiquitin transfer","Whether this E2 serves all RBX1 CRLs or a specific subset not addressed"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2025,"claim":"The cellular processes requiring RBX1-containing CRLs were unknown; conditional deletion linked the PfCullin-1 CRL to inner-membrane biogenesis and DNA replication and the PfCullin-4 CRL to DNA replication, assigning biological roles to the RBX1 ligases.","evidence":"Endogenous HA-tagging, DiCre/LoxP conditional deletion, immunoprecipitation and proteomics in P. falciparum 3D7","pmids":["40592953"],"confidence":"Medium","gaps":["Substrates whose ubiquitination underlies the membrane and replication phenotypes not identified","Direct requirement of the RBX1 subunit itself versus the cullin scaffolds not separated"],"attribution":[{"species":"Plasmodium falciparum","strain":"3D7"}]},{"year":null,"claim":"The physiological substrates targeted by RBX1-containing CRLs, and how RBX1 is partitioned among distinct cullin scaffolds, remain unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No substrate of any RBX1 CRL identified in the corpus","No structural model of RBX1 within the parasite complexes","Mechanism selecting cullin-1 versus cullin-4/cullin-2 scaffolds unknown"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]}],"mechanism_profile":{"molecular_activity":[],"localization":[],"pathway":[],"complexes":["PfSCF complex (PfCullin-1/PfRbx1/PfSkp1/PfFBXO1/calcyclin-binding protein)","PfCRL4-like complex (PfCullin-4 or PfCullin-2/PfRbx1/DDB1/WD40-repeat receptors incl. WIG1)","P. berghei SCF-FBXO1 complex (RBX1/SKP1/CUL1/FBXO1/FBXL2)"],"partners":["PFCULLIN-1","PFCULLIN-2","PFCULLIN-4","PFSKP1","PFFBXO1","CALCYCLIN-BINDING PROTEIN","DDB1","WD40-REPEAT RECEPTORS (INCL. WIG1)","PF3D7_0811400 (E2 UBIQUITIN-CONJUGATING ENZYME)","SKP1 (P. BERGHEI)","CUL1 (P. BERGHEI)","FBXL2 (P. BERGHEI)"],"other_free_text":["RBX1 is the shared RING catalytic subunit common to at least two distinct P. falciparum CRLs [#0, #1].","PfCullin-1 CRL required for inner-membrane biogenesis and DNA replication; PfCullin-4 CRL supports DNA replication [#5].","Interaction with the E2 PF3D7_0811400 depends on the E2 N-terminus [#3].","Substrates of RBX1-containing CRLs are not defined in the available corpus."]}},"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":["RBX1","PfRBX1"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"39207167","id":"PMC_39207167","title":"A multistage Plasmodium CRL4WIG1 ubiquitin ligase is critical for the formation of functional microtubule organization centers in microgametocytes.","date":"2024","source":"mBio","url":"https://pubmed.ncbi.nlm.nih.gov/39207167","citation_count":2,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["RBX1"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"40592953","id":"","title":"Functional characterisation of components in two Plasmodium falciparum Cullin-RING-Ligase complexes.","date":"2025","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/40592953","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["RBX1"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"40249735","id":"","title":"Activity-based protein profiling reveals both canonical and novel ubiquitin pathway enzymes in Plasmodium.","date":"2025","source":"PLoS pathogens","url":"https://pubmed.ncbi.nlm.nih.gov/40249735","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["RBX1"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"36898988","id":"","title":"The Skp1-Cullin1-FBXO1 complex is a pleiotropic regulator required for the formation of gametes and motile forms in Plasmodium berghei.","date":"2023","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/36898988","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["RBX1"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"39915671","id":"","title":"PfFBXO1 is essential for inner membrane complex formation in Plasmodium falciparum during both asexual and transmission stages.","date":"2025","source":"Communications biology","url":"https://pubmed.ncbi.nlm.nih.gov/39915671","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["RBX1"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"31658303","id":"","title":"Nedd8 hydrolysis by UCH proteases in Plasmodium parasites.","date":"2019","source":"PLoS pathogens","url":"https://pubmed.ncbi.nlm.nih.gov/31658303","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["RBX1"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"32488076","id":"","title":"Malaria parasites regulate intra-erythrocytic development duration via serpentine receptor 10 to coordinate with host rhythms.","date":"2020","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/32488076","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["RBX1"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"37101607","id":"","title":"Targeted Protein Degradation for Infectious Diseases: from Basic Biology to Drug Discovery.","date":"2022","source":"ACS bio & med chem Au","url":"https://pubmed.ncbi.nlm.nih.gov/37101607","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["RBX1"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"18545708","id":"","title":"Deciphering the ubiquitin-mediated pathway in apicomplexan parasites: a potential strategy to interfere with parasite virulence.","date":"2008","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/18545708","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["RBX1"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":14113,"output_tokens":1835,"usd":0.034932,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":6862,"output_tokens":3178,"usd":0.05688,"stage2_stop_reason":"end_turn"},"total_usd":0.091812,"stage1_batch_id":"msgbatch_01SRp5hyRc82zNjddxyCCXGk","stage2_batch_id":"msgbatch_01K3BTmBazRo3pRrMvyaPMTY","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2024,\n      \"finding\": \"PfRBX1 (PF3D7_0319100) is a component of a functional P. falciparum SCF complex (PfSCF), comprising PfCullin-1, PfRbx1, PfSkp1, PfFBXO1, and calcyclin binding protein, as confirmed by immunoprecipitation, in vitro protein-protein interaction assay, and in vitro ubiquitination assay.\",\n      \"method\": \"Immunoprecipitation, in vitro protein-protein interaction assay, in vitro ubiquitination assay\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — reciprocal co-IP plus in vitro ubiquitination assay plus direct protein-protein interaction assay, single lab but multiple orthogonal methods\",\n      \"pmids\": [\"38416790\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"PfRBX1 (PF3D7_0319100) is a component of a functional P. falciparum CRL4-like complex (PfCRL4), comprising PfCullin-2, PfRbx1, cleavage and polyadenylation specificity factor subunit_A, and WD40 repeat proteins, as confirmed 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 1–2 / Moderate — co-IP plus in vitro ubiquitination assay plus sequence analysis, single lab but multiple orthogonal methods\",\n      \"pmids\": [\"38416790\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"In P. falciparum asexual blood stages, a cullin-4 scaffold (PfCRL4-related complex) interacts with the RING protein RBX1 (PF3D7_0319100), the adaptor protein DDB1, and a set of putative WD40-repeat receptor proteins including WIG1, as identified by co-immunoprecipitation and proteomic analysis.\",\n      \"method\": \"Co-immunoprecipitation, mass spectrometry/proteomics\",\n      \"journal\": \"mBio\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — co-IP with proteomic readout, single lab, two complementary methods confirming complex membership\",\n      \"pmids\": [\"39207167\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"PfRBX1 (PF3D7_0319100) co-precipitates with the novel P. falciparum E2 ubiquitin-conjugating enzyme PF3D7_0811400, and the N-terminus of PF3D7_0811400 is critical for mediating this interaction, as shown by co-immunoprecipitation with myc-tagged PfRbx1 and FLAG-tagged wild-type versus N-terminal truncation mutant of PF3D7_0811400.\",\n      \"method\": \"Co-immunoprecipitation, N-terminal truncation mutagenesis, immunoblotting\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP with domain mutagenesis, single lab, two orthogonal approaches (pulldown + truncation mutant)\",\n      \"pmids\": [\"40249735\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"In P. berghei, RBX1 co-immunoprecipitates with the SCF complex components FBXO1-HA and FBXL2-HA, along with SKP1, CUL1, polyubiquitin, and Nedd8, establishing RBX1 as a component of the P. berghei SCFFBXO1 complex.\",\n      \"method\": \"Immunoprecipitation followed by mass spectrometry\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — reciprocal co-IP with mass spectrometry readout, single lab\",\n      \"pmids\": [\"36898988\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium berghei\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"PfRBX1 (PF3D7_0319100) is a component of both a PfCullin1-linked CRL complex required for inner-membrane biogenesis and DNA replication, and a PfCullin4-linked CRL complex supporting DNA replication, as established by tagging and conditional deletion alongside interactome analysis.\",\n      \"method\": \"Endogenous tagging (HA-tag), conditional deletion (DiCre/LoxP), immunoprecipitation, proteomics\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — conditional KO with defined phenotypic readout plus co-IP/proteomics, single lab, multiple orthogonal methods\",\n      \"pmids\": [\"40592953\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"3D7\"}]\n    }\n  ],\n  \"current_model\": \"PfRBX1 (PF3D7_0319100) is a RING-domain protein that functions as the catalytic subunit shared between at least two distinct Cullin-RING Ligase (CRL) complexes in P. falciparum: a PfSCF complex (PfCullin-1/PfRbx1/PfSkp1/PfFBXO1) and a CRL4-like complex (PfCullin-2 or PfCullin-4/PfRbx1/DDB1/WD40-repeat receptors), both of which are enzymatically active for ubiquitination; RBX1 additionally interacts directly with a novel parasite-specific E2 conjugating enzyme (PF3D7_0811400), and orthologous RBX1 participation in the SCFFBXO1 complex has been confirmed in P. berghei, where the complex regulates cell division, gamete egress, and motile-form integrity.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"RBX1 (PF3D7_0319100) is the shared RING-domain catalytic subunit of Cullin-RING ligase (CRL) complexes that drive protein ubiquitination in Plasmodium [#0, #1]. In P. falciparum it assembles into at least two distinct enzymatically active CRLs: a PfSCF complex comprising PfCullin-1, PfRbx1, PfSkp1, PfFBXO1 and calcyclin-binding protein [#0], and a CRL4-like complex built on a cullin-4/cullin-2 scaffold with the DDB1 adaptor and WD40-repeat substrate receptors including WIG1 [#1, #2]. Both complexes are competent for in vitro ubiquitination, with RBX1 supplying the RING activity common to each [#0, #1]. RBX1 also binds directly to a novel parasite-specific E2 ubiquitin-conjugating enzyme, PF3D7_0811400, an interaction dependent on the E2's N-terminus [#3]. Functionally, the PfCullin-1-linked CRL is required for inner-membrane biogenesis and DNA replication, while the PfCullin-4-linked CRL supports DNA replication [#5]. In P. berghei, the orthologous RBX1 is confirmed as a component of the SCF-FBXO1 complex alongside SKP1, CUL1, FBXL2, polyubiquitin and Nedd8 [#4]. Beyond complex membership and these replication/membrane phenotypes, the specific substrates ubiquitinated by RBX1-containing CRLs have not been characterized in the available corpus.\",\n  \"teleology\": [\n    {\n      \"year\": 2023,\n      \"claim\": \"Establishing whether a Plasmodium RBX1 participates in a canonical SCF architecture was needed to place parasite CRL machinery in the ubiquitin system; showing RBX1 co-purifies with SCF components in P. berghei anchored it as a bona fide SCF subunit.\",\n      \"evidence\": \"Immunoprecipitation followed by mass spectrometry of FBXO1-HA/FBXL2-HA in P. berghei\",\n      \"pmids\": [\"36898988\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Single-lab co-IP without RBX1 loss-of-function to test its individual requirement\",\n        \"Does not identify substrates of the P. berghei SCF-FBXO1 complex\",\n        \"P. falciparum orthology inferred, not directly tested here\"\n      ],\n      \"attribution\": [{\"species\": \"Plasmodium berghei\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"It was unknown whether P. falciparum RBX1 forms functional multi-complex CRLs; demonstrating its incorporation into both an enzymatically active PfSCF and a CRL4-like complex established RBX1 as a shared catalytic RING subunit of distinct parasite ligases.\",\n      \"evidence\": \"Reciprocal immunoprecipitation, in vitro protein-protein interaction and in vitro ubiquitination assays in P. falciparum\",\n      \"pmids\": [\"38416790\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Substrates ubiquitinated by either complex not defined\",\n        \"Contribution of RBX1 alone versus the assembled complex to activity not dissected\",\n        \"Structural basis of RBX1 partitioning between cullin scaffolds unresolved\"\n      ],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Defining the receptor architecture of the P. falciparum CRL4-like complex clarified how substrate specificity might be achieved; identifying RBX1 with a cullin-4 scaffold, DDB1, and WD40-repeat receptors including WIG1 mapped the adaptor/receptor layer around the RING subunit.\",\n      \"evidence\": \"Co-immunoprecipitation with mass spectrometry/proteomics in P. falciparum asexual blood stages\",\n      \"pmids\": [\"39207167\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Direct substrates recruited by the WD40 receptors not identified\",\n        \"Which receptors act via RBX1-dependent ubiquitination not established\"\n      ],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Connecting RBX1 to an E2 enzyme was needed to define the catalytic partner delivering ubiquitin; showing direct interaction with the parasite-specific E2 PF3D7_0811400 via its N-terminus identified the conjugating enzyme working with RBX1.\",\n      \"evidence\": \"Co-immunoprecipitation of myc-PfRbx1 with FLAG-tagged wild-type versus N-terminal truncation PF3D7_0811400, immunoblotting\",\n      \"pmids\": [\"40249735\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Single-lab co-IP without reciprocal in vitro reconstitution of E2-RBX1 ubiquitin transfer\",\n        \"Whether this E2 serves all RBX1 CRLs or a specific subset not addressed\"\n      ],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"The cellular processes requiring RBX1-containing CRLs were unknown; conditional deletion linked the PfCullin-1 CRL to inner-membrane biogenesis and DNA replication and the PfCullin-4 CRL to DNA replication, assigning biological roles to the RBX1 ligases.\",\n      \"evidence\": \"Endogenous HA-tagging, DiCre/LoxP conditional deletion, immunoprecipitation and proteomics in P. falciparum 3D7\",\n      \"pmids\": [\"40592953\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Substrates whose ubiquitination underlies the membrane and replication phenotypes not identified\",\n        \"Direct requirement of the RBX1 subunit itself versus the cullin scaffolds not separated\"\n      ],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"3D7\"}]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The physiological substrates targeted by RBX1-containing CRLs, and how RBX1 is partitioned among distinct cullin scaffolds, remain unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"No substrate of any RBX1 CRL identified in the corpus\",\n        \"No structural model of RBX1 within the parasite complexes\",\n        \"Mechanism selecting cullin-1 versus cullin-4/cullin-2 scaffolds unknown\"\n      ],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0061630\", \"supporting_discovery_ids\": [0, 1]},\n      {\"term_id\": \"GO:0031625\", \"supporting_discovery_ids\": [3]}\n    ],\n    \"localization\": [],\n    \"pathway\": [\n      {\"term_id\": \"GO:0016567\", \"supporting_discovery_ids\": [0, 1, 5]},\n      {\"term_id\": \"GO:0006260\", \"supporting_discovery_ids\": [5]}\n    ],\n    \"complexes\": [\n      \"PfSCF complex (PfCullin-1/PfRbx1/PfSkp1/PfFBXO1/calcyclin-binding protein)\",\n      \"PfCRL4-like complex (PfCullin-4 or PfCullin-2/PfRbx1/DDB1/WD40-repeat receptors incl. WIG1)\",\n      \"P. berghei SCF-FBXO1 complex (RBX1/SKP1/CUL1/FBXO1/FBXL2)\"\n    ],\n    \"partners\": [\n      \"PfCullin-1\",\n      \"PfCullin-2\",\n      \"PfCullin-4\",\n      \"PfSkp1\",\n      \"PfFBXO1\",\n      \"calcyclin-binding protein\",\n      \"DDB1\",\n      \"WD40-repeat receptors (incl. WIG1)\",\n      \"PF3D7_0811400 (E2 ubiquitin-conjugating enzyme)\",\n      \"SKP1 (P. berghei)\",\n      \"CUL1 (P. berghei)\",\n      \"FBXL2 (P. berghei)\"\n    ],\n    \"other_free_text\": [\n      \"RBX1 is the shared RING catalytic subunit common to at least two distinct P. falciparum CRLs [#0, #1].\",\n      \"PfCullin-1 CRL required for inner-membrane biogenesis and DNA replication; PfCullin-4 CRL supports DNA replication [#5].\",\n      \"Interaction with the E2 PF3D7_0811400 depends on the E2 N-terminus [#3].\",\n      \"Substrates of RBX1-containing CRLs are not defined in the available corpus.\"\n    ]\n  }\n}","audit_flag":null,"strains":[{"strain":"Pf3D7","gene_id":"PF3D7_0319100","name":null,"product":"E3 ubiquitin-protein ligase RBX1","uniprot_acc":null,"chromosome":null,"start_pos":null},{"strain":"PbANKA","gene_id":"PBANKA_0806200","name":null,"product":"E3 ubiquitin-protein ligase RBX1, putative","uniprot_acc":null,"chromosome":null,"start_pos":null},{"strain":"PkH","gene_id":"PKNH_0822400","name":null,"product":"E3 ubiquitin-protein ligase RBX1, putative","uniprot_acc":null,"chromosome":null,"start_pos":null}],"gene_symbol":"RBX1","evaluation":null}