{"gene":"PRPF4","run_date":"2026-06-10T06:43:36","timeline":{"discoveries":[{"year":1997,"finding":"Human U4/U6-60kD protein (PRPF4/SNRNP60) was identified as a component of the U4/U6.U5 tri-snRNP that specifically associates with U4/U6 snRNP even at salt concentrations where the tri-snRNP complex dissociates; its primary structure contains seven WD repeats, predicting a beta-propeller topology homologous to G-protein beta subunits.","method":"Immunoaffinity chromatography, cDNA cloning and sequencing, sequence homology analysis","journal":"RNA (New York, N.Y.)","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal immunoaffinity purification with direct biochemical characterization; replicated by independent group (PMID:9328476)","pmids":["9257651"],"is_preprint":false},{"year":1997,"finding":"Hprp3p and Hprp4p interact with each other; Hprp3p co-immunoprecipitates with U4, U6, and U5 snRNAs, placing both proteins in the U4/U6.U5 tri-snRNP; the first 100 amino acids of Hprp3p are not essential for the Hprp3p–Hprp4p interaction.","method":"Co-immunoprecipitation, antibody pulldown, snRNA co-precipitation","journal":"Human molecular genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal co-IP with snRNA detection, single lab but two orthogonal methods","pmids":["9328476"],"is_preprint":false},{"year":2002,"finding":"The central region of Hprp3p (lacking N-terminal 194 aa or C-terminal 240 aa) is necessary and sufficient for binding Hprp4p; this Hprp3p–Hprp4p interaction is required for association of the complex with U4/U6 snRNAs, indicating that Hprp3p recruits Hprp4p to the U4/U6 snRNP.","method":"Co-immunoprecipitation, isothermal titration calorimetry, primer extension analysis, bacterial and mammalian expression of deletion mutants","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro ITC (thermodynamic binding), co-IP in HeLa nuclear extracts, primer extension for functional readout; single lab but three orthogonal methods","pmids":["11971898"],"is_preprint":false},{"year":2000,"finding":"Human hPRP4, a CDK-like kinase homologous to S. pombe Prp4p, is activated by EGF or forskolin treatment and phosphorylates Elk-1 at Thr-417 (a site distinct from other MAPKs), leading to Elk-1 transactivation; this demonstrates a signaling/transcription-factor activation function for hPRP4 beyond splicing.","method":"Kinase activity assay in COS-7 cells, EGF/forskolin stimulation, site-specific phosphorylation analysis, luciferase reporter (Elk-1 trans-activation)","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — direct kinase assay with substrate phosphorylation and reporter readout, single lab, single study; note this concerns the kinase paralog of PRPF4 (CDK-like), context is mammalian cells","pmids":["10799319"],"is_preprint":false},{"year":2001,"finding":"S. pombe Prp4p kinase activity is required for pre-mRNA splicing in vivo; Prp4p phosphorylates the non-SR splicing factor Prp1p in vitro and interacts with Prp1p in two-hybrid assays; loss of Prp4p kinase activity impairs G1-S and G2-M cell cycle progression.","method":"In vitro kinase assay with bacterially expressed Prp1p, yeast two-hybrid, in vivo phosphorylation labeling, genetic analysis","journal":"EMBO reports","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — in vitro kinase reconstitution plus in vivo labeling and two-hybrid; fission yeast ortholog, single lab","pmids":["11252721"],"is_preprint":false},{"year":2002,"finding":"The deubiquitinating enzyme Ubp21p of S. pombe stabilizes a mutant form of Prp4p kinase; Ubp21 was identified as a high-copy suppressor of a mutation in the ALKHP motif of Prp4p's kinase subdomain XI, indicating that ubiquitin-mediated proteolysis regulates Prp4p steady-state levels.","method":"High-copy suppressor screen, mutational analysis of kinase domain, Ubp21 characterization","journal":"Molecular genetics and genomics","confidence":"Low","confidence_rationale":"Tier 3 / Weak — genetic suppressor screen in fission yeast, single lab, limited direct biochemical validation of the ubiquitination mechanism","pmids":["11919719"],"is_preprint":false},{"year":2014,"finding":"The PRPF4 p.Pro315Leu variant causes dominant-negative effects: overexpression in cells upregulates tri-snRNP components (PRPF4 itself included) and alters SC35 splicing-factor localization as a compensatory response; overexpression of mutant but not wild-type PRPF4 causes retinal deformities in zebrafish and worsens phenotypes in prpf4 morphants, establishing PRPF4 as a U4/U6-U5 tri-snRNP component whose dysfunction leads to adRP via haploinsufficiency and dominant-negative mechanisms.","method":"Luciferase promoter assay, overexpression in fibroblasts and HeLa cells, SC35 immunostaining, zebrafish overexpression and morpholino knockdown","journal":"Human molecular genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal methods (reporter assay, cell overexpression, in vivo zebrafish), single lab","pmids":["24419317"],"is_preprint":false},{"year":2014,"finding":"The PRPF4 p.R192H variant disrupts the binding interface between PRPF4 and PRPF3, preventing PRPF4 from integrating into the tri-snRNP; introduction of the equivalent mutation in zebrafish prpf4 causes complete loss of function in vivo, and the variant behaves as a functional null (haploinsufficiency) in a human cell line.","method":"Biochemical binding assays (co-IP/pulldown), cell-line tri-snRNP integration assay, zebrafish in vivo functional rescue experiments","journal":"PloS one","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal biochemical assays plus in vivo zebrafish validation; independently supports and extends findings from PMID:11971898 on PRPF3–PRPF4 interface","pmids":["25383878"],"is_preprint":false},{"year":2017,"finding":"PPIH (cyclophilin H) binds the N-terminus of PRPF4 at two distinct interaction sites (bipartite binding); the N-terminal region of PRPF4 is intrinsically disordered and does not adopt secondary structure upon PPIH binding; mutations in both sites are required to abrogate complex formation.","method":"Recombinant protein expression and purification, complex formation assay, mutational analysis (point mutations uncoupling binding sites), NMR/biophysical characterization","journal":"The Biochemical journal","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — in vitro reconstitution with mutagenesis, single lab, no replication","pmids":["28935721"],"is_preprint":false},{"year":2018,"finding":"In zebrafish, prpf4 loss-of-function leads to p53-dependent apoptosis in neural cells and defects in posterior lateral line primordium (pLLP) migration; RNA-Seq shows prpf4 deficiency impairs spliceosome assembly, causes compensatory upregulation of spliceosomal genes, and alters pre-mRNA splicing; Fgf, Wnt/β-catenin, and chemokine signaling pathways are disrupted.","method":"Gene-trap zebrafish mutant, p53 rescue experiments, RNA-Seq splicing analysis, signaling pathway analysis","journal":"Journal of genetics and genomics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic loss-of-function with defined cellular and molecular phenotypes plus transcriptome-level splicing analysis; single lab","pmids":["30174136"],"is_preprint":false},{"year":2019,"finding":"PRPF4 knockdown in breast cancer cell lines reduces proliferation, migration, invasion, and induces apoptosis; microarray analysis indicates reduced phosphorylation of p38 MAPK associated with decreased expression of PPIP5K1, PPIPK2, and YWHAE, placing PRPF4 upstream of p38 MAPK signaling.","method":"Stable shRNA knockdown in MCF7 and MDA-MB-468 cells, colony formation assay, migration/invasion assays, microarray, Western blot for p38 MAPK phosphorylation","journal":"Molecular and cellular probes","confidence":"Low","confidence_rationale":"Tier 3 / Weak — knockdown with phenotypic readouts and pathway inference by microarray, no direct mechanistic reconstitution; single lab","pmids":["31445970"],"is_preprint":false},{"year":2025,"finding":"In zebrafish, prpf4 mutation causes DNA damage and activates the ATM/CHK2-p53 signaling pathway, arresting early erythrocytes at S and G2/M phases and inducing apoptosis; separately, prpf4 mutation causes skipped-exon splicing defects including in slc25a39 pre-mRNA, reducing slc25a39 mRNA and impairing late erythrocyte maturation.","method":"Zebrafish prpf4 mutant analysis, cell cycle assay (FACS), DNA damage markers, RNA-Seq splice-site analysis, slc25a39 expression analysis","journal":"Cell death discovery","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic loss-of-function with cell cycle FACS, DNA damage markers, and RNA-Seq splicing analysis; single lab, two distinct mechanistic readouts","pmids":["41360772"],"is_preprint":false}],"current_model":"PRPF4 (SNRNP60/HPrp4p) is a WD-repeat (seven-bladed beta-propeller) protein that integrates into the U4/U6 snRNP and U4/U6.U5 tri-snRNP via a direct interaction with PRPF3 (mediated by PRPF3's central region) and a bipartite interaction with cyclophilin H (PPIH) at its intrinsically disordered N-terminus; disruption of the PRPF4–PRPF3 interface abrogates tri-snRNP assembly and causes retinitis pigmentosa through haploinsufficiency or dominant-negative effects; in zebrafish, prpf4 loss impairs pre-mRNA splicing broadly (including slc25a39), triggers ATM/CHK2-p53-dependent DNA-damage responses arresting cell-cycle progression, and disrupts downstream Fgf, Wnt, and chemokine signaling, with a mammalian CDK-like paralog additionally shown to phosphorylate Elk-1 downstream of EGF and activate p38 MAPK/ERK cascades."},"narrative":{"mechanistic_narrative":"PRPF4 (SNRNP60/HPrp4p) is a WD-repeat protein that forms a seven-bladed beta-propeller and functions as a structural component of the U4/U6.U5 tri-snRNP, where it remains stably associated with the U4/U6 snRNP even under salt conditions that dissociate the tri-snRNP [PMID:9257651]. It is recruited to the U4/U6 snRNP through a direct interaction with PRPF3 (Hprp3p): the central region of PRPF3 is necessary and sufficient for binding PRPF4, and this interaction is required for the complex to associate with U4/U6 snRNAs [PMID:9328476, PMID:11971898]. The intrinsically disordered N-terminus of PRPF4 additionally engages cyclophilin H (PPIH) through a bipartite, two-site interaction [PMID:28935721]. The functional importance of these contacts is established by adRP-causing variants: PRPF4 p.R192H disrupts the PRPF4–PRPF3 interface and prevents tri-snRNP integration, behaving as a functional null, while p.Pro315Leu acts through dominant-negative effects on tri-snRNP component levels, together causing autosomal dominant retinitis pigmentosa via haploinsufficiency and dominant-negative mechanisms [PMID:24419317, PMID:25383878]. Consistent with a broad role in splicing, zebrafish prpf4 loss impairs spliceosome assembly and pre-mRNA splicing (including slc25a39), activates ATM/CHK2-p53-dependent DNA-damage and cell-cycle arrest responses, and disrupts downstream Fgf, Wnt/β-catenin, and chemokine signaling, leading to apoptosis in neural and erythroid lineages [PMID:30174136, PMID:41360772]. A distinct CDK-like kinase paralog (hPRP4/Prp4p) phosphorylates Elk-1 downstream of EGF and the splicing factor Prp1p, linking kinase activity to splicing and cell-cycle progression [PMID:10799319, PMID:11252721].","teleology":[{"year":1997,"claim":"Identifying PRPF4 as a salt-stable U4/U6 snRNP component established it as a core structural subunit of the tri-snRNP rather than a transient factor, and the WD-repeat architecture predicted a beta-propeller scaffold.","evidence":"Immunoaffinity purification, cDNA cloning/sequencing, and homology analysis of the human U4/U6-60kD protein","pmids":["9257651"],"confidence":"High","gaps":["Did not define which surface of the propeller mediates snRNP contacts","No direct binding partner identified at this stage"]},{"year":1997,"claim":"Demonstrating a physical PRPF4–PRPF3 interaction and PRPF3's co-precipitation with U4/U6/U5 snRNAs placed both proteins together in the tri-snRNP and pointed to PRPF3 as the link to the snRNP.","evidence":"Co-immunoprecipitation, antibody pulldown, and snRNA co-precipitation","pmids":["9328476"],"confidence":"Medium","gaps":["Did not map the minimal interaction region","Single lab, no thermodynamic quantification"]},{"year":2002,"claim":"Mapping the PRPF3 central region as necessary and sufficient for PRPF4 binding, and showing this interaction is required for U4/U6 snRNA association, established that PRPF3 recruits PRPF4 onto the snRNP.","evidence":"Deletion-mutant co-IP, isothermal titration calorimetry, and primer extension in HeLa nuclear extracts","pmids":["11971898"],"confidence":"High","gaps":["No atomic structure of the PRPF3–PRPF4 interface","Did not address PPIH contribution"]},{"year":2017,"claim":"Resolving that PPIH binds the intrinsically disordered PRPF4 N-terminus at two distinct sites defined a second, bipartite assembly contact distinct from the PRPF3 interface.","evidence":"Recombinant reconstitution, point-mutation uncoupling of binding sites, and biophysical/NMR characterization","pmids":["28935721"],"confidence":"Medium","gaps":["In vitro only; cellular requirement of each site untested","No structure of the full PRPF4–PPIH–PRPF3 module"]},{"year":2014,"claim":"Two disease variants clarified how PRPF4 dysfunction causes adRP: R192H abolishes PRPF3 binding and tri-snRNP integration (functional null/haploinsufficiency), while P315L acts dominant-negatively, linking the assembly biochemistry directly to human pathology.","evidence":"Binding/integration assays in human cells plus zebrafish overexpression, morpholino knockdown, and equivalent-mutation rescue","pmids":["25383878","24419317"],"confidence":"High","gaps":["Why a ubiquitously expressed splicing factor causes a retina-specific phenotype is unresolved","Dominant-negative mechanism not reconstituted biochemically"]},{"year":2018,"claim":"Genetic prpf4 loss in zebrafish showed that splicing failure has tissue-level consequences, coupling impaired spliceosome assembly to p53-dependent apoptosis and disrupted Fgf/Wnt/chemokine signaling.","evidence":"Gene-trap mutant with p53 rescue, RNA-Seq splicing analysis, and pathway analysis","pmids":["30174136"],"confidence":"Medium","gaps":["Whether disrupted signaling is direct or secondary to global mis-splicing unclear","Specific mis-spliced effector transcripts not defined"]},{"year":2025,"claim":"Refined zebrafish analysis connected prpf4 loss to ATM/CHK2-p53-driven cell-cycle arrest and to a specific splicing defect in slc25a39, providing concrete molecular routes from splicing failure to lineage-specific (erythroid) defects.","evidence":"prpf4 mutant cell-cycle FACS, DNA-damage markers, RNA-Seq splice-site analysis, and slc25a39 expression","pmids":["41360772"],"confidence":"Medium","gaps":["Causal link between DNA damage and which mis-spliced transcripts is not established","Relevance to mammalian/human erythropoiesis untested"]},{"year":2000,"claim":"A CDK-like kinase paralog (hPRP4) was shown to phosphorylate Elk-1 at Thr-417 downstream of EGF/forskolin, defining a signaling/transcription role separate from the WD-repeat splicing protein.","evidence":"Kinase assays in COS-7 cells with site-specific phosphorylation and Elk-1 luciferase reporter","pmids":["10799319"],"confidence":"Medium","gaps":["This concerns the CDK-like kinase paralog, not the WD-repeat PRPF4","Single study, no in vivo confirmation"]},{"year":2001,"claim":"S. pombe Prp4p kinase activity was shown to be required for splicing and to phosphorylate the splicing factor Prp1p, linking the kinase ortholog to spliceosome function and cell-cycle progression.","evidence":"In vitro kinase reconstitution with Prp1p, yeast two-hybrid, in vivo labeling, and genetic analysis","pmids":["11252721"],"confidence":"Medium","gaps":["Fission-yeast ortholog; mammalian relevance to WD-repeat PRPF4 unclear","Substrate set beyond Prp1p undefined"]},{"year":null,"claim":"How tri-snRNP assembly biochemistry translates into a retina-restricted disease phenotype, and whether the kinase-associated signaling functions reflect PRPF4 itself or a distinct paralog, remain unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No atomic-resolution structure of the PRPF4–PRPF3–PPIH assembly module","Tissue specificity of adRP unexplained","Identity/separation of WD-repeat PRPF4 versus CDK-like kinase functions not clarified within the corpus"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0005198","term_label":"structural molecule activity","supporting_discovery_ids":[0]},{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[2,8]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[0,6]}],"pathway":[{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[0,2,9]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[6,7]}],"complexes":["U4/U6.U5 tri-snRNP","U4/U6 snRNP"],"partners":["PRPF3","PPIH"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"O43172","full_name":"U4/U6 small nuclear ribonucleoprotein Prp4","aliases":["PRP4 homolog","hPrp4","U4/U6 snRNP 60 kDa protein","WD splicing factor Prp4"],"length_aa":522,"mass_kda":58.4,"function":"Plays a role in pre-mRNA splicing as component of the U4/U6-U5 tri-snRNP complex that is involved in spliceosome assembly, and as component of the precatalytic spliceosome (spliceosome B complex)","subcellular_location":"Nucleus; Nucleus speckle","url":"https://www.uniprot.org/uniprotkb/O43172/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/PRPF4","classification":"Common Essential","n_dependent_lines":1205,"n_total_lines":1208,"dependency_fraction":0.9975165562913907},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"PRPF4B","stoichiometry":4.0},{"gene":"PRPF8","stoichiometry":4.0},{"gene":"SNRNP40","stoichiometry":4.0},{"gene":"SNRPF","stoichiometry":4.0},{"gene":"CD2BP2","stoichiometry":0.2},{"gene":"CPSF6","stoichiometry":0.2},{"gene":"DDX21","stoichiometry":0.2},{"gene":"DDX39B","stoichiometry":0.2},{"gene":"EFTUD2","stoichiometry":0.2},{"gene":"RBM39","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/PRPF4","total_profiled":1310},"omim":[{"mim_id":"615922","title":"RETINITIS PIGMENTOSA 70; RP70","url":"https://www.omim.org/entry/615922"},{"mim_id":"607795","title":"PRE-mRNA-PROCESSING FACTOR 4; PRPF4","url":"https://www.omim.org/entry/607795"},{"mim_id":"607301","title":"PRE-mRNA PROCESSING FACTOR 3; PRPF3","url":"https://www.omim.org/entry/607301"},{"mim_id":"607001","title":"EUCHROMATIC HISTONE METHYLTRANSFERASE 1; EHMT1","url":"https://www.omim.org/entry/607001"},{"mim_id":"268000","title":"RETINITIS PIGMENTOSA; RP","url":"https://www.omim.org/entry/268000"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Enhanced","locations":[{"location":"Nuclear speckles","reliability":"Enhanced"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/PRPF4"},"hgnc":{"alias_symbol":["Prp4p","HPRP4","HPRP4P","PRP4","SNRNP60"],"prev_symbol":[]},"alphafold":{"accession":"O43172","domains":[{"cath_id":"-","chopping":"109-149","consensus_level":"medium","plddt":83.9846,"start":109,"end":149},{"cath_id":"2.130.10.10","chopping":"173-522","consensus_level":"high","plddt":90.0789,"start":173,"end":522}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/O43172","model_url":"https://alphafold.ebi.ac.uk/files/AF-O43172-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-O43172-F1-predicted_aligned_error_v6.png","plddt_mean":82.06},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=PRPF4","jax_strain_url":"https://www.jax.org/strain/search?query=PRPF4"},"sequence":{"accession":"O43172","fasta_url":"https://rest.uniprot.org/uniprotkb/O43172.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/O43172/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/O43172"}},"corpus_meta":[{"pmid":"24419317","id":"PMC_24419317","title":"PRPF4 mutations cause autosomal dominant retinitis pigmentosa.","date":"2014","source":"Human molecular genetics","url":"https://pubmed.ncbi.nlm.nih.gov/24419317","citation_count":108,"is_preprint":false},{"pmid":"9257651","id":"PMC_9257651","title":"The human U4/U6 snRNP contains 60 and 90kD proteins that are structurally homologous to the yeast splicing factors Prp4p and Prp3p.","date":"1997","source":"RNA (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/9257651","citation_count":62,"is_preprint":false},{"pmid":"25383878","id":"PMC_25383878","title":"Identification of a PRPF4 loss-of-function variant that abrogates U4/U6.U5 tri-snRNP integration and is associated with retinitis pigmentosa.","date":"2014","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/25383878","citation_count":36,"is_preprint":false},{"pmid":"9328476","id":"PMC_9328476","title":"Identification and characterization of human genes encoding Hprp3p and Hprp4p, interacting components of the spliceosome.","date":"1997","source":"Human molecular genetics","url":"https://pubmed.ncbi.nlm.nih.gov/9328476","citation_count":33,"is_preprint":false},{"pmid":"11252721","id":"PMC_11252721","title":"Fission yeast Prp4p kinase regulates pre-mRNA splicing by phosphorylating a non-SR-splicing factor.","date":"2001","source":"EMBO reports","url":"https://pubmed.ncbi.nlm.nih.gov/11252721","citation_count":33,"is_preprint":false},{"pmid":"11971898","id":"PMC_11971898","title":"Central region of the human splicing factor Hprp3p interacts with Hprp4p.","date":"2002","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/11971898","citation_count":28,"is_preprint":false},{"pmid":"10799319","id":"PMC_10799319","title":"Characterization of hPRP4 kinase activation: potential role in signaling.","date":"2000","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/10799319","citation_count":15,"is_preprint":false},{"pmid":"31445970","id":"PMC_31445970","title":"PRPF4 is a novel therapeutic target for the treatment of breast cancer by influencing growth, migration, invasion, and apoptosis of breast cancer cells via p38 MAPK signaling pathway.","date":"2019","source":"Molecular and cellular probes","url":"https://pubmed.ncbi.nlm.nih.gov/31445970","citation_count":15,"is_preprint":false},{"pmid":"28935721","id":"PMC_28935721","title":"The spliceosomal proteins PPIH and PRPF4 exhibit bi-partite binding.","date":"2017","source":"The Biochemical journal","url":"https://pubmed.ncbi.nlm.nih.gov/28935721","citation_count":9,"is_preprint":false},{"pmid":"30174136","id":"PMC_30174136","title":"prpf4 is essential for cell survival and posterior lateral line primordium migration in zebrafish.","date":"2018","source":"Journal of genetics and genomics = Yi chuan xue bao","url":"https://pubmed.ncbi.nlm.nih.gov/30174136","citation_count":8,"is_preprint":false},{"pmid":"31502671","id":"PMC_31502671","title":"Suppression of PRPF4 regulates pluripotency, proliferation, and differentiation in mouse embryonic stem cells.","date":"2019","source":"Cell biochemistry and function","url":"https://pubmed.ncbi.nlm.nih.gov/31502671","citation_count":6,"is_preprint":false},{"pmid":"11919719","id":"PMC_11919719","title":"The deubiquitinating enzyme Ubp21p of fission yeast stabilizes a mutant form of protein kinase Prp4p.","date":"2002","source":"Molecular genetics and genomics : MGG","url":"https://pubmed.ncbi.nlm.nih.gov/11919719","citation_count":6,"is_preprint":false},{"pmid":"39890169","id":"PMC_39890169","title":"PRPF4 Knockdown Suppresses Glioblastoma Progression via the p38 MAPK and ERK Signaling Pathways.","date":"2025","source":"Anticancer research","url":"https://pubmed.ncbi.nlm.nih.gov/39890169","citation_count":1,"is_preprint":false},{"pmid":"41360772","id":"PMC_41360772","title":"Prpf4 sequentially regulates the expansion and maturation of erythrocyte through distinct mechanisms.","date":"2025","source":"Cell death discovery","url":"https://pubmed.ncbi.nlm.nih.gov/41360772","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":9753,"output_tokens":3349,"usd":0.039747,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":10753,"output_tokens":3388,"usd":0.069233,"stage2_stop_reason":"end_turn"},"total_usd":0.10898,"stage1_batch_id":"msgbatch_01CNoDbkChHEkgrJnL7BwtGJ","stage2_batch_id":"msgbatch_016Y4x9E9Wg5VQDANj9quSQD","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1997,\n      \"finding\": \"Human U4/U6-60kD protein (PRPF4/SNRNP60) was identified as a component of the U4/U6.U5 tri-snRNP that specifically associates with U4/U6 snRNP even at salt concentrations where the tri-snRNP complex dissociates; its primary structure contains seven WD repeats, predicting a beta-propeller topology homologous to G-protein beta subunits.\",\n      \"method\": \"Immunoaffinity chromatography, cDNA cloning and sequencing, sequence homology analysis\",\n      \"journal\": \"RNA (New York, N.Y.)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal immunoaffinity purification with direct biochemical characterization; replicated by independent group (PMID:9328476)\",\n      \"pmids\": [\"9257651\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1997,\n      \"finding\": \"Hprp3p and Hprp4p interact with each other; Hprp3p co-immunoprecipitates with U4, U6, and U5 snRNAs, placing both proteins in the U4/U6.U5 tri-snRNP; the first 100 amino acids of Hprp3p are not essential for the Hprp3p–Hprp4p interaction.\",\n      \"method\": \"Co-immunoprecipitation, antibody pulldown, snRNA co-precipitation\",\n      \"journal\": \"Human molecular genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal co-IP with snRNA detection, single lab but two orthogonal methods\",\n      \"pmids\": [\"9328476\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"The central region of Hprp3p (lacking N-terminal 194 aa or C-terminal 240 aa) is necessary and sufficient for binding Hprp4p; this Hprp3p–Hprp4p interaction is required for association of the complex with U4/U6 snRNAs, indicating that Hprp3p recruits Hprp4p to the U4/U6 snRNP.\",\n      \"method\": \"Co-immunoprecipitation, isothermal titration calorimetry, primer extension analysis, bacterial and mammalian expression of deletion mutants\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro ITC (thermodynamic binding), co-IP in HeLa nuclear extracts, primer extension for functional readout; single lab but three orthogonal methods\",\n      \"pmids\": [\"11971898\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"Human hPRP4, a CDK-like kinase homologous to S. pombe Prp4p, is activated by EGF or forskolin treatment and phosphorylates Elk-1 at Thr-417 (a site distinct from other MAPKs), leading to Elk-1 transactivation; this demonstrates a signaling/transcription-factor activation function for hPRP4 beyond splicing.\",\n      \"method\": \"Kinase activity assay in COS-7 cells, EGF/forskolin stimulation, site-specific phosphorylation analysis, luciferase reporter (Elk-1 trans-activation)\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — direct kinase assay with substrate phosphorylation and reporter readout, single lab, single study; note this concerns the kinase paralog of PRPF4 (CDK-like), context is mammalian cells\",\n      \"pmids\": [\"10799319\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"S. pombe Prp4p kinase activity is required for pre-mRNA splicing in vivo; Prp4p phosphorylates the non-SR splicing factor Prp1p in vitro and interacts with Prp1p in two-hybrid assays; loss of Prp4p kinase activity impairs G1-S and G2-M cell cycle progression.\",\n      \"method\": \"In vitro kinase assay with bacterially expressed Prp1p, yeast two-hybrid, in vivo phosphorylation labeling, genetic analysis\",\n      \"journal\": \"EMBO reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro kinase reconstitution plus in vivo labeling and two-hybrid; fission yeast ortholog, single lab\",\n      \"pmids\": [\"11252721\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"The deubiquitinating enzyme Ubp21p of S. pombe stabilizes a mutant form of Prp4p kinase; Ubp21 was identified as a high-copy suppressor of a mutation in the ALKHP motif of Prp4p's kinase subdomain XI, indicating that ubiquitin-mediated proteolysis regulates Prp4p steady-state levels.\",\n      \"method\": \"High-copy suppressor screen, mutational analysis of kinase domain, Ubp21 characterization\",\n      \"journal\": \"Molecular genetics and genomics\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — genetic suppressor screen in fission yeast, single lab, limited direct biochemical validation of the ubiquitination mechanism\",\n      \"pmids\": [\"11919719\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"The PRPF4 p.Pro315Leu variant causes dominant-negative effects: overexpression in cells upregulates tri-snRNP components (PRPF4 itself included) and alters SC35 splicing-factor localization as a compensatory response; overexpression of mutant but not wild-type PRPF4 causes retinal deformities in zebrafish and worsens phenotypes in prpf4 morphants, establishing PRPF4 as a U4/U6-U5 tri-snRNP component whose dysfunction leads to adRP via haploinsufficiency and dominant-negative mechanisms.\",\n      \"method\": \"Luciferase promoter assay, overexpression in fibroblasts and HeLa cells, SC35 immunostaining, zebrafish overexpression and morpholino knockdown\",\n      \"journal\": \"Human molecular genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal methods (reporter assay, cell overexpression, in vivo zebrafish), single lab\",\n      \"pmids\": [\"24419317\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"The PRPF4 p.R192H variant disrupts the binding interface between PRPF4 and PRPF3, preventing PRPF4 from integrating into the tri-snRNP; introduction of the equivalent mutation in zebrafish prpf4 causes complete loss of function in vivo, and the variant behaves as a functional null (haploinsufficiency) in a human cell line.\",\n      \"method\": \"Biochemical binding assays (co-IP/pulldown), cell-line tri-snRNP integration assay, zebrafish in vivo functional rescue experiments\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal biochemical assays plus in vivo zebrafish validation; independently supports and extends findings from PMID:11971898 on PRPF3–PRPF4 interface\",\n      \"pmids\": [\"25383878\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"PPIH (cyclophilin H) binds the N-terminus of PRPF4 at two distinct interaction sites (bipartite binding); the N-terminal region of PRPF4 is intrinsically disordered and does not adopt secondary structure upon PPIH binding; mutations in both sites are required to abrogate complex formation.\",\n      \"method\": \"Recombinant protein expression and purification, complex formation assay, mutational analysis (point mutations uncoupling binding sites), NMR/biophysical characterization\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — in vitro reconstitution with mutagenesis, single lab, no replication\",\n      \"pmids\": [\"28935721\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"In zebrafish, prpf4 loss-of-function leads to p53-dependent apoptosis in neural cells and defects in posterior lateral line primordium (pLLP) migration; RNA-Seq shows prpf4 deficiency impairs spliceosome assembly, causes compensatory upregulation of spliceosomal genes, and alters pre-mRNA splicing; Fgf, Wnt/β-catenin, and chemokine signaling pathways are disrupted.\",\n      \"method\": \"Gene-trap zebrafish mutant, p53 rescue experiments, RNA-Seq splicing analysis, signaling pathway analysis\",\n      \"journal\": \"Journal of genetics and genomics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic loss-of-function with defined cellular and molecular phenotypes plus transcriptome-level splicing analysis; single lab\",\n      \"pmids\": [\"30174136\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"PRPF4 knockdown in breast cancer cell lines reduces proliferation, migration, invasion, and induces apoptosis; microarray analysis indicates reduced phosphorylation of p38 MAPK associated with decreased expression of PPIP5K1, PPIPK2, and YWHAE, placing PRPF4 upstream of p38 MAPK signaling.\",\n      \"method\": \"Stable shRNA knockdown in MCF7 and MDA-MB-468 cells, colony formation assay, migration/invasion assays, microarray, Western blot for p38 MAPK phosphorylation\",\n      \"journal\": \"Molecular and cellular probes\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — knockdown with phenotypic readouts and pathway inference by microarray, no direct mechanistic reconstitution; single lab\",\n      \"pmids\": [\"31445970\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"In zebrafish, prpf4 mutation causes DNA damage and activates the ATM/CHK2-p53 signaling pathway, arresting early erythrocytes at S and G2/M phases and inducing apoptosis; separately, prpf4 mutation causes skipped-exon splicing defects including in slc25a39 pre-mRNA, reducing slc25a39 mRNA and impairing late erythrocyte maturation.\",\n      \"method\": \"Zebrafish prpf4 mutant analysis, cell cycle assay (FACS), DNA damage markers, RNA-Seq splice-site analysis, slc25a39 expression analysis\",\n      \"journal\": \"Cell death discovery\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic loss-of-function with cell cycle FACS, DNA damage markers, and RNA-Seq splicing analysis; single lab, two distinct mechanistic readouts\",\n      \"pmids\": [\"41360772\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"PRPF4 (SNRNP60/HPrp4p) is a WD-repeat (seven-bladed beta-propeller) protein that integrates into the U4/U6 snRNP and U4/U6.U5 tri-snRNP via a direct interaction with PRPF3 (mediated by PRPF3's central region) and a bipartite interaction with cyclophilin H (PPIH) at its intrinsically disordered N-terminus; disruption of the PRPF4–PRPF3 interface abrogates tri-snRNP assembly and causes retinitis pigmentosa through haploinsufficiency or dominant-negative effects; in zebrafish, prpf4 loss impairs pre-mRNA splicing broadly (including slc25a39), triggers ATM/CHK2-p53-dependent DNA-damage responses arresting cell-cycle progression, and disrupts downstream Fgf, Wnt, and chemokine signaling, with a mammalian CDK-like paralog additionally shown to phosphorylate Elk-1 downstream of EGF and activate p38 MAPK/ERK cascades.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"PRPF4 (SNRNP60/HPrp4p) is a WD-repeat protein that forms a seven-bladed beta-propeller and functions as a structural component of the U4/U6.U5 tri-snRNP, where it remains stably associated with the U4/U6 snRNP even under salt conditions that dissociate the tri-snRNP [#0]. It is recruited to the U4/U6 snRNP through a direct interaction with PRPF3 (Hprp3p): the central region of PRPF3 is necessary and sufficient for binding PRPF4, and this interaction is required for the complex to associate with U4/U6 snRNAs [#1, #2]. The intrinsically disordered N-terminus of PRPF4 additionally engages cyclophilin H (PPIH) through a bipartite, two-site interaction [#8]. The functional importance of these contacts is established by adRP-causing variants: PRPF4 p.R192H disrupts the PRPF4–PRPF3 interface and prevents tri-snRNP integration, behaving as a functional null, while p.Pro315Leu acts through dominant-negative effects on tri-snRNP component levels, together causing autosomal dominant retinitis pigmentosa via haploinsufficiency and dominant-negative mechanisms [#6, #7]. Consistent with a broad role in splicing, zebrafish prpf4 loss impairs spliceosome assembly and pre-mRNA splicing (including slc25a39), activates ATM/CHK2-p53-dependent DNA-damage and cell-cycle arrest responses, and disrupts downstream Fgf, Wnt/\\u03b2-catenin, and chemokine signaling, leading to apoptosis in neural and erythroid lineages [#9, #11]. A distinct CDK-like kinase paralog (hPRP4/Prp4p) phosphorylates Elk-1 downstream of EGF and the splicing factor Prp1p, linking kinase activity to splicing and cell-cycle progression [#3, #4].\",\n  \"teleology\": [\n    {\n      \"year\": 1997,\n      \"claim\": \"Identifying PRPF4 as a salt-stable U4/U6 snRNP component established it as a core structural subunit of the tri-snRNP rather than a transient factor, and the WD-repeat architecture predicted a beta-propeller scaffold.\",\n      \"evidence\": \"Immunoaffinity purification, cDNA cloning/sequencing, and homology analysis of the human U4/U6-60kD protein\",\n      \"pmids\": [\"9257651\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define which surface of the propeller mediates snRNP contacts\", \"No direct binding partner identified at this stage\"]\n    },\n    {\n      \"year\": 1997,\n      \"claim\": \"Demonstrating a physical PRPF4–PRPF3 interaction and PRPF3's co-precipitation with U4/U6/U5 snRNAs placed both proteins together in the tri-snRNP and pointed to PRPF3 as the link to the snRNP.\",\n      \"evidence\": \"Co-immunoprecipitation, antibody pulldown, and snRNA co-precipitation\",\n      \"pmids\": [\"9328476\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Did not map the minimal interaction region\", \"Single lab, no thermodynamic quantification\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Mapping the PRPF3 central region as necessary and sufficient for PRPF4 binding, and showing this interaction is required for U4/U6 snRNA association, established that PRPF3 recruits PRPF4 onto the snRNP.\",\n      \"evidence\": \"Deletion-mutant co-IP, isothermal titration calorimetry, and primer extension in HeLa nuclear extracts\",\n      \"pmids\": [\"11971898\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No atomic structure of the PRPF3–PRPF4 interface\", \"Did not address PPIH contribution\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Resolving that PPIH binds the intrinsically disordered PRPF4 N-terminus at two distinct sites defined a second, bipartite assembly contact distinct from the PRPF3 interface.\",\n      \"evidence\": \"Recombinant reconstitution, point-mutation uncoupling of binding sites, and biophysical/NMR characterization\",\n      \"pmids\": [\"28935721\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"In vitro only; cellular requirement of each site untested\", \"No structure of the full PRPF4–PPIH–PRPF3 module\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Two disease variants clarified how PRPF4 dysfunction causes adRP: R192H abolishes PRPF3 binding and tri-snRNP integration (functional null/haploinsufficiency), while P315L acts dominant-negatively, linking the assembly biochemistry directly to human pathology.\",\n      \"evidence\": \"Binding/integration assays in human cells plus zebrafish overexpression, morpholino knockdown, and equivalent-mutation rescue\",\n      \"pmids\": [\"25383878\", \"24419317\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Why a ubiquitously expressed splicing factor causes a retina-specific phenotype is unresolved\", \"Dominant-negative mechanism not reconstituted biochemically\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Genetic prpf4 loss in zebrafish showed that splicing failure has tissue-level consequences, coupling impaired spliceosome assembly to p53-dependent apoptosis and disrupted Fgf/Wnt/chemokine signaling.\",\n      \"evidence\": \"Gene-trap mutant with p53 rescue, RNA-Seq splicing analysis, and pathway analysis\",\n      \"pmids\": [\"30174136\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether disrupted signaling is direct or secondary to global mis-splicing unclear\", \"Specific mis-spliced effector transcripts not defined\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Refined zebrafish analysis connected prpf4 loss to ATM/CHK2-p53-driven cell-cycle arrest and to a specific splicing defect in slc25a39, providing concrete molecular routes from splicing failure to lineage-specific (erythroid) defects.\",\n      \"evidence\": \"prpf4 mutant cell-cycle FACS, DNA-damage markers, RNA-Seq splice-site analysis, and slc25a39 expression\",\n      \"pmids\": [\"41360772\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Causal link between DNA damage and which mis-spliced transcripts is not established\", \"Relevance to mammalian/human erythropoiesis untested\"]\n    },\n    {\n      \"year\": 2000,\n      \"claim\": \"A CDK-like kinase paralog (hPRP4) was shown to phosphorylate Elk-1 at Thr-417 downstream of EGF/forskolin, defining a signaling/transcription role separate from the WD-repeat splicing protein.\",\n      \"evidence\": \"Kinase assays in COS-7 cells with site-specific phosphorylation and Elk-1 luciferase reporter\",\n      \"pmids\": [\"10799319\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"This concerns the CDK-like kinase paralog, not the WD-repeat PRPF4\", \"Single study, no in vivo confirmation\"]\n    },\n    {\n      \"year\": 2001,\n      \"claim\": \"S. pombe Prp4p kinase activity was shown to be required for splicing and to phosphorylate the splicing factor Prp1p, linking the kinase ortholog to spliceosome function and cell-cycle progression.\",\n      \"evidence\": \"In vitro kinase reconstitution with Prp1p, yeast two-hybrid, in vivo labeling, and genetic analysis\",\n      \"pmids\": [\"11252721\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Fission-yeast ortholog; mammalian relevance to WD-repeat PRPF4 unclear\", \"Substrate set beyond Prp1p undefined\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How tri-snRNP assembly biochemistry translates into a retina-restricted disease phenotype, and whether the kinase-associated signaling functions reflect PRPF4 itself or a distinct paralog, remain unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No atomic-resolution structure of the PRPF4–PRPF3–PPIH assembly module\", \"Tissue specificity of adRP unexplained\", \"Identity/separation of WD-repeat PRPF4 versus CDK-like kinase functions not clarified within the corpus\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [0]},\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [2, 8]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [0, 6]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [0, 2, 9]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [6, 7]}\n    ],\n    \"complexes\": [\n      \"U4/U6.U5 tri-snRNP\",\n      \"U4/U6 snRNP\"\n    ],\n    \"partners\": [\n      \"PRPF3\",\n      \"PPIH\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}