{"gene":"PRPF40B","run_date":"2026-06-10T06:43:36","timeline":{"discoveries":[{"year":2015,"finding":"PRPF40B directly interacts with the splicing factor SF1 and associates with U2AF65, colocalizing with these factors in nuclear speckles, and modulates alternative pre-mRNA splice site selection in vivo.","method":"Co-immunoprecipitation, direct interaction assays, colocalization by immunofluorescence, splicing reporter minigene assays, siRNA knockdown","journal":"RNA (New York, N.Y.)","confidence":"High","confidence_rationale":"Tier 2 / Moderate — reciprocal interaction assays demonstrating direct binding to SF1 and association with U2AF65, combined with colocalization and functional splicing assays in a single study","pmids":["25605964"],"is_preprint":false},{"year":2015,"finding":"PRPF40B is highly enriched in nuclear speckles, displaying localization behavior similar to established splicing factors, and this localization is functionally coupled to its role in splice site regulation.","method":"Immunofluorescence microscopy and colocalization with splicing factor markers","journal":"RNA (New York, N.Y.)","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — single-lab localization experiment tied to functional splicing assays but not FRAP or fractionation","pmids":["25605964"],"is_preprint":false},{"year":2015,"finding":"PRPF40B depletion increases Fas/CD95 receptor number and promotes cell apoptosis, demonstrating that PRPF40B regulates alternative splicing of apoptotic genes to control cell survival; weak 5' and 3' splice sites and exonic sequences are required for PRPF40B function at the Fas locus.","method":"siRNA knockdown, splicing reporter minigene assays, flow cytometry for Fas surface expression, apoptosis assays","journal":"RNA (New York, N.Y.)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function with defined cellular phenotype and mechanistic splice-site requirements determined, single lab","pmids":["25605964"],"is_preprint":false},{"year":2019,"finding":"PRPF40B knockout in K562 cells causes a net increase in exon inclusion among hundreds of alternative splicing events, indicating that PRPF40B primarily acts as a splicing repressor; regulated events are enriched for A-rich downstream intronic motifs and weak 5' splice sites.","method":"CRISPR/Cas9 knockout, RNA-seq, rescue with wild-type and MDS mutant alleles","journal":"RNA (New York, N.Y.)","confidence":"High","confidence_rationale":"Tier 2 / Moderate — full KO with RNA-seq and allelic rescue using multiple variants, identifying mechanistic sequence features; single lab but multiple orthogonal approaches","pmids":["31088860"],"is_preprint":false},{"year":2019,"finding":"Loss of PRPF40B in K562 cells induces a KLF1 transcriptional signature involving iron metabolism and hypoxia-related pathways (including cholesterol biosynthesis and Akt/MAPK signaling), suggesting PRPF40B represses hypoxia-associated gene expression in myeloid cells.","method":"CRISPR/Cas9 knockout, RNA-seq gene expression analysis, rescue experiments","journal":"RNA (New York, N.Y.)","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — clean KO with defined transcriptional phenotype, single lab, no direct molecular mechanism for the hypoxia repression established","pmids":["31088860"],"is_preprint":false},{"year":2019,"finding":"PRPF40B-regulated alternative splicing events are likely cotranscriptional, consistent with its ortholog Prp40 being part of the U1 snRNP.","method":"RNA-seq analysis of splice-site features in PRPF40B KO cells","journal":"RNA (New York, N.Y.)","confidence":"Low","confidence_rationale":"Tier 4 / Weak — inferred from sequence feature analysis, not directly demonstrated by cotranscriptional splicing assay","pmids":["31088860"],"is_preprint":false},{"year":2025,"finding":"PRPF40B promotes production of the full-length TRKB (TRKB-FL) isoform over the truncated TRKB-T1 isoform during neuronal differentiation by regulating alternative splicing of NTRK2; silencing PRPF40B shifts the balance toward TRKB-T1 and impairs neuronal differentiation and synaptic plasticity gene expression both in vitro and in vivo during early embryogenesis.","method":"siRNA knockdown, in vitro neuronal differentiation assays, in vivo embryogenesis models, RT-PCR/RNA-seq for isoform quantification","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function with defined molecular (isoform switch) and cellular phenotype validated in multiple model systems, single lab","pmids":["41360765"],"is_preprint":false},{"year":2025,"finding":"Human PRPF40B (and its paralog PRPF40A) are not integral components of U1 snRNP, in contrast to their yeast ortholog Prp40 which is stably associated with U1 snRNP; instead, human PRPF40B functions as an alternative splicing factor.","method":"Comparative biochemical analysis and cryo-EM/structural context from yeast Prp40 studies; functional annotation from published literature reviewed in the context of yeast experiments","journal":"bioRxiv","confidence":"Low","confidence_rationale":"Tier 3 / Weak — claim about human PRPF40B is an inference from yeast comparative data in a preprint, not directly demonstrated for the human protein","pmids":[],"is_preprint":true}],"current_model":"PRPF40B is a nuclear splicing factor that localizes to nuclear speckles, directly interacts with SF1 and associates with U2AF65, and primarily acts as a repressor of exon inclusion for hundreds of alternative splicing targets by recognizing weak splice sites with A-rich downstream intronic motifs; functionally, it regulates alternative splicing of apoptotic genes (including Fas/CD95), NTRK2 (controlling TRKB isoform balance during neuronal differentiation), and hypoxia-related genes in myeloid cells."},"narrative":{"mechanistic_narrative":"PRPF40B is a nuclear splicing factor that regulates alternative pre-mRNA splice site selection and controls splicing-dependent cell fate decisions [PMID:25605964, PMID:31088860]. It directly binds the early spliceosome assembly factor SF1 and associates with U2AF65, colocalizing with these factors in nuclear speckles, where its enrichment is coupled to its splicing-regulatory function [PMID:25605964]. Genome-wide loss-of-function defines PRPF40B as predominantly a splicing repressor: its depletion produces a net increase in exon inclusion across hundreds of events, and the regulated targets are enriched for weak 5' splice sites and A-rich downstream intronic motifs, indicating recognition of suboptimal splice sites [PMID:31088860]. Through this activity PRPF40B shapes specific cell-fate programs — it represses Fas/CD95 splicing such that its loss raises Fas surface expression and promotes apoptosis [PMID:25605964], directs the NTRK2 isoform balance toward full-length TRKB (TRKB-FL) over the truncated TRKB-T1 form to support neuronal differentiation and synaptic plasticity gene expression [PMID:41360765], and restrains a KLF1-associated transcriptional program linked to iron metabolism and hypoxia in myeloid cells [PMID:31088860].","teleology":[{"year":2015,"claim":"Establishing PRPF40B's molecular partners and subcellular context was needed to place it within spliceosome assembly; demonstrating direct SF1 binding, U2AF65 association, and nuclear speckle localization positioned it as a bona fide splice-site regulator.","evidence":"Co-immunoprecipitation, direct interaction assays, immunofluorescence colocalization, and minigene splicing reporters with siRNA knockdown","pmids":["25605964"],"confidence":"High","gaps":["Structural basis of the SF1 interaction not resolved","Whether U2AF65 association is direct or bridged is undefined","Localization tied to function but not tested by FRAP or fractionation"]},{"year":2015,"claim":"Connecting PRPF40B to a cellular outcome showed that its splicing activity is functionally consequential; depletion raised Fas/CD95 receptor levels and promoted apoptosis, with weak splice sites and exonic sequences required at the Fas locus.","evidence":"siRNA knockdown, splicing minigene reporters, flow cytometry for Fas, and apoptosis assays","pmids":["25605964"],"confidence":"Medium","gaps":["Direct binding of PRPF40B to the Fas pre-mRNA not demonstrated","Single lab and single cellular context"]},{"year":2019,"claim":"The directionality and sequence logic of PRPF40B regulation were unknown; transcriptome-wide knockout showed it is primarily a repressor of exon inclusion that recognizes weak 5' splice sites and A-rich downstream intronic motifs.","evidence":"CRISPR/Cas9 knockout in K562 with RNA-seq and rescue using wild-type and MDS mutant alleles","pmids":["31088860"],"confidence":"High","gaps":["Direct RNA-binding map (e.g., CLIP) not generated","Mechanism by which weak splice sites are repressed not resolved","Functional impact of MDS mutant alleles on splicing outcomes only partly characterized"]},{"year":2019,"claim":"Whether PRPF40B loss had broader gene-expression consequences was untested; knockout induced a KLF1 signature spanning iron metabolism and hypoxia-related pathways, implicating it in myeloid transcriptional programs.","evidence":"CRISPR/Cas9 knockout with RNA-seq gene expression and rescue analysis","pmids":["31088860"],"confidence":"Medium","gaps":["No direct molecular mechanism linking PRPF40B to hypoxia gene repression","Whether the effect is splicing-driven or secondary is unresolved"]},{"year":2025,"claim":"A developmental role for PRPF40B splicing control was unknown; it was shown to favor the TRKB-FL over TRKB-T1 isoform of NTRK2, with silencing impairing neuronal differentiation and synaptic plasticity gene expression in vitro and in vivo.","evidence":"siRNA knockdown, in vitro neuronal differentiation, in vivo embryogenesis models, and RT-PCR/RNA-seq isoform quantification","pmids":["41360765"],"confidence":"Medium","gaps":["Direct binding of PRPF40B to NTRK2 pre-mRNA not shown","Single lab","Mechanistic link between isoform switch and the differentiation phenotype not fully dissected"]},{"year":null,"claim":"It remains unresolved how human PRPF40B associates with the spliceosome relative to its yeast ortholog Prp40 and whether its regulation is genuinely cotranscriptional.","evidence":"Comparative inference from yeast Prp40 structural/biochemical context; not directly demonstrated for human PRPF40B","pmids":[],"confidence":"Low","gaps":["Whether human PRPF40B is a U1 snRNP component is inferred, not directly tested","Cotranscriptional coupling inferred from sequence features, not measured directly","No direct RNA-binding or recruitment mechanism established"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0003723","term_label":"RNA binding","supporting_discovery_ids":[3]}],"localization":[{"term_id":"GO:0005654","term_label":"nucleoplasm","supporting_discovery_ids":[0,1]}],"pathway":[{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[0,3]}],"complexes":[],"partners":["SF1","U2AF65"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q6NWY9","full_name":"Pre-mRNA-processing factor 40 homolog B","aliases":["Huntingtin yeast partner C","Huntingtin-interacting protein C"],"length_aa":871,"mass_kda":99.4,"function":"May be involved in pre-mRNA splicing","subcellular_location":"Nucleus speckle","url":"https://www.uniprot.org/uniprotkb/Q6NWY9/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/PRPF40B","classification":"Not Classified","n_dependent_lines":7,"n_total_lines":1208,"dependency_fraction":0.005794701986754967},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/PRPF40B","total_profiled":1310},"omim":[{"mim_id":"621019","title":"PRE-mRNA-PROCESSING FACTOR 40 HOMOLOG B; PRPF40B","url":"https://www.omim.org/entry/621019"},{"mim_id":"612568","title":"SPIC TRANSCRIPTION FACTOR; SPIC","url":"https://www.omim.org/entry/612568"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Nuclear bodies","reliability":"Approved"},{"location":"Nucleoplasm","reliability":"Additional"},{"location":"Aggresome","reliability":"Additional"},{"location":"Cytosol","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in many","driving_tissues":[],"url":"https://www.proteinatlas.org/search/PRPF40B"},"hgnc":{"alias_symbol":["HYPC"],"prev_symbol":[]},"alphafold":{"accession":"Q6NWY9","domains":[{"cath_id":"-","chopping":"99-177","consensus_level":"medium","plddt":79.1653,"start":99,"end":177},{"cath_id":"-","chopping":"314-422","consensus_level":"medium","plddt":92.063,"start":314,"end":422},{"cath_id":"-","chopping":"481-616","consensus_level":"medium","plddt":93.7079,"start":481,"end":616},{"cath_id":"1.10.10.440","chopping":"618-688","consensus_level":"medium","plddt":87.8301,"start":618,"end":688}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q6NWY9","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q6NWY9-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q6NWY9-F1-predicted_aligned_error_v6.png","plddt_mean":67.75},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=PRPF40B","jax_strain_url":"https://www.jax.org/strain/search?query=PRPF40B"},"sequence":{"accession":"Q6NWY9","fasta_url":"https://rest.uniprot.org/uniprotkb/Q6NWY9.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q6NWY9/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q6NWY9"}},"corpus_meta":[{"pmid":"9485446","id":"PMC_9485446","title":"Interaction of the hydrogenase accessory protein HypC with HycE, the large subunit of Escherichia coli hydrogenase 3 during enzyme maturation.","date":"1998","source":"Biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/9485446","citation_count":90,"is_preprint":false},{"pmid":"15504408","id":"PMC_15504408","title":"The complex between hydrogenase-maturation proteins HypC and HypD is an intermediate in the supply of cyanide to the active site iron of [NiFe]-hydrogenases.","date":"2004","source":"Journal of molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/15504408","citation_count":88,"is_preprint":false},{"pmid":"12441107","id":"PMC_12441107","title":"Maturation of [NiFe]-hydrogenases in Escherichia coli: the HypC cycle.","date":"2002","source":"Journal of molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/12441107","citation_count":79,"is_preprint":false},{"pmid":"10783387","id":"PMC_10783387","title":"Analysis of the HypC-hycE complex, a key intermediate in the assembly of the metal center of the Escherichia coli hydrogenase 3.","date":"2000","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/10783387","citation_count":73,"is_preprint":false},{"pmid":"11292801","id":"PMC_11292801","title":"Interplay between the specific chaperone-like proteins HybG and HypC in maturation of hydrogenases 1, 2, and 3 from Escherichia coli.","date":"2001","source":"Journal of bacteriology","url":"https://pubmed.ncbi.nlm.nih.gov/11292801","citation_count":54,"is_preprint":false},{"pmid":"20348292","id":"PMC_20348292","title":"HypC, the anthrone oxidase involved in aflatoxin biosynthesis.","date":"2010","source":"Applied and environmental microbiology","url":"https://pubmed.ncbi.nlm.nih.gov/20348292","citation_count":45,"is_preprint":false},{"pmid":"25605964","id":"PMC_25605964","title":"Prp40 pre-mRNA processing factor 40 homolog B (PRPF40B) associates with SF1 and U2AF65 and modulates alternative pre-mRNA splicing in vivo.","date":"2015","source":"RNA (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/25605964","citation_count":30,"is_preprint":false},{"pmid":"14726233","id":"PMC_14726233","title":"Requirement of hydD, hydE, hypC and hypE genes for hydrogenase activity in Helicobacter pylori.","date":"2004","source":"Microbial pathogenesis","url":"https://pubmed.ncbi.nlm.nih.gov/14726233","citation_count":25,"is_preprint":false},{"pmid":"20023111","id":"PMC_20023111","title":"CalA, a cyanobacterial AbrB protein, interacts with the upstream region of hypC and acts as a repressor of its transcription in the cyanobacterium Nostoc sp. strain PCC 7120.","date":"2009","source":"Applied and environmental microbiology","url":"https://pubmed.ncbi.nlm.nih.gov/20023111","citation_count":19,"is_preprint":false},{"pmid":"31088860","id":"PMC_31088860","title":"Human PRPF40B regulates hundreds of alternative splicing targets and represses a hypoxia expression signature.","date":"2019","source":"RNA (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/31088860","citation_count":17,"is_preprint":false},{"pmid":"26364315","id":"PMC_26364315","title":"Heterologous complementation studies in Escherichia coli with the Hyp accessory protein machinery from Chloroflexi provide insight into [NiFe]-hydrogenase large subunit recognition by the HypC protein family.","date":"2015","source":"Microbiology (Reading, England)","url":"https://pubmed.ncbi.nlm.nih.gov/26364315","citation_count":9,"is_preprint":false},{"pmid":"17669368","id":"PMC_17669368","title":"Solution structure of Escherichia coli HypC.","date":"2007","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/17669368","citation_count":9,"is_preprint":false},{"pmid":"9358044","id":"PMC_9358044","title":"The sequences of hypF, hypC and hypD complete the hyp gene cluster required for hydrogenase activity in Bradyrhizobium japonicum.","date":"1997","source":"Gene","url":"https://pubmed.ncbi.nlm.nih.gov/9358044","citation_count":8,"is_preprint":false},{"pmid":"35271275","id":"PMC_35271275","title":"[NiFe] Hydrogenase Accessory Proteins HypB-HypC Accelerate Proton Conversion to Enhance the Acid Resistance and d-Lactic Acid Production of Escherichia coli.","date":"2022","source":"ACS synthetic biology","url":"https://pubmed.ncbi.nlm.nih.gov/35271275","citation_count":6,"is_preprint":false},{"pmid":"24942742","id":"PMC_24942742","title":"Maturation of Rhizobium leguminosarum hydrogenase in the presence of oxygen requires the interaction of the chaperone HypC and the scaffolding protein HupK.","date":"2014","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/24942742","citation_count":5,"is_preprint":false},{"pmid":"17554182","id":"PMC_17554182","title":"Crystallization and preliminary X-ray crystallographic studies of the [NiFe] hydrogenase maturation proteins HypC and HypD.","date":"2007","source":"Acta crystallographica. Section F, Structural biology and crystallization communications","url":"https://pubmed.ncbi.nlm.nih.gov/17554182","citation_count":5,"is_preprint":false},{"pmid":"24094065","id":"PMC_24094065","title":"Computational study of the Fe(CN)2CO cofactor and its binding to HypC protein.","date":"2013","source":"The journal of physical chemistry. B","url":"https://pubmed.ncbi.nlm.nih.gov/24094065","citation_count":5,"is_preprint":false},{"pmid":"29996137","id":"PMC_29996137","title":"The Extended C-Terminal α-Helix of the HypC Chaperone Restricts Recognition of Large Subunit Precursors by the Hyp-Scaffold Machinery during [NiFe]-Hydrogenase Maturation in Escherichia coli.","date":"2018","source":"Journal of molecular microbiology and biotechnology","url":"https://pubmed.ncbi.nlm.nih.gov/29996137","citation_count":2,"is_preprint":false},{"pmid":"41360765","id":"PMC_41360765","title":"Regulation of NTRK2 alternative splicing by PRPF40B controls neural differentiation and synaptic plasticity.","date":"2025","source":"Cell death & disease","url":"https://pubmed.ncbi.nlm.nih.gov/41360765","citation_count":1,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.04.07.647526","title":"Regulation of  <i>NTRK2</i>  alternative splicing by PRPF40B controls neural differentiation and synaptic plasticity","date":"2025-04-07","source":"bioRxiv","url":"https://doi.org/10.1101/2025.04.07.647526","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2025.08.28.672894","title":"U1 snRNP and RNA polymerase II interaction is predominantly mediated by Prp40 rather than U1-70K in yeast","date":"2025-08-30","source":"bioRxiv","url":"https://doi.org/10.1101/2025.08.28.672894","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":11704,"output_tokens":1846,"usd":0.031401,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":8714,"output_tokens":2242,"usd":0.04981,"stage2_stop_reason":"end_turn"},"total_usd":0.081211,"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\": 2015,\n      \"finding\": \"PRPF40B directly interacts with the splicing factor SF1 and associates with U2AF65, colocalizing with these factors in nuclear speckles, and modulates alternative pre-mRNA splice site selection in vivo.\",\n      \"method\": \"Co-immunoprecipitation, direct interaction assays, colocalization by immunofluorescence, splicing reporter minigene assays, siRNA knockdown\",\n      \"journal\": \"RNA (New York, N.Y.)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal interaction assays demonstrating direct binding to SF1 and association with U2AF65, combined with colocalization and functional splicing assays in a single study\",\n      \"pmids\": [\"25605964\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"PRPF40B is highly enriched in nuclear speckles, displaying localization behavior similar to established splicing factors, and this localization is functionally coupled to its role in splice site regulation.\",\n      \"method\": \"Immunofluorescence microscopy and colocalization with splicing factor markers\",\n      \"journal\": \"RNA (New York, N.Y.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — single-lab localization experiment tied to functional splicing assays but not FRAP or fractionation\",\n      \"pmids\": [\"25605964\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"PRPF40B depletion increases Fas/CD95 receptor number and promotes cell apoptosis, demonstrating that PRPF40B regulates alternative splicing of apoptotic genes to control cell survival; weak 5' and 3' splice sites and exonic sequences are required for PRPF40B function at the Fas locus.\",\n      \"method\": \"siRNA knockdown, splicing reporter minigene assays, flow cytometry for Fas surface expression, apoptosis assays\",\n      \"journal\": \"RNA (New York, N.Y.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function with defined cellular phenotype and mechanistic splice-site requirements determined, single lab\",\n      \"pmids\": [\"25605964\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"PRPF40B knockout in K562 cells causes a net increase in exon inclusion among hundreds of alternative splicing events, indicating that PRPF40B primarily acts as a splicing repressor; regulated events are enriched for A-rich downstream intronic motifs and weak 5' splice sites.\",\n      \"method\": \"CRISPR/Cas9 knockout, RNA-seq, rescue with wild-type and MDS mutant alleles\",\n      \"journal\": \"RNA (New York, N.Y.)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — full KO with RNA-seq and allelic rescue using multiple variants, identifying mechanistic sequence features; single lab but multiple orthogonal approaches\",\n      \"pmids\": [\"31088860\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Loss of PRPF40B in K562 cells induces a KLF1 transcriptional signature involving iron metabolism and hypoxia-related pathways (including cholesterol biosynthesis and Akt/MAPK signaling), suggesting PRPF40B represses hypoxia-associated gene expression in myeloid cells.\",\n      \"method\": \"CRISPR/Cas9 knockout, RNA-seq gene expression analysis, rescue experiments\",\n      \"journal\": \"RNA (New York, N.Y.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — clean KO with defined transcriptional phenotype, single lab, no direct molecular mechanism for the hypoxia repression established\",\n      \"pmids\": [\"31088860\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"PRPF40B-regulated alternative splicing events are likely cotranscriptional, consistent with its ortholog Prp40 being part of the U1 snRNP.\",\n      \"method\": \"RNA-seq analysis of splice-site features in PRPF40B KO cells\",\n      \"journal\": \"RNA (New York, N.Y.)\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 4 / Weak — inferred from sequence feature analysis, not directly demonstrated by cotranscriptional splicing assay\",\n      \"pmids\": [\"31088860\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"PRPF40B promotes production of the full-length TRKB (TRKB-FL) isoform over the truncated TRKB-T1 isoform during neuronal differentiation by regulating alternative splicing of NTRK2; silencing PRPF40B shifts the balance toward TRKB-T1 and impairs neuronal differentiation and synaptic plasticity gene expression both in vitro and in vivo during early embryogenesis.\",\n      \"method\": \"siRNA knockdown, in vitro neuronal differentiation assays, in vivo embryogenesis models, RT-PCR/RNA-seq for isoform quantification\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function with defined molecular (isoform switch) and cellular phenotype validated in multiple model systems, single lab\",\n      \"pmids\": [\"41360765\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Human PRPF40B (and its paralog PRPF40A) are not integral components of U1 snRNP, in contrast to their yeast ortholog Prp40 which is stably associated with U1 snRNP; instead, human PRPF40B functions as an alternative splicing factor.\",\n      \"method\": \"Comparative biochemical analysis and cryo-EM/structural context from yeast Prp40 studies; functional annotation from published literature reviewed in the context of yeast experiments\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — claim about human PRPF40B is an inference from yeast comparative data in a preprint, not directly demonstrated for the human protein\",\n      \"pmids\": [],\n      \"is_preprint\": true\n    }\n  ],\n  \"current_model\": \"PRPF40B is a nuclear splicing factor that localizes to nuclear speckles, directly interacts with SF1 and associates with U2AF65, and primarily acts as a repressor of exon inclusion for hundreds of alternative splicing targets by recognizing weak splice sites with A-rich downstream intronic motifs; functionally, it regulates alternative splicing of apoptotic genes (including Fas/CD95), NTRK2 (controlling TRKB isoform balance during neuronal differentiation), and hypoxia-related genes in myeloid cells.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"PRPF40B is a nuclear splicing factor that regulates alternative pre-mRNA splice site selection and controls splicing-dependent cell fate decisions [#0, #3]. It directly binds the early spliceosome assembly factor SF1 and associates with U2AF65, colocalizing with these factors in nuclear speckles, where its enrichment is coupled to its splicing-regulatory function [#0, #1]. Genome-wide loss-of-function defines PRPF40B as predominantly a splicing repressor: its depletion produces a net increase in exon inclusion across hundreds of events, and the regulated targets are enriched for weak 5' splice sites and A-rich downstream intronic motifs, indicating recognition of suboptimal splice sites [#3]. Through this activity PRPF40B shapes specific cell-fate programs — it represses Fas/CD95 splicing such that its loss raises Fas surface expression and promotes apoptosis [#2], directs the NTRK2 isoform balance toward full-length TRKB (TRKB-FL) over the truncated TRKB-T1 form to support neuronal differentiation and synaptic plasticity gene expression [#6], and restrains a KLF1-associated transcriptional program linked to iron metabolism and hypoxia in myeloid cells [#4].\",\n  \"teleology\": [\n    {\n      \"year\": 2015,\n      \"claim\": \"Establishing PRPF40B's molecular partners and subcellular context was needed to place it within spliceosome assembly; demonstrating direct SF1 binding, U2AF65 association, and nuclear speckle localization positioned it as a bona fide splice-site regulator.\",\n      \"evidence\": \"Co-immunoprecipitation, direct interaction assays, immunofluorescence colocalization, and minigene splicing reporters with siRNA knockdown\",\n      \"pmids\": [\"25605964\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Structural basis of the SF1 interaction not resolved\",\n        \"Whether U2AF65 association is direct or bridged is undefined\",\n        \"Localization tied to function but not tested by FRAP or fractionation\"\n      ]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Connecting PRPF40B to a cellular outcome showed that its splicing activity is functionally consequential; depletion raised Fas/CD95 receptor levels and promoted apoptosis, with weak splice sites and exonic sequences required at the Fas locus.\",\n      \"evidence\": \"siRNA knockdown, splicing minigene reporters, flow cytometry for Fas, and apoptosis assays\",\n      \"pmids\": [\"25605964\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Direct binding of PRPF40B to the Fas pre-mRNA not demonstrated\",\n        \"Single lab and single cellular context\"\n      ]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"The directionality and sequence logic of PRPF40B regulation were unknown; transcriptome-wide knockout showed it is primarily a repressor of exon inclusion that recognizes weak 5' splice sites and A-rich downstream intronic motifs.\",\n      \"evidence\": \"CRISPR/Cas9 knockout in K562 with RNA-seq and rescue using wild-type and MDS mutant alleles\",\n      \"pmids\": [\"31088860\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Direct RNA-binding map (e.g., CLIP) not generated\",\n        \"Mechanism by which weak splice sites are repressed not resolved\",\n        \"Functional impact of MDS mutant alleles on splicing outcomes only partly characterized\"\n      ]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Whether PRPF40B loss had broader gene-expression consequences was untested; knockout induced a KLF1 signature spanning iron metabolism and hypoxia-related pathways, implicating it in myeloid transcriptional programs.\",\n      \"evidence\": \"CRISPR/Cas9 knockout with RNA-seq gene expression and rescue analysis\",\n      \"pmids\": [\"31088860\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"No direct molecular mechanism linking PRPF40B to hypoxia gene repression\",\n        \"Whether the effect is splicing-driven or secondary is unresolved\"\n      ]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"A developmental role for PRPF40B splicing control was unknown; it was shown to favor the TRKB-FL over TRKB-T1 isoform of NTRK2, with silencing impairing neuronal differentiation and synaptic plasticity gene expression in vitro and in vivo.\",\n      \"evidence\": \"siRNA knockdown, in vitro neuronal differentiation, in vivo embryogenesis models, and RT-PCR/RNA-seq isoform quantification\",\n      \"pmids\": [\"41360765\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Direct binding of PRPF40B to NTRK2 pre-mRNA not shown\",\n        \"Single lab\",\n        \"Mechanistic link between isoform switch and the differentiation phenotype not fully dissected\"\n      ]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"It remains unresolved how human PRPF40B associates with the spliceosome relative to its yeast ortholog Prp40 and whether its regulation is genuinely cotranscriptional.\",\n      \"evidence\": \"Comparative inference from yeast Prp40 structural/biochemical context; not directly demonstrated for human PRPF40B\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\n        \"Whether human PRPF40B is a U1 snRNP component is inferred, not directly tested\",\n        \"Cotranscriptional coupling inferred from sequence features, not measured directly\",\n        \"No direct RNA-binding or recruitment mechanism established\"\n      ]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0003723\", \"supporting_discovery_ids\": [3]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005654\", \"supporting_discovery_ids\": [0, 1]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [0, 3]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"SF1\", \"U2AF65\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"faith_supported":4,"faith_total":4,"faith_pct":100.0}}