{"gene":"SF3B6","run_date":"2026-06-10T07:46:31","timeline":{"discoveries":[{"year":2021,"finding":"SF3B6 stabilizes the branch site (BS):U2 snRNA duplex within the substrate-bound U2 snRNP, which could aid binding of introns with poor sequence complementarity to U2 snRNA. High-resolution cryo-EM structures (2.0–2.2 Å) of human 17S U2 snRNP during ATP-dependent remodeling and pre-mRNA binding provided direct snapshots of SF3B6's role in branch site selection.","method":"Cryo-EM structure determination (2.0–2.2 Å resolution) of human 17S U2 snRNP reconstituted in vitro with ATP-dependent remodeling and pre-mRNA substrate binding","journal":"Science","confidence":"High","confidence_rationale":"Tier 1 / Strong — high-resolution cryo-EM structures with in vitro reconstitution of U2 snRNP remodeling and substrate binding, multiple structural snapshots providing direct mechanistic insight","pmids":["34822310"],"is_preprint":false},{"year":2006,"finding":"SF3b14a/p14 (SF3B6) contacts U2 snRNA near the branch site-interacting region (BSiR) in purified human 17S U2 snRNPs, and these RNA-protein interactions are dynamic — changing between 17S U2 snRNP and spliceosomal A and B complexes.","method":"Lead(II)-induced RNA cleavage and UV cross-linking of immunoaffinity-purified human 17S U2 snRNPs, compared across spliceosomal A and B complexes","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal UV cross-linking and chemical probing on purified native complexes; findings replicated across multiple spliceosomal assembly states","pmids":["16537922"],"is_preprint":false},{"year":2007,"finding":"p14/SF3b14a (SF3B6) directly contacts the branch site-interacting region (BSiR) of U2 snRNA, as identified by MS analysis of UV cross-linked peptide-RNA conjugates from a partial U2 snRNP complex.","method":"UV cross-linking combined with IMAC enrichment and MALDI-MS/MS analysis of peptide-RNA heteroconjugates from purified partial U2 snRNP complex","journal":"Nucleic acids research","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — direct biochemical identification of contact site by MS, but single lab, no mutagenesis validation reported in abstract","pmids":["17652325"],"is_preprint":false},{"year":2021,"finding":"SF3B6 interacts with the N-terminal extension of SF3B1 and, through computational structural and normal mode analyses, promotes a functionally relevant 'open-to-close' conformational transition in SF3B1 by enhancing concerted residue motions, thereby acting as an allosteric regulator of SF3B1 for branch point sequence (BPS) selection. SF3B6 influences motions of 16 SF3B1 residues that interact with U2 snRNA/branchpoint duplex.","method":"Comparative homology modeling, anisotropic network model (ANM) normal mode analysis, and inter-residue correlated motion analysis of human vs. yeast SF3b complexes","journal":"Current research in structural biology","confidence":"Low","confidence_rationale":"Tier 4 / Weak — purely computational analysis (normal mode modeling), no experimental validation reported; functional inference is speculative","pmids":["35028595"],"is_preprint":false},{"year":2024,"finding":"In the human spliceosomal B complex, SF3B6 was localized to a previously unknown position, revealing new molecular contacts and functions during pre-mRNA splicing at the B complex stage.","method":"Cryo-EM structure determination of dimerized human spliceosomal B complexes formed in the presence of ATP-γS, at improved resolution","journal":"The EMBO journal","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — high-resolution cryo-EM structural localization, single study, no mutagenesis or functional validation of SF3B6-specific contacts reported in abstract","pmids":["38383864"],"is_preprint":false},{"year":2022,"finding":"An additional SF3B6 subunit had no detectable effect on U2AF2-SF3B1 binding affinities, as measured by quantitative binding assays.","method":"Quantitative binding affinity measurements (fluorescence anisotropy or ITC) of U2AF2 UHM with SF3B1 ULM in the presence or absence of SF3B6","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — direct quantitative binding assay with crystal structure context, but this is a negative result (SF3B6 had no effect), single lab","pmids":["35780835"],"is_preprint":false},{"year":2021,"finding":"The lncRNA DKFZp434J0226 physically interacts with SF3B6 and promotes its phosphorylation, which further regulates alternative splicing of pre-mRNA in pancreatic ductal adenocarcinoma (PDAC) cells.","method":"RNA pull-down, RNA immunoprecipitation (RIP), immunofluorescence, and western blotting in PDAC cells; tumor xenograft in vivo assays","journal":"Molecular medicine","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, RNA pull-down and RIP without mutagenesis; phosphorylation claim is not directly validated with kinase identification or phosphosite mapping in abstract","pmids":["34470609"],"is_preprint":false},{"year":2023,"finding":"In fibroblasts with PHF5A loss-of-function (LOF) variants, SF3B6 protein levels and SF3B complex formation were unaffected, suggesting SF3B6 protein stability does not require PHF5A and that compensatory mechanisms maintain normal SF3B component levels.","method":"Western blotting and SF3B complex analysis in subject-derived fibroblasts with PHF5A LOF variants","journal":"Genetics in medicine","confidence":"Low","confidence_rationale":"Tier 3 / Weak — indirect finding about SF3B6, single lab, limited mechanistic follow-up for SF3B6 specifically","pmids":["37422718"],"is_preprint":false},{"year":2025,"finding":"SF3B6 knockdown in MDA-MB-231 breast cancer cells inhibits cellular proliferation and migration and increases apoptosis. iRIP-seq demonstrated that SF3B6 directly binds target mRNAs and regulates their expression and alternative splicing, including key oncogenes (PPM1F, FASN) and tumor suppressor genes (RLF, RECQL4), and modulates NF-κB signaling pathway-associated transcripts.","method":"siRNA knockdown combined with RNA-seq and improved RIP-seq (iRIP-seq) in MDA-MB-231 cells; RT-qPCR validation of specific targets","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — genome-wide RIP-seq with functional KD phenotype and target validation by RT-qPCR, but single lab with no structural or biochemical reconstitution","pmids":["41233405"],"is_preprint":false}],"current_model":"SF3B6 (also known as SF3b14a/p14) is an integral component of the human SF3b subcomplex within the 17S U2 snRNP, where it directly contacts the branch site-interacting region of U2 snRNA and stabilizes the branch site:U2 snRNA duplex—thereby facilitating recognition of poorly conserved intron branch sites during spliceosome assembly; structural and biochemical evidence further places SF3B6 at the B complex stage and indicates it may allosterically promote open-to-close conformational transitions in SF3B1 to regulate branch point selection, while cell-based studies show it regulates alternative splicing of diverse pre-mRNA targets and promotes cancer cell proliferation and migration."},"narrative":{"mechanistic_narrative":"SF3B6 (SF3b14a/p14) is an integral component of the human SF3b subcomplex within the 17S U2 snRNP that participates in branch site recognition during spliceosome assembly [PMID:34822310, PMID:16537922]. It directly contacts the branch site-interacting region of U2 snRNA, and these RNA-protein interactions are dynamic, changing as the complex transitions between the 17S U2 snRNP and the spliceosomal A and B complexes [PMID:16537922, PMID:17652325]. High-resolution cryo-EM of the substrate-bound U2 snRNP shows that SF3B6 stabilizes the branch site:U2 snRNA duplex, a function expected to aid recognition of introns with poor complementarity to U2 snRNA [PMID:34822310], and structural work further localizes SF3B6 to a defined position within the spliceosomal B complex [PMID:38383864]. Beyond core splicing, SF3B6 directly binds diverse target mRNAs and regulates their expression and alternative splicing, including oncogene and tumor suppressor transcripts, and its knockdown impairs breast cancer cell proliferation and migration while increasing apoptosis [PMID:41233405].","teleology":[{"year":2006,"claim":"Established that SF3B6 makes physical contact with U2 snRNA near the branch site and that this contact is not static, framing SF3B6 as a dynamic participant in branch site engagement across assembly stages.","evidence":"Lead(II)-induced RNA cleavage and UV cross-linking of immunoaffinity-purified human 17S U2 snRNPs compared across A and B complexes","pmids":["16537922"],"confidence":"High","gaps":["Did not define the precise residues mediating the contact","Functional consequence of the dynamic rearrangement not tested by perturbation"]},{"year":2007,"claim":"Pinpointed the molecular contact, confirming SF3B6 directly cross-links to the branch site-interacting region of U2 snRNA rather than contacting it indirectly.","evidence":"UV cross-linking with IMAC enrichment and MALDI-MS/MS of peptide-RNA heteroconjugates from a partial U2 snRNP","pmids":["17652325"],"confidence":"Medium","gaps":["No mutagenesis to validate the cross-link contribution to function","Single-lab biochemical assignment"]},{"year":2021,"claim":"Resolved at near-atomic detail how SF3B6 acts within the substrate-bound U2 snRNP, showing it stabilizes the branch site:U2 snRNA duplex and thereby could license introns with weak U2 complementarity.","evidence":"Cryo-EM (2.0–2.2 Å) of in vitro-reconstituted human 17S U2 snRNP during ATP-dependent remodeling and pre-mRNA binding","pmids":["34822310"],"confidence":"High","gaps":["Direct demonstration that SF3B6 loss broadens or restricts branch site usage in cells not shown","Does not address regulation of SF3B6 within the complex"]},{"year":2021,"claim":"Proposed a regulatory mechanism in which SF3B6 allosterically promotes an open-to-close conformational transition in SF3B1 to influence branch point selection.","evidence":"Comparative homology modeling, anisotropic network model normal mode analysis, and correlated-motion analysis of human vs. yeast SF3b","pmids":["35028595"],"confidence":"Low","gaps":["Purely computational with no experimental validation of the predicted allostery","Functional inference about residue motions not tested by mutagenesis"]},{"year":2021,"claim":"Linked SF3B6 to a cancer-associated regulatory axis, showing a lncRNA binds SF3B6 and promotes its phosphorylation to alter alternative splicing in pancreatic cancer cells.","evidence":"RNA pull-down, RIP, immunofluorescence, western blotting, and xenograft assays in PDAC cells","pmids":["34470609"],"confidence":"Low","gaps":["Phosphorylation claim lacks kinase identification and phosphosite mapping","No mutagenesis to establish causality of the interaction"]},{"year":2022,"claim":"Tested whether SF3B6 modulates the U2AF2-SF3B1 interface and found it does not, delimiting where SF3B6's influence does and does not lie.","evidence":"Quantitative binding assays of U2AF2 UHM with SF3B1 ULM ± SF3B6","pmids":["35780835"],"confidence":"Medium","gaps":["Negative result confined to a single interface","Does not address SF3B6 effects on other steps of U2 recruitment"]},{"year":2023,"claim":"Showed SF3B6 protein stability and SF3B complex incorporation are independent of PHF5A, indicating SF3B6 levels are buffered against loss of another SF3b subunit.","evidence":"Western blotting and SF3B complex analysis in patient-derived fibroblasts with PHF5A LOF variants","pmids":["37422718"],"confidence":"Low","gaps":["Indirect observation of SF3B6, no direct test of stability determinants","Compensatory mechanism not identified"]},{"year":2024,"claim":"Updated the structural model by localizing SF3B6 to a previously unknown position within the spliceosomal B complex, revealing new contacts at a later assembly stage.","evidence":"Cryo-EM of dimerized human spliceosomal B complexes formed with ATP-γS at improved resolution","pmids":["38383864"],"confidence":"Medium","gaps":["SF3B6-specific contacts not validated by mutagenesis or function","Functional role of the new position inferred from structure alone"]},{"year":2025,"claim":"Extended SF3B6 beyond core splicing, demonstrating it directly binds target mRNAs to regulate their expression and splicing and is required for breast cancer cell proliferation and migration.","evidence":"siRNA knockdown with RNA-seq and iRIP-seq in MDA-MB-231 cells with RT-qPCR target validation","pmids":["41233405"],"confidence":"Medium","gaps":["Direct regulation of named oncogene/suppressor targets not reconstituted biochemically","Mechanism linking SF3B6 to NF-κB-associated transcripts not resolved"]},{"year":null,"claim":"How SF3B6's branch-site duplex stabilization translates into genome-wide branch point and alternative splicing choices, and how its cancer-associated regulation feeds back on this activity, remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No cell-based perturbation linking SF3B6 structural function to defined branch site usage changes","Allosteric regulation of SF3B1 lacks experimental confirmation","Kinase and phosphosites governing SF3B6 regulation unidentified"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0003723","term_label":"RNA binding","supporting_discovery_ids":[0,1,2,8]}],"localization":[],"pathway":[{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[0,1,4,8]}],"complexes":["SF3b subcomplex","17S U2 snRNP","spliceosomal B complex"],"partners":["SF3B1","U2 SNRNA"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9Y3B4","full_name":"Splicing factor 3B subunit 6","aliases":["Pre-mRNA branch site protein p14","SF3b 14 kDa subunit","SF3B14a","Spliceosome-associated protein, 14-kDa","Splicing factor 3b, subunit 6, 14kDa"],"length_aa":125,"mass_kda":14.6,"function":"Component of the 17S U2 SnRNP complex of the spliceosome, a large ribonucleoprotein complex that removes introns from transcribed pre-mRNAs (PubMed:12234937, PubMed:27720643, PubMed:32494006, PubMed:34822310). The 17S U2 SnRNP complex (1) directly participates in early spliceosome assembly and (2) mediates recognition of the intron branch site during pre-mRNA splicing by promoting the selection of the pre-mRNA branch-site adenosine, the nucleophile for the first step of splicing (PubMed:12234937, PubMed:32494006, PubMed:34822310). Within the 17S U2 SnRNP complex, SF3B6 is part of the SF3B subcomplex, which is required for 'A' complex assembly formed by the stable binding of U2 snRNP to the branchpoint sequence in pre-mRNA (PubMed:12234937, PubMed:27720643). Sequence independent binding of SF3A and SF3B subcomplexes upstream of the branch site is essential, it may anchor U2 snRNP to the pre-mRNA (PubMed:12234937). Within the 17S U2 SnRNP complex, SF3B6 directly contacts the pre-mRNA branch site adenosine for the first catalytic step of splicing (PubMed:16432215). SF3B6 stabilizes the intron branch site-U2 snRNA duplex, thereby promoting-binding of introns with poor sequence complementarity (PubMed:34822310). Also acts as a component of the minor spliceosome, which is involved in the splicing of U12-type introns in pre-mRNAs (PubMed:15146077, PubMed:33509932)","subcellular_location":"Nucleus","url":"https://www.uniprot.org/uniprotkb/Q9Y3B4/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/SF3B6","classification":"Common Essential","n_dependent_lines":1203,"n_total_lines":1208,"dependency_fraction":0.9958609271523179},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000115128","cell_line_id":"CID001452","localizations":[{"compartment":"chromatin","grade":3}],"interactors":[{"gene":"SF3A1","stoichiometry":10.0},{"gene":"SF3A2","stoichiometry":10.0},{"gene":"SF3B1","stoichiometry":10.0},{"gene":"SF3B2","stoichiometry":10.0},{"gene":"SF3B3","stoichiometry":10.0},{"gene":"SF3B5","stoichiometry":10.0},{"gene":"TTC33","stoichiometry":10.0},{"gene":"DDX42","stoichiometry":10.0},{"gene":"SNRPD1","stoichiometry":10.0},{"gene":"SNRPD2","stoichiometry":10.0}],"url":"https://opencell.sf.czbiohub.org/target/CID001452","total_profiled":1310},"omim":[{"mim_id":"607835","title":"SPLICING FACTOR 3B, SUBUNIT 6; SF3B6","url":"https://www.omim.org/entry/607835"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Nucleoplasm","reliability":"Supported"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/SF3B6"},"hgnc":{"alias_symbol":["P14","SF3B14a","Ht006","CGI-110","SAP14a"],"prev_symbol":[]},"alphafold":{"accession":"Q9Y3B4","domains":[{"cath_id":"3.30.70.330","chopping":"15-92","consensus_level":"medium","plddt":94.4536,"start":15,"end":92},{"cath_id":"1.10.40","chopping":"94-125","consensus_level":"medium","plddt":88.4987,"start":94,"end":125}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9Y3B4","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9Y3B4-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9Y3B4-F1-predicted_aligned_error_v6.png","plddt_mean":90.12},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=SF3B6","jax_strain_url":"https://www.jax.org/strain/search?query=SF3B6"},"sequence":{"accession":"Q9Y3B4","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9Y3B4.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9Y3B4/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9Y3B4"}},"corpus_meta":[{"pmid":"34822310","id":"PMC_34822310","title":"Structural basis of branch site recognition by the human spliceosome.","date":"2021","source":"Science (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/34822310","citation_count":57,"is_preprint":false},{"pmid":"28296343","id":"PMC_28296343","title":"A genome-wide siRNA screen for regulators of tumor suppressor p53 activity in human non-small cell lung cancer cells identifies components of the RNA splicing machinery as targets for anticancer treatment.","date":"2017","source":"Molecular oncology","url":"https://pubmed.ncbi.nlm.nih.gov/28296343","citation_count":55,"is_preprint":false},{"pmid":"16537922","id":"PMC_16537922","title":"U2 snRNA-protein contacts in purified human 17S U2 snRNPs and in spliceosomal A and B complexes.","date":"2006","source":"Molecular and cellular biology","url":"https://pubmed.ncbi.nlm.nih.gov/16537922","citation_count":41,"is_preprint":false},{"pmid":"34344401","id":"PMC_34344401","title":"Identification of proteins associated with development of psoriatic arthritis in peripheral blood mononuclear cells: a quantitative iTRAQ-based proteomics study.","date":"2021","source":"Journal of translational medicine","url":"https://pubmed.ncbi.nlm.nih.gov/34344401","citation_count":18,"is_preprint":false},{"pmid":"31285391","id":"PMC_31285391","title":"Co-expression of key gene modules and pathways of human breast cancer cell lines.","date":"2019","source":"Bioscience reports","url":"https://pubmed.ncbi.nlm.nih.gov/31285391","citation_count":17,"is_preprint":false},{"pmid":"35780835","id":"PMC_35780835","title":"A UHM-ULM interface with unusual structural features contributes to U2AF2 and SF3B1 association for pre-mRNA splicing.","date":"2022","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/35780835","citation_count":15,"is_preprint":false},{"pmid":"38383864","id":"PMC_38383864","title":"Cryo-EM analyses of dimerized spliceosomes provide new insights into the functions of B complex proteins.","date":"2024","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/38383864","citation_count":14,"is_preprint":false},{"pmid":"17652325","id":"PMC_17652325","title":"Improved identification of enriched peptide RNA cross-links from ribonucleoprotein particles (RNPs) by mass spectrometry.","date":"2007","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/17652325","citation_count":14,"is_preprint":false},{"pmid":"34470609","id":"PMC_34470609","title":"The long non-coding RNA DKFZp434J0226 regulates the alternative splicing process through phosphorylation of SF3B6 in PDAC.","date":"2021","source":"Molecular medicine (Cambridge, Mass.)","url":"https://pubmed.ncbi.nlm.nih.gov/34470609","citation_count":5,"is_preprint":false},{"pmid":"37422718","id":"PMC_37422718","title":"De novo PHF5A variants are associated with craniofacial abnormalities, developmental delay, and hypospadias.","date":"2023","source":"Genetics in medicine : official journal of the American College of Medical Genetics","url":"https://pubmed.ncbi.nlm.nih.gov/37422718","citation_count":5,"is_preprint":false},{"pmid":"35028595","id":"PMC_35028595","title":"Rewards of divergence in sequences, 3-D structures and dynamics of yeast and human spliceosome SF3b complexes.","date":"2021","source":"Current research in structural biology","url":"https://pubmed.ncbi.nlm.nih.gov/35028595","citation_count":3,"is_preprint":false},{"pmid":"41233405","id":"PMC_41233405","title":"SF3B6 promotes the oncogenic phenotypes of MDA-MB-231 cells by extensively interacting with and regulating transcripts expression and alternative splicing.","date":"2025","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/41233405","citation_count":1,"is_preprint":false},{"pmid":"35082828","id":"PMC_35082828","title":"Sequence Divergence and Functional Specializations of the Ancient Spliceosomal SF3b: Implications in Flexibility and Adaptations of the Multi-Protein Complex.","date":"2022","source":"Frontiers in genetics","url":"https://pubmed.ncbi.nlm.nih.gov/35082828","citation_count":1,"is_preprint":false},{"pmid":"39570445","id":"PMC_39570445","title":"Reference gene selection for real-time qPCR in European flounder (Platichthys flesus) using organ-specific RNA-seq data.","date":"2024","source":"Molecular biology reports","url":"https://pubmed.ncbi.nlm.nih.gov/39570445","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":9441,"output_tokens":2305,"usd":0.031449,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":9383,"output_tokens":2924,"usd":0.060007,"stage2_stop_reason":"end_turn"},"total_usd":0.091456,"stage1_batch_id":"msgbatch_013LWZZT6MWaqB9qyTSeQBmU","stage2_batch_id":"msgbatch_0194ZzQaTGNpruoNJBbEMbt3","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2021,\n      \"finding\": \"SF3B6 stabilizes the branch site (BS):U2 snRNA duplex within the substrate-bound U2 snRNP, which could aid binding of introns with poor sequence complementarity to U2 snRNA. High-resolution cryo-EM structures (2.0–2.2 Å) of human 17S U2 snRNP during ATP-dependent remodeling and pre-mRNA binding provided direct snapshots of SF3B6's role in branch site selection.\",\n      \"method\": \"Cryo-EM structure determination (2.0–2.2 Å resolution) of human 17S U2 snRNP reconstituted in vitro with ATP-dependent remodeling and pre-mRNA substrate binding\",\n      \"journal\": \"Science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — high-resolution cryo-EM structures with in vitro reconstitution of U2 snRNP remodeling and substrate binding, multiple structural snapshots providing direct mechanistic insight\",\n      \"pmids\": [\"34822310\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"SF3b14a/p14 (SF3B6) contacts U2 snRNA near the branch site-interacting region (BSiR) in purified human 17S U2 snRNPs, and these RNA-protein interactions are dynamic — changing between 17S U2 snRNP and spliceosomal A and B complexes.\",\n      \"method\": \"Lead(II)-induced RNA cleavage and UV cross-linking of immunoaffinity-purified human 17S U2 snRNPs, compared across spliceosomal A and B complexes\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal UV cross-linking and chemical probing on purified native complexes; findings replicated across multiple spliceosomal assembly states\",\n      \"pmids\": [\"16537922\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"p14/SF3b14a (SF3B6) directly contacts the branch site-interacting region (BSiR) of U2 snRNA, as identified by MS analysis of UV cross-linked peptide-RNA conjugates from a partial U2 snRNP complex.\",\n      \"method\": \"UV cross-linking combined with IMAC enrichment and MALDI-MS/MS analysis of peptide-RNA heteroconjugates from purified partial U2 snRNP complex\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — direct biochemical identification of contact site by MS, but single lab, no mutagenesis validation reported in abstract\",\n      \"pmids\": [\"17652325\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"SF3B6 interacts with the N-terminal extension of SF3B1 and, through computational structural and normal mode analyses, promotes a functionally relevant 'open-to-close' conformational transition in SF3B1 by enhancing concerted residue motions, thereby acting as an allosteric regulator of SF3B1 for branch point sequence (BPS) selection. SF3B6 influences motions of 16 SF3B1 residues that interact with U2 snRNA/branchpoint duplex.\",\n      \"method\": \"Comparative homology modeling, anisotropic network model (ANM) normal mode analysis, and inter-residue correlated motion analysis of human vs. yeast SF3b complexes\",\n      \"journal\": \"Current research in structural biology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 4 / Weak — purely computational analysis (normal mode modeling), no experimental validation reported; functional inference is speculative\",\n      \"pmids\": [\"35028595\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"In the human spliceosomal B complex, SF3B6 was localized to a previously unknown position, revealing new molecular contacts and functions during pre-mRNA splicing at the B complex stage.\",\n      \"method\": \"Cryo-EM structure determination of dimerized human spliceosomal B complexes formed in the presence of ATP-γS, at improved resolution\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — high-resolution cryo-EM structural localization, single study, no mutagenesis or functional validation of SF3B6-specific contacts reported in abstract\",\n      \"pmids\": [\"38383864\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"An additional SF3B6 subunit had no detectable effect on U2AF2-SF3B1 binding affinities, as measured by quantitative binding assays.\",\n      \"method\": \"Quantitative binding affinity measurements (fluorescence anisotropy or ITC) of U2AF2 UHM with SF3B1 ULM in the presence or absence of SF3B6\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — direct quantitative binding assay with crystal structure context, but this is a negative result (SF3B6 had no effect), single lab\",\n      \"pmids\": [\"35780835\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"The lncRNA DKFZp434J0226 physically interacts with SF3B6 and promotes its phosphorylation, which further regulates alternative splicing of pre-mRNA in pancreatic ductal adenocarcinoma (PDAC) cells.\",\n      \"method\": \"RNA pull-down, RNA immunoprecipitation (RIP), immunofluorescence, and western blotting in PDAC cells; tumor xenograft in vivo assays\",\n      \"journal\": \"Molecular medicine\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, RNA pull-down and RIP without mutagenesis; phosphorylation claim is not directly validated with kinase identification or phosphosite mapping in abstract\",\n      \"pmids\": [\"34470609\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"In fibroblasts with PHF5A loss-of-function (LOF) variants, SF3B6 protein levels and SF3B complex formation were unaffected, suggesting SF3B6 protein stability does not require PHF5A and that compensatory mechanisms maintain normal SF3B component levels.\",\n      \"method\": \"Western blotting and SF3B complex analysis in subject-derived fibroblasts with PHF5A LOF variants\",\n      \"journal\": \"Genetics in medicine\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — indirect finding about SF3B6, single lab, limited mechanistic follow-up for SF3B6 specifically\",\n      \"pmids\": [\"37422718\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"SF3B6 knockdown in MDA-MB-231 breast cancer cells inhibits cellular proliferation and migration and increases apoptosis. iRIP-seq demonstrated that SF3B6 directly binds target mRNAs and regulates their expression and alternative splicing, including key oncogenes (PPM1F, FASN) and tumor suppressor genes (RLF, RECQL4), and modulates NF-κB signaling pathway-associated transcripts.\",\n      \"method\": \"siRNA knockdown combined with RNA-seq and improved RIP-seq (iRIP-seq) in MDA-MB-231 cells; RT-qPCR validation of specific targets\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — genome-wide RIP-seq with functional KD phenotype and target validation by RT-qPCR, but single lab with no structural or biochemical reconstitution\",\n      \"pmids\": [\"41233405\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"SF3B6 (also known as SF3b14a/p14) is an integral component of the human SF3b subcomplex within the 17S U2 snRNP, where it directly contacts the branch site-interacting region of U2 snRNA and stabilizes the branch site:U2 snRNA duplex—thereby facilitating recognition of poorly conserved intron branch sites during spliceosome assembly; structural and biochemical evidence further places SF3B6 at the B complex stage and indicates it may allosterically promote open-to-close conformational transitions in SF3B1 to regulate branch point selection, while cell-based studies show it regulates alternative splicing of diverse pre-mRNA targets and promotes cancer cell proliferation and migration.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"SF3B6 (SF3b14a/p14) is an integral component of the human SF3b subcomplex within the 17S U2 snRNP that participates in branch site recognition during spliceosome assembly [#0, #1]. It directly contacts the branch site-interacting region of U2 snRNA, and these RNA-protein interactions are dynamic, changing as the complex transitions between the 17S U2 snRNP and the spliceosomal A and B complexes [#1, #2]. High-resolution cryo-EM of the substrate-bound U2 snRNP shows that SF3B6 stabilizes the branch site:U2 snRNA duplex, a function expected to aid recognition of introns with poor complementarity to U2 snRNA [#0], and structural work further localizes SF3B6 to a defined position within the spliceosomal B complex [#4]. Beyond core splicing, SF3B6 directly binds diverse target mRNAs and regulates their expression and alternative splicing, including oncogene and tumor suppressor transcripts, and its knockdown impairs breast cancer cell proliferation and migration while increasing apoptosis [#8].\",\n  \"teleology\": [\n    {\n      \"year\": 2006,\n      \"claim\": \"Established that SF3B6 makes physical contact with U2 snRNA near the branch site and that this contact is not static, framing SF3B6 as a dynamic participant in branch site engagement across assembly stages.\",\n      \"evidence\": \"Lead(II)-induced RNA cleavage and UV cross-linking of immunoaffinity-purified human 17S U2 snRNPs compared across A and B complexes\",\n      \"pmids\": [\"16537922\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Did not define the precise residues mediating the contact\",\n        \"Functional consequence of the dynamic rearrangement not tested by perturbation\"\n      ]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Pinpointed the molecular contact, confirming SF3B6 directly cross-links to the branch site-interacting region of U2 snRNA rather than contacting it indirectly.\",\n      \"evidence\": \"UV cross-linking with IMAC enrichment and MALDI-MS/MS of peptide-RNA heteroconjugates from a partial U2 snRNP\",\n      \"pmids\": [\"17652325\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"No mutagenesis to validate the cross-link contribution to function\",\n        \"Single-lab biochemical assignment\"\n      ]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Resolved at near-atomic detail how SF3B6 acts within the substrate-bound U2 snRNP, showing it stabilizes the branch site:U2 snRNA duplex and thereby could license introns with weak U2 complementarity.\",\n      \"evidence\": \"Cryo-EM (2.0\\u20132.2 \\u00c5) of in vitro-reconstituted human 17S U2 snRNP during ATP-dependent remodeling and pre-mRNA binding\",\n      \"pmids\": [\"34822310\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Direct demonstration that SF3B6 loss broadens or restricts branch site usage in cells not shown\",\n        \"Does not address regulation of SF3B6 within the complex\"\n      ]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Proposed a regulatory mechanism in which SF3B6 allosterically promotes an open-to-close conformational transition in SF3B1 to influence branch point selection.\",\n      \"evidence\": \"Comparative homology modeling, anisotropic network model normal mode analysis, and correlated-motion analysis of human vs. yeast SF3b\",\n      \"pmids\": [\"35028595\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\n        \"Purely computational with no experimental validation of the predicted allostery\",\n        \"Functional inference about residue motions not tested by mutagenesis\"\n      ]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Linked SF3B6 to a cancer-associated regulatory axis, showing a lncRNA binds SF3B6 and promotes its phosphorylation to alter alternative splicing in pancreatic cancer cells.\",\n      \"evidence\": \"RNA pull-down, RIP, immunofluorescence, western blotting, and xenograft assays in PDAC cells\",\n      \"pmids\": [\"34470609\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\n        \"Phosphorylation claim lacks kinase identification and phosphosite mapping\",\n        \"No mutagenesis to establish causality of the interaction\"\n      ]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Tested whether SF3B6 modulates the U2AF2-SF3B1 interface and found it does not, delimiting where SF3B6's influence does and does not lie.\",\n      \"evidence\": \"Quantitative binding assays of U2AF2 UHM with SF3B1 ULM \\u00b1 SF3B6\",\n      \"pmids\": [\"35780835\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Negative result confined to a single interface\",\n        \"Does not address SF3B6 effects on other steps of U2 recruitment\"\n      ]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Showed SF3B6 protein stability and SF3B complex incorporation are independent of PHF5A, indicating SF3B6 levels are buffered against loss of another SF3b subunit.\",\n      \"evidence\": \"Western blotting and SF3B complex analysis in patient-derived fibroblasts with PHF5A LOF variants\",\n      \"pmids\": [\"37422718\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\n        \"Indirect observation of SF3B6, no direct test of stability determinants\",\n        \"Compensatory mechanism not identified\"\n      ]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Updated the structural model by localizing SF3B6 to a previously unknown position within the spliceosomal B complex, revealing new contacts at a later assembly stage.\",\n      \"evidence\": \"Cryo-EM of dimerized human spliceosomal B complexes formed with ATP-\\u03b3S at improved resolution\",\n      \"pmids\": [\"38383864\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"SF3B6-specific contacts not validated by mutagenesis or function\",\n        \"Functional role of the new position inferred from structure alone\"\n      ]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Extended SF3B6 beyond core splicing, demonstrating it directly binds target mRNAs to regulate their expression and splicing and is required for breast cancer cell proliferation and migration.\",\n      \"evidence\": \"siRNA knockdown with RNA-seq and iRIP-seq in MDA-MB-231 cells with RT-qPCR target validation\",\n      \"pmids\": [\"41233405\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Direct regulation of named oncogene/suppressor targets not reconstituted biochemically\",\n        \"Mechanism linking SF3B6 to NF-\\u03baB-associated transcripts not resolved\"\n      ]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How SF3B6's branch-site duplex stabilization translates into genome-wide branch point and alternative splicing choices, and how its cancer-associated regulation feeds back on this activity, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"No cell-based perturbation linking SF3B6 structural function to defined branch site usage changes\",\n        \"Allosteric regulation of SF3B1 lacks experimental confirmation\",\n        \"Kinase and phosphosites governing SF3B6 regulation unidentified\"\n      ]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0003723\", \"supporting_discovery_ids\": [0, 1, 2, 8]}\n    ],\n    \"localization\": [],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [0, 1, 4, 8]}\n    ],\n    \"complexes\": [\n      \"SF3b subcomplex\",\n      \"17S U2 snRNP\",\n      \"spliceosomal B complex\"\n    ],\n    \"partners\": [\n      \"SF3B1\",\n      \"U2 snRNA\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":4,"faith_total":4,"faith_pct":100.0}}