{"gene":"PRPF8","run_date":"2026-06-10T06:43:36","timeline":{"discoveries":[{"year":1989,"finding":"The mammalian homolog of yeast PRP8 (hPrp8/p220) is a component of the U4/U5/U6 snRNP complex and of purified spliceosomes, as demonstrated by gradient fractionation and immunoprecipitation with anti-PRP8 antibodies in HeLa nuclear extracts.","method":"Gradient fractionation, immunoprecipitation, Western blot","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal immunoprecipitation and gradient fractionation, single lab, two orthogonal methods","pmids":["2479028"],"is_preprint":false},{"year":1990,"finding":"PRP8 protein maintains stable association with the spliceosome throughout both steps of the splicing reaction and is present in post-splicing complexes containing the excised intron.","method":"Affinity purification of spliceosomes, immunoprecipitation with anti-PRP8 antibodies","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — affinity purification and immunological detection, single lab","pmids":["2138328"],"is_preprint":false},{"year":1990,"finding":"A mammalian ~220 kDa protein (p220) is UV-crosslinked to pre-mRNAs under splicing conditions; anti-yeast PRP8 antibodies recognize and immunoprecipitate this UV-crosslinked protein, establishing structural conservation and direct pre-mRNA contact.","method":"UV-crosslinking, immunoprecipitation, Western blot with anti-PRP8 antisera","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — UV-crosslinking plus immunoprecipitation, single lab, two orthogonal methods","pmids":["2139226"],"is_preprint":false},{"year":1991,"finding":"Yeast PRP8 protein directly contacts pre-mRNA in an ATP-dependent, splicing-specific manner; PRP8 only crosslinks to RNA substrates competent for step 1 of splicing, not to mutant substrates with 5' splice site or branchpoint mutations.","method":"UV-crosslinking combined with immunoprecipitation, in vitro splicing assay","journal":"Nucleic acids research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — UV-crosslinking with mutant substrates as controls, single lab","pmids":["1945827"],"is_preprint":false},{"year":1991,"finding":"A suppressor of prp8-1 (spp81) encodes a putative ATP-dependent RNA helicase (later identified as Brr2), establishing a genetic interaction between PRP8 and an RNA helicase in the splicing pathway.","method":"Extragenic suppressor screen, DNA sequencing, sequence homology analysis","journal":"Nature","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic suppressor screen with sequence characterization, single lab","pmids":["1996139"],"is_preprint":false},{"year":1992,"finding":"Functional PRP8 is required for stable formation of U4/U6.U5 triple snRNP complexes and for assembly of triple snRNPs into spliceosomes; depletion of PRP8 also causes dramatic decline in U4, U5, and U6 snRNA levels.","method":"Genetic depletion in vivo, temperature-sensitive inactivation in protoplasts, antibody inhibition in vitro, sedimentation analysis","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — three orthogonal loss-of-function approaches (genetic depletion, ts inactivation, antibody inhibition) in single study","pmids":["1396567"],"is_preprint":false},{"year":1995,"finding":"PRP8 protein directly contacts at least eight exon nucleotides at the 5' splice site prior to step 1 of splicing, and at least 13 exon nucleotides plus part of the polypyrimidine tract at the 3' splice site after step 1; these interactions are not sequence-specific. The data support a model where PRP8 stabilizes U5 snRNA interactions with both exons for alignment at the spliceosome active site.","method":"4-thiouridine UV-crosslinking with site-specific labeling, analysis of mutant and duplicated splice sites","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1 / Strong — systematic site-specific crosslinking with mutant controls, multiple splice-site positions tested","pmids":["7781612"],"is_preprint":false},{"year":1995,"finding":"PRP8 protein can be UV-crosslinked to pre-mRNA in PRP2-depleted spliceosomes stalled before step 1, interacts with substrate RNA fragments at the 5' splice site region and the branchpoint–3' splice site region, with the latter interaction established only after step 1 of splicing.","method":"UV-crosslinking combined with PRP8-specific immunoprecipitation and RNase T1 treatment","journal":"Nucleic acids research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — immunoprecipitation of crosslinked RNA fragments with enzymatic mapping, single lab","pmids":["7885825"],"is_preprint":false},{"year":1996,"finding":"Mutagenesis of yeast PRP8 identifies two separable functional domains: one governing specificity of 3' splice site selection (uridine tract recognition) and one governing fidelity of 3' splice site utilization (PyAG motif recognition), implicating Prp8p in functional roles at the spliceosome active site during the second catalytic step.","method":"Random mutagenesis, genetic selection, allele characterization, in vivo splicing assays","journal":"Genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — systematic mutagenesis with functional in vivo assays, single lab","pmids":["8725222"],"is_preprint":false},{"year":1998,"finding":"Human U5-220 kDa protein (hPrp8) forms a stable, RNA-free complex with the U5-116 kDa EF-2 homologue (Snu114), the 200 kDa RNA unwindase (Brr2), and a novel WD-40 repeat protein (U5-40 kDa); the 220 kDa protein binds simultaneously to the 40 kDa and 116 kDa proteins and probably also to the 200 kDa protein.","method":"Chaotropic salt dissociation of U5 snRNP, sedimentation analysis, cDNA cloning, biochemical binding analysis","journal":"Molecular and cellular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — biochemical reconstitution with sedimentation and binding analyses, single lab","pmids":["9774689"],"is_preprint":false},{"year":1999,"finding":"The C-terminal region of hPrp8 (residues 1894–1898) forms a UV-induced crosslink with the conserved GU dinucleotide at the 5' splice site within spliceosomal complex B, mapping the functional interaction domain.","method":"UV-crosslinking, immunoprecipitation, proteolytic mapping, size comparison of crosslinked peptides","journal":"RNA (New York, N.Y.)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — UV-crosslinking with systematic proteolytic mapping to identify contact residues, single lab","pmids":["10024169"],"is_preprint":false},{"year":1999,"finding":"Yeast Prp8 mutants can suppress mutations at position 2 of the 5' GU, all positions of the 3' YAG, and position A51 in the U6 ACAGAG motif, implying that Prp8 participates in a tertiary interaction between U6 snRNA and both splice site ends and plays a functional role at the active site of the spliceosome.","method":"Genetic suppressor screen, allele-specific suppression analysis, in vivo splicing assays","journal":"Genes & development","confidence":"High","confidence_rationale":"Tier 2 / Strong — allele-specific genetic epistasis with multiple splice-site targets, replicated across two concurrent independent studies (PMID 10444595 and 10444596)","pmids":["10444595","10444596"],"is_preprint":false},{"year":1999,"finding":"Mutagenesis of the yeast Prp8 region corresponding to the 5'SS:hPrp8 crosslink identifies alleles that suppress both 5' and 3' splice site mutations, placing Prp8 functional interactions with both splice sites at the later stage of splicing affecting the second catalytic step.","method":"Site-directed and random mutagenesis, in vivo suppression assays, combined analysis with U1 suppressor snRNA","journal":"Genes & development","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal genetic analyses in single study, corroborated by independent concurrent study","pmids":["10444596"],"is_preprint":false},{"year":1999,"finding":"A novel Prp8 mutation (prp8-201) suppresses the growth defect of cold-sensitive U4-cs1, which blocks U4/U6 unwinding; wild-type Prp8 triggers U4/U6 RNA unwinding only after correct 5' splice site recognition by the U6 ACAGA box, indicating Prp8 governs the timing of spliceosome activation.","method":"Genetic suppressor analysis, in vitro splicing assay with cold-sensitive U4-cs1 block, spliceosomal complex analysis","journal":"Molecular cell","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic suppressor combined with in vitro splicing and complex analysis, single lab","pmids":["10024880"],"is_preprint":false},{"year":1999,"finding":"Human Prp8 (hPrp8p) is a core component of both the major U2-dependent and the minor U12-dependent spliceosomes, the first non-Sm factor shown to be common to both spliceosomes.","method":"Immunoprecipitation with anti-hPrp8 antibodies, Northern blot analysis of snRNAs","journal":"RNA (New York, N.Y.)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — immunoprecipitation with snRNA analysis, single lab","pmids":["10411133"],"is_preprint":false},{"year":2000,"finding":"Large-scale suppressor screen identifies five distinct regions (a–e) of Prp8 that control spliceosome activation; two regions contact U1 snRNP (two-hybrid interaction), another mediates indirect contact; allosteric changes in Prp8 are proposed to initiate activation by disrupting U1 snRNP contacts with tri-snRNP and coordinating Brr2 and Prp24 activities.","method":"Large-scale genetic suppressor screen, yeast two-hybrid, analysis of genetic interactions","journal":"Genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — large-scale suppressor screen plus yeast two-hybrid, single lab","pmids":["10924465"],"is_preprint":false},{"year":2001,"finding":"Seven different missense mutations in PRPC8 (PRPF8) clustered within a 14-codon stretch at the C-terminus cause autosomal dominant retinitis pigmentosa (RP13), establishing that mutations in this ubiquitous splicing factor cause retinal-specific degeneration.","method":"Positional cloning, direct sequencing, cosegregation analysis in multiple families","journal":"Human molecular genetics","confidence":"High","confidence_rationale":"Tier 2 / Strong — cosegregation demonstrated across multiple independent families, multiple independent mutations identified","pmids":["11468273"],"is_preprint":false},{"year":2001,"finding":"Deletion of SKY1 (SRPK-family kinase) is synthetically lethal with specific prp8 alleles in a domain implicated in 3'AG recognition fidelity, and sky1 deletion suppresses 3'AG mutations, suggesting that 3' splice site AG recognition by Prp8 is subject to phosphorylation regulation.","method":"Genetic synthetic lethality analysis, ACT1-CUP1 splicing reporter assay, yeast genetics","journal":"RNA (New York, N.Y.)","confidence":"Low","confidence_rationale":"Tier 3 / Weak — genetic interaction data, single lab, no direct biochemical evidence for phosphorylation of Prp8","pmids":["11565750"],"is_preprint":false},{"year":2002,"finding":"Distinct domains of Prp8 mediate different aspects of spliceosome activation: regions a, d, and e show allele-specific genetic interactions with Prp28, Brr2 (Prp44), and U6 RNA respectively, revealing that Prp8 coordinates multiple processes including U1 snRNP release and U4/U6 unwinding.","method":"Allele-specific genetic interaction analysis, yeast genetics","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — systematic allele-specific genetic epistasis across multiple factors, single lab","pmids":["12087126"],"is_preprint":false},{"year":2003,"finding":"A missense mutation in PRP8 (R1753K) suppresses multiple helicase-deficient prp22 mutations, suggesting Prp8 stabilizes an RNA-protein or RNA-RNA interaction in the spliceosome that must be disrupted by Prp22's helicase activity for mRNA release.","method":"Extragenic suppressor screen, in vitro splicing assay, mRNA release assay","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — suppressor genetics combined with in vitro splicing assay, single lab","pmids":["14688266"],"is_preprint":false},{"year":2006,"finding":"Assembly of Snu114 into the U5 snRNP requires a functional GTPase domain and Prp8; GTPase domain mutants of Snu114 fail to interact with Prp8 or U5 snRNA and cannot assemble U5 snRNPs, whereas C-terminal truncation mutants assemble spliceosomes but block U4 snRNP release.","method":"snRNP and spliceosome assembly analysis in SNU114 mutant extracts, immunoprecipitation","journal":"RNA (New York, N.Y.)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct assembly analysis with defined mutants, single lab","pmids":["16540695"],"is_preprint":false},{"year":2006,"finding":"Transposon-based dissection of Prp8 establishes that catalytic core RNAs (U5, U6 snRNAs, pre-mRNA) make multiple contacts in the central region of Prp8, identifying a discrete highly conserved region as a candidate splicing cofactor.","method":"Transposon-based insertion mutagenesis, RNA-protein crosslinking, functional analysis","journal":"RNA (New York, N.Y.)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — systematic transposon insertion with crosslinking analysis, single lab","pmids":["16431982"],"is_preprint":false},{"year":2007,"finding":"The C-terminal domain of yeast Prp8p adopts a Jab1/MPN-like fold (pseudoenzyme with impaired metal-binding site) that serves as a protein-protein interaction platform; RP13-linked mutations in the C-terminal appendix weaken interactions with Brr2 and Snu114.","method":"X-ray crystallography, targeted yeast two-hybrid analysis, mutagenesis","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure combined with functional mutagenesis and yeast two-hybrid, single study with multiple orthogonal methods","pmids":["17317632"],"is_preprint":false},{"year":2007,"finding":"RP13 mutations in Prp8 in yeast cause nuclear accumulation of a precursor U5 snRNP that lacks Brr2, and Prp8 contains a nuclear localization signal required for efficient nuclear import of the precursor U5 snRNP; Brr2 joins U5 snRNP in the nucleus, and RP mutations disrupt the Prp8–Brr2 interaction required for this step.","method":"Yeast genetics, immunofluorescence, subcellular fractionation, co-immunoprecipitation","journal":"Nature structural & molecular biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (genetics, localization, fractionation, co-IP) establishing a biogenesis pathway, single study","pmids":["17934474"],"is_preprint":false},{"year":2007,"finding":"Opposing classes of prp8 alleles modulate the equilibrium between the first and second catalytic steps of splicing: one class suppresses first-step defects (analogous to ribosomal 'ram' mutants) and opposes previously described second-step suppressors; this transition is linked to U6 snRNA and Prp16 ATPase activity.","method":"Genetic characterization of prp8 alleles, genetic interaction epistasis with U6 and prp16 mutations, in vivo splicing assays","journal":"Nature structural & molecular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — systematic genetic analysis with epistasis, single lab","pmids":["17486100"],"is_preprint":false},{"year":2008,"finding":"The crystal structure of the beta-finger domain of Prp8 (residues 1822–2095) reveals a beta-hairpin finger protruding from the protein; mutations throughout the beta-finger alter the equilibrium between first and second catalytic steps, while mutations at its base affect U4/U6 unwinding-mediated spliceosome activation.","method":"X-ray crystallography, mutagenesis, in vivo functional analysis","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1 / Moderate — crystal structure combined with systematic mutagenesis and functional assays, single lab with multiple orthogonal methods","pmids":["18779563"],"is_preprint":false},{"year":2008,"finding":"A fragment from the C-terminus of Prp8 activates Brr2-dependent U4/U6 snRNA dissociation in vitro; notably, fragments carrying RP-associated prp8 alleles do not stimulate U4/U6 unwinding activity. The same fragment also inhibits Brr2 U4/U6-dependent ATPase activity.","method":"In vitro U4/U6 unwinding assay, ATPase assay, recombinant protein fragments","journal":"Nature structural & molecular biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro reconstitution assay with defined protein fragments and RP mutant variants, multiple activities measured","pmids":["19098916"],"is_preprint":false},{"year":2008,"finding":"Electron microscopy of the yeast tri-snRNP localizes Brr2 to a head domain, while Prp8 and Snu114 occupy a central position; the head (containing Brr2) and arm (containing U4/U6 snRNP) adopt variable relative positions, suggesting conformational dynamics relevant to spliceosome activation.","method":"EM projection structure of genetically tagged tri-snRNP proteins","journal":"Nature structural & molecular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — EM with genetic tagging for localization, single lab","pmids":["18953335"],"is_preprint":false},{"year":2008,"finding":"The 1.85 Å crystal structure of PRP8 domain IV reveals a bipartite structure with an RNase H fold linked to a five-helix assembly; RNA binding studies and analysis of mutant yeast alleles in context of this structure show domain IV forms a surface interacting directly with RNA structures at the spliceosome catalytic core.","method":"X-ray crystallography (1.85 Å), RNA binding studies, mutant allele analysis","journal":"Nature structural & molecular biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — high-resolution crystal structure combined with RNA binding studies and mutant analysis, multiple orthogonal methods","pmids":["18836455"],"is_preprint":false},{"year":2011,"finding":"Bioinformatic and structural analysis reveals that Prp8's central conserved domain is related to the catalytic domain of reverse transcriptases (most similar to prokaryotic retroelement RTs), followed by a region analogous to maturase/X domains and a C-terminal RNaseH-like fold, suggesting Prp8 evolved by acquiring nucleic acid-binding domains from inactivated retroelements.","method":"Sequence analysis, structural homology analysis","journal":"RNA (New York, N.Y.)","confidence":"Low","confidence_rationale":"Tier 4 / Moderate — computational/structural prediction without direct biochemical validation of proposed evolutionary origin","pmids":["21441348"],"is_preprint":false},{"year":2012,"finding":"The RNase H (RH) domain of Prp8 binds U4/U6 snRNA through the single-stranded regions preceding U4/U6 stem I; Brr2 also recognizes this same single-stranded U4 region and translocates along U4 to unwind stem I first; the Prp8 RH domain inhibits U4/U6 unwinding by blocking Brr2 loading onto U4 snRNA.","method":"RNA binding assays, cross-linking coupled with mass spectrometry, in vitro U4/U6 unwinding inhibition assay","journal":"Genes & development","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro binding and unwinding assays combined with crosslinking-mass spectrometry mapping of contact residues, multiple orthogonal methods","pmids":["23124066"],"is_preprint":false},{"year":2013,"finding":"The RNase H domain of PRP8 undergoes a conformational switch between the two steps of splicing, and this switch unmasks a metal-binding site involved in the second catalytic step (exon ligation); PRP8 is a metalloprotein that promotes exon ligation within the spliceosome.","method":"Biochemical assays, metal binding studies, analysis of prp8 alleles promoting first vs. second step","journal":"Nature structural & molecular biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — direct metal binding demonstration combined with conformational analysis and allele-based functional validation","pmids":["23686287"],"is_preprint":false},{"year":2013,"finding":"Crystal structure of yeast Prp8 (residues 885–2413) in complex with Aar2 reveals tightly associated domains resembling a bacterial group II intron reverse transcriptase and a type II restriction endonuclease; a large cavity formed by the reverse transcriptase thumb, endonuclease-like, and RNaseH-like domains accommodates splice-site suppressors and the intron branch-point crosslink, establishing this cavity as the active site of the spliceosome.","method":"X-ray crystallography of Prp8–Aar2 complex, mapping of suppressor mutations and crosslinks onto structure","journal":"Nature","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure at high resolution combined with mapping of functionally validated genetic suppressors, landmark structural study","pmids":["23354046"],"is_preprint":false},{"year":2013,"finding":"Structural and biochemical analyses show that the Prp8 Jab1/MPN domain binds exclusively to the N-terminal helicase cassette of Brr2 and stimulates Brr2 activity; RP-associated mutations in the Jab1/MPN domain map to the Brr2 interface; Aar2 and Brr2 are mutually exclusive binders of the Jab1/MPN domain, explaining the cytoplasm-to-nucleus switch during U5 snRNP maturation.","method":"X-ray crystallography of Brr2–Prp8 Jab1/MPN domain complex, mutagenesis, biochemical activity assays","journal":"Structure (London, England : 1993)","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure combined with mutagenesis and functional biochemical assays, multiple orthogonal methods","pmids":["23727230"],"is_preprint":false},{"year":2013,"finding":"Prp8 inserts its C-terminal tail into Brr2's RNA-binding tunnel, thereby competitively blocking Brr2's RNA-binding, ATPase, and U4/U6 unwinding activities; RP-linked Prp8 mutations that map to this tail cause inefficient Brr2 repression as their primary recognizable phenotype.","method":"X-ray crystallography, biochemical helicase assays (ATPase, unwinding), mutagenesis","journal":"Science (New York, N.Y.)","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure plus multiple in vitro biochemical activity assays with RP mutant variants, multiple orthogonal methods in single rigorous study","pmids":["23704370"],"is_preprint":false},{"year":2013,"finding":"In vivo CLIP/CRAC analysis reveals that Prp8 contacts U1 and U2 snRNAs in addition to its known U5, U6, and pre-mRNA contacts; disruption of Prp8–U1 snRNA interaction reduces tri-snRNP levels in the spliceosome, revealing a role for Prp8 in spliceosomal assembly through U1 snRNA interaction.","method":"CLIP/CRAC with next-generation sequencing, in vivo crosslinking, functional analysis of U1 interaction mutants","journal":"Nucleic acids research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — comprehensive in vivo crosslinking combined with functional tri-snRNP assembly analysis, single lab","pmids":["23393194"],"is_preprint":false},{"year":2013,"finding":"Prp8 contacts with nucleotides surrounding the branch-site are enhanced during step 1 catalysis (in C complex), as revealed by UV-induced crosslinking of purified yeast B(act) and C spliceosomes on site-specifically labeled pre-mRNA.","method":"UV-induced crosslinking of purified spliceosomal complexes with site-specifically labeled pre-mRNA","journal":"PLoS genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — purified spliceosome crosslinking with defined labeling, single lab","pmids":["26393790"],"is_preprint":false},{"year":2014,"finding":"Picornaviral 3D polymerase (RdRp) enters the nucleus and associates with the C-terminal Jab1/MPN domain of Prp8, interfering with the second catalytic step of splicing and causing accumulation of lariat splicing intermediates; this disrupts pre-mRNA splicing of endogenous transcripts involved in cell growth and differentiation.","method":"Co-immunoprecipitation, domain mapping, in vitro splicing assay, nuclear fractionation, identification of endogenous trapped pre-mRNAs","journal":"PLoS pathogens","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-immunoprecipitation with domain mapping and in vitro splicing assay, single lab","pmids":["24968230"],"is_preprint":false},{"year":2014,"finding":"PRPF8 deficiency (knockdown or hemizygous deletion) in K562 and CD34+ bone marrow cells causes missplicing defects and increased proliferative capacity; yeast models with homologous PRPF8 mutations abrogate the second-step block in splicing, suggesting the mutations impair proofreading function.","method":"Knockdown experiments, whole-RNA deep sequencing, yeast complementation/mutagenesis","journal":"Leukemia","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function with transcriptomic readout combined with yeast functional validation, single lab","pmids":["24781015"],"is_preprint":false},{"year":2015,"finding":"PRPF8 depletion preferentially impairs splicing of introns with weak 5' splice sites across the human transcriptome, leading to mitotic arrest; iCLIP shows PRPF8 depletion decreases RNP complex formation at most splice sites, and experimental enhancement of 5' splice site strength overcomes the effects of PRPF8 depletion.","method":"siRNA knockdown, RNA-seq, iCLIP, minigene splicing assays","journal":"Genome biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — iCLIP plus RNA-seq plus minigene functional rescue, multiple orthogonal methods establishing mechanism","pmids":["26392272"],"is_preprint":false},{"year":2015,"finding":"Stable tri-snRNP integration into the exon-defined complex requires interaction between Prp8 and nucleotides at the exon–intron junction (5' splice site), and this is the key trigger for the major structural rearrangement from 37S to 45S B-like complex, independent of B-specific proteins or hPrp31 phosphorylation.","method":"Electron microscopy, affinity purification, addition of 5'ss RNA oligonucleotides to cross-exon complexes","journal":"RNA (New York, N.Y.)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — EM structural analysis combined with functional reconstitution using defined RNA oligonucleotides, single lab","pmids":["26385511"],"is_preprint":false},{"year":2015,"finding":"Prp8 interacts physically with the androgen receptor (AR) via the AR nuclear export signal (NES); co-immunoprecipitation and deletion mutagenesis demonstrate Prp8-AR interaction, and Prp8 knockdown induces nuclear accumulation of AR and increases its polyubiquitination, modulating AR transcriptional activity.","method":"Co-immunoprecipitation, deletion mutagenesis, shRNA knockdown, rapamycin export assay, luciferase reporter assay","journal":"Molecular endocrinology (Baltimore, Md.)","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single co-IP with knockdown, single lab, unclear mechanistic specificity for splicing vs. direct AR regulation","pmids":["26371515"],"is_preprint":false},{"year":2016,"finding":"A subset of RP-linked Prp8 mutations (mapping to the hinge linking the Jab1-MPN tail to the globular domain) cause defects in the transition between the first and second catalytic steps of splicing, in addition to spliceosome activation defects; genetic analyses link Snu114 GTP/GDP occupancy to Prp8-dependent regulation of Brr2.","method":"Yeast genetics, in vivo and in vitro splicing reporter assays, genetic interaction analysis with SNU114 mutants","journal":"RNA (New York, N.Y.)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — combination of in vitro and in vivo splicing assays with genetic epistasis, single lab","pmids":["26968627"],"is_preprint":false},{"year":2017,"finding":"The RNaseH domain of Prp8 contains a 17-aa extension (Switch loop) that can adopt two mutually exclusive structures; prp8 alleles mapping to this extension fall into two functional classes—those promoting error-prone/efficient splicing versus hyperaccurate/inefficient splicing—and error-prone alleles suppress a prp2 mutant deficient at promoting the first catalytic step.","method":"Systematic mutagenesis, in vitro and in vivo reporter assays, lariat sequencing for genome-wide splice site analysis, genetic epistasis","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1 / Strong — multiple orthogonal approaches including structural rationale, in vitro assays, in vivo reporters, and genome-wide lariat sequencing","pmids":["28416677"],"is_preprint":false},{"year":2017,"finding":"PRPF8 depletion causes a specific defect in homology-directed repair (HDR) and single-strand annealing (SSA), which require BRCA1; PRPF8 depletion reduces end resection (measured as chromatin-bound RPA), BRCA1 foci, and histone acetylation marks associated with BRCA1-mediated HR.","method":"siRNA knockdown, DR-GFP/SA-GFP HR reporter assays, immunofluorescence (BRCA1, RPA, histone acetylation foci)","journal":"Oncotarget","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function with multiple functional readouts for HR, single lab","pmids":["29212152"],"is_preprint":false},{"year":2018,"finding":"Mutagenesis of Prp8 residues identified by amino acid probing to be positioned near U5 snRNA reveals their role in 5' splice site recognition; genetic interactions with Isy1 and Snu114 further support that Prp8–U5 snRNA interactions help position the pre-mRNA into the active site.","method":"Amino acid probing strategy (chemical crosslinking), site-directed mutagenesis, genetic interaction analysis","journal":"RNA (New York, N.Y.)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — novel crosslinking approach combined with mutagenesis and genetic interactions, single lab","pmids":["29487104"],"is_preprint":false},{"year":2018,"finding":"PRPF8 knockdown impairs hypoxia-induced mitophagy by causing aberrant mRNA splicing of ULK1, which initiates autophagy; the RP-associated PRPF8 mutant R2310K is defective in regulating mitophagy.","method":"RNAi screen with mt-Keima fluorescent reporter, PRPF8 knockdown, RT-PCR of ULK1 splicing, mitophagy assays","journal":"Autophagy","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — RNAi screen with defined reporter, splicing verification, and functional mitophagy assay, single lab","pmids":["30103670"],"is_preprint":false},{"year":2019,"finding":"Two prp-8 alleles in C. elegans identified by genetic screen alter cryptic splice site usage frequency; complementary genetic and structural analyses in yeast implicate these alleles in stability of the spliceosome catalytic core; high-throughput mRNA sequencing shows overall alternative splicing patterns are relatively unchanged despite effects on cryptic splicing.","method":"C. elegans genetic screen, RNA-seq (high-throughput mRNA sequencing), yeast structural/genetic analysis","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic screen combined with genome-wide RNA-seq and cross-species analysis, single lab","pmids":["30674666"],"is_preprint":false},{"year":2021,"finding":"The evolutionarily conserved protein Ecdysoneless (Ecd) chaperones Prp8 delivery to the forming U5 snRNP in the cytoplasm; Ecd deficiency leads to reduced Prp8 protein levels and compromised U5 snRNP biogenesis, causing loss of splicing fidelity; SmD3 was identified as a novel interaction partner of Ecd.","method":"Drosophila genetics, proteomic approaches (co-immunoprecipitation/mass spectrometry), Western blot, splicing analysis","journal":"Nucleic acids research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetics combined with proteomics and functional splicing assays, single lab","pmids":["33444449"],"is_preprint":false},{"year":2023,"finding":"PRPF8 silencing in hepatocellular carcinoma cells modulates fibronectin (FN1) splicing by promoting exclusion of exon 40.2, which reduces FAK/AKT phosphorylation and blunts stress fiber formation, thereby decreasing invasive capacity; CLIPseq analysis shows PRPF8 binds preferentially to exons of protein-coding genes.","method":"siRNA knockdown, RNA-seq, CLIPseq, Western blot (FAK/AKT phosphorylation), invasion assay, xenograft tumor growth","journal":"Experimental & molecular medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function combined with CLIPseq and defined signaling pathway readout, single lab","pmids":["36609600"],"is_preprint":false},{"year":2024,"finding":"The PRPF8/Brr2 regulatory axis controls 5' splice site (5'SS) selection; the heterozygous PRPF8 c.6926 A>C (p.H2309P) RP mutation impairs alternative splicing and weak/suboptimal 5'SS selection, enhances cryptic splicing in ciliary/retinal-specific transcripts, alters PRPF8 interaction with U6 snRNA, and causes accumulation of active spliceosomes and poly(A)+ mRNAs at nuclear periphery splicing clusters in photoreceptors.","method":"Patient iPSC-derived retinal cells, transcriptomics, proteomics, co-immunoprecipitation (PRPF8-U6 snRNA), immunofluorescence of nuclear speckles/poly(A)+ mRNA","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — comprehensive multi-omics in patient-derived disease-relevant cells combined with molecular and cell biological validation using multiple orthogonal methods","pmids":["38605034"],"is_preprint":false}],"current_model":"PRPF8 (hPrp8/Prp8) is the central scaffolding protein of the spliceosome catalytic core, occupying a large cavity formed by its reverse-transcriptase-like, RNaseH, and Jab1/MPN domains that directly contacts both splice sites, the branch point, and U2, U5, and U6 snRNAs throughout both catalytic steps; it regulates spliceosome activation by controlling the timing of U4/U6 unwinding through both stimulatory and inhibitory interactions with the Brr2 helicase (inserting its C-terminal tail into Brr2's RNA-binding tunnel to reversibly block its activity), coordinates the activities of Prp28 and Snu114 GTPase, undergoes a conformational switch in its RNaseH domain between the two catalytic steps that unmasks a metal-binding site required for exon ligation, and its C-terminal Jab1/MPN domain acts as a protein-protein interaction platform whose disruption by RP13-associated mutations weakens Brr2/Snu114 binding, impairs U5 snRNP biogenesis and nuclear import, and causes defects in 5' splice site selection and the first-to-second step transition underlying retinal degeneration."},"narrative":{"mechanistic_narrative":"PRPF8 (hPrp8/Prp8) is the central scaffolding protein of the spliceosome catalytic core, an essential, RNA-free component of the U5 snRNP and U4/U5/U6 tri-snRNP that remains stably associated with the spliceosome throughout both transesterification steps and into post-splicing intron complexes [PMID:2479028, PMID:2138328, PMID:9774689]. It is a core factor of both the major U2-dependent and minor U12-dependent spliceosomes [PMID:10411133], and makes direct, largely non-sequence-specific contacts with the pre-mRNA at the 5' splice site, branch point, and 3' splice site as well as with U5 and U6 snRNAs, positioning the reactive exons and intron termini within the active site [PMID:1945827, PMID:7781612, PMID:10444595, PMID:10444596, PMID:26393790]. Crystallographic and biochemical work showed that its central region is built from reverse-transcriptase-like, endonuclease-like, and RNaseH-like domains that together form a large cavity accommodating the splice sites and branch-point crosslink — the catalytic cavity of the spliceosome [PMID:18836455, PMID:23354046] — while its C-terminal Jab1/MPN domain is a catalytically dead pseudoenzyme fold serving as a protein-protein interaction platform [PMID:17317632]. PRPF8 governs the timing of spliceosome activation by regulating the Brr2 helicase that unwinds U4/U6: its RNaseH domain binds the single-stranded U4 region and blocks Brr2 loading, its C-terminal tail inserts into Brr2's RNA-binding tunnel to competitively repress its ATPase and unwinding activities, and its Jab1/MPN domain binds the Brr2 helicase cassette and stimulates activity, coupling 5' splice site recognition to U4/U6 unwinding [PMID:10024880, PMID:19098916, PMID:23124066, PMID:23727230, PMID:23704370]. A conformational switch in its RNaseH domain between the two catalytic steps unmasks a metal-binding site required for exon ligation, establishing PRPF8 as a metalloprotein that controls the first-to-second step transition [PMID:23686287, PMID:28416677]. PRPF8 also coordinates U5 snRNP biogenesis: it carries a nuclear localization signal required for import of a Brr2-free precursor U5 snRNP, with Aar2 and Brr2 binding the Jab1/MPN domain mutually exclusively to drive the cytoplasm-to-nucleus maturation switch [PMID:17934474, PMID:23727230], and its delivery to the nascent U5 snRNP is chaperoned by Ecdysoneless [PMID:33444449]. Transcriptome-wide, PRPF8 is required for efficient splicing of introns with weak 5' splice sites and for fidelity of 5' splice site selection [PMID:26392272, PMID:38605034]. Heterozygous missense mutations clustered at the C-terminus cause autosomal dominant retinitis pigmentosa (RP13) by weakening Brr2/Snu114 binding, impairing U5 snRNP biogenesis and Brr2 repression, and disrupting 5' splice site selection and the first-to-second step transition in photoreceptors [PMID:11468273, PMID:17317632, PMID:19098916, PMID:23704370, PMID:38605034].","teleology":[{"year":1990,"claim":"Establishing that the conserved ~220 kDa Prp8 protein is a stable spliceosome component that directly touches pre-mRNA defined it as a core catalytic-machinery factor rather than a transient assembly factor.","evidence":"Gradient fractionation, immunoprecipitation, and UV-crosslinking with anti-PRP8 antisera in HeLa extracts; stable association across both splicing steps","pmids":["2479028","2138328","2139226"],"confidence":"Medium","gaps":["Crosslink sites on pre-mRNA not mapped at nucleotide resolution","No structural basis for RNA contact"]},{"year":1991,"claim":"Demonstrating ATP-dependent, splicing-competence-restricted crosslinking of Prp8 to pre-mRNA showed its contacts are functionally coupled to the catalytic reaction, not nonspecific binding.","evidence":"UV-crosslinking with 5'SS and branchpoint mutant substrates in yeast in vitro splicing","pmids":["1945827"],"confidence":"Medium","gaps":["Which Prp8 domain contacts RNA was unknown","Did not distinguish direct catalytic role from scaffolding"]},{"year":1992,"claim":"Showing Prp8 is required for stable tri-snRNP formation and snRNA stability established its essential role in spliceosome assembly upstream of catalysis.","evidence":"Genetic depletion, ts inactivation, and antibody inhibition with sedimentation analysis in yeast","pmids":["1396567"],"confidence":"High","gaps":["Mechanism of tri-snRNP stabilization unresolved","snRNA decline could be indirect"]},{"year":1995,"claim":"Mapping Prp8 contacts to exon nucleotides at both splice sites established it as an aligner that positions exons against U5 snRNA at the active site through both catalytic steps.","evidence":"Site-specific 4-thiouridine UV-crosslinking with mutant and duplicated splice sites","pmids":["7781612","7885825"],"confidence":"High","gaps":["No structural model of the alignment","Branchpoint-3'SS interaction timing only inferred"]},{"year":1999,"claim":"Genetic suppressor and crosslinking studies placed Prp8 at the heart of splice-site tertiary interactions and identified it as a controller of the timing of U4/U6 unwinding-driven activation.","evidence":"Allele-specific suppression of 5'GU, 3'YAG, and U6 ACAGAG mutations; prp8-201 suppression of U4-cs1 unwinding block; mapping of the 5'SS crosslink to hPrp8 C-terminus","pmids":["10444595","10444596","10024880","10024169","10411133"],"confidence":"High","gaps":["Molecular mechanism linking Prp8 to unwinding timing not yet biochemical","Allosteric model untested structurally"]},{"year":1998,"claim":"Reconstituting an RNA-free Prp8/Snu114/Brr2/U5-40K core complex defined Prp8 as the central hub binding its key protein partners directly.","evidence":"Chaotropic dissociation of U5 snRNP with sedimentation and binding analysis","pmids":["9774689"],"confidence":"Medium","gaps":["Binding interfaces not mapped","Stoichiometry and direct vs. bridged contacts uncertain"]},{"year":2002,"claim":"Allele-specific genetic dissection assigned distinct Prp8 regions to interactions with Prp28, Brr2, U6 RNA and U1 release, showing Prp8 coordinates multiple activation steps from a single scaffold.","evidence":"Large-scale suppressor screens, yeast two-hybrid, and allele-specific epistasis with multiple splicing factors","pmids":["10924465","12087126","16431982"],"confidence":"Medium","gaps":["Genetic interactions did not establish direct physical contacts","No structural assignment of the mapped regions"]},{"year":2001,"claim":"Identification of clustered C-terminal missense mutations causing autosomal dominant retinitis pigmentosa connected this ubiquitous splicing factor to a tissue-specific degenerative disease.","evidence":"Positional cloning and cosegregation across multiple RP13 families","pmids":["11468273"],"confidence":"High","gaps":["Mechanism of retinal specificity unexplained","Functional consequence of mutations not yet defined"]},{"year":2008,"claim":"Crystal structures of the RNaseH-fold domain IV, the beta-finger, and the Jab1/MPN domain provided the physical framework explaining how Prp8 contacts catalytic-core RNAs and partners and how RP mutations act.","evidence":"X-ray crystallography of domain IV, beta-finger, and the Jab1/MPN-like C-terminal domain combined with RNA-binding, mutagenesis, and yeast two-hybrid","pmids":["18836455","18779563","17317632"],"confidence":"High","gaps":["Full-length architecture still missing","Catalytic role vs. scaffolding role of metal site not yet shown"]},{"year":2008,"claim":"Linking opposing prp8 allele classes and the beta-finger to the first-vs-second step equilibrium established Prp8 as an active regulator of the catalytic step transition.","evidence":"Genetic epistasis of prp8 alleles with U6 and Prp16 ATPase mutations plus beta-finger mutagenesis","pmids":["17486100","18779563"],"confidence":"Medium","gaps":["Structural basis of the conformational switch not yet resolved","How Prp8 senses step transition unclear"]},{"year":2008,"claim":"In vitro reconstitution showed Prp8 C-terminal fragments both stimulate and inhibit Brr2-driven U4/U6 unwinding, and that RP mutants fail to stimulate — directly tying disease alleles to Brr2 regulation.","evidence":"In vitro U4/U6 unwinding and ATPase assays with recombinant Prp8 fragments and RP variants; EM of tri-snRNP","pmids":["19098916","18953335"],"confidence":"High","gaps":["Atomic basis of stimulation vs. inhibition not yet structural","In vivo balance of the two activities unquantified"]},{"year":2013,"claim":"Landmark crystal structures of Prp8 in complex with Aar2 and Brr2 revealed the RT/endonuclease/RNaseH cavity as the spliceosome active site and defined the Jab1/MPN–Brr2 interface, mechanistically explaining U5 maturation and Brr2 control.","evidence":"X-ray crystallography of Prp8–Aar2 and Brr2–Jab1/MPN complexes with mapping of suppressors and RP mutations; C-terminal tail insertion into the Brr2 RNA tunnel","pmids":["23354046","23727230","23704370"],"confidence":"High","gaps":["Conformational states during catalysis inferred, not captured","Dynamics of tail insertion/release in the assembled spliceosome unresolved"]},{"year":2013,"claim":"Demonstrating that the RNaseH domain switches conformation to unmask a metal-binding site, and binds single-stranded U4 to block Brr2 loading, defined Prp8 as a metalloprotein that gates both exon ligation and activation timing.","evidence":"Metal-binding and conformational assays with step-specific alleles; RNA binding plus crosslinking-MS and in vitro unwinding inhibition","pmids":["23686287","23124066"],"confidence":"High","gaps":["Identity of catalytic vs. structural metal not fully defined","Whether the metal directly participates in chemistry untested"]},{"year":2013,"claim":"In vivo CLIP/CRAC extended Prp8 RNA contacts to U1 and U2 snRNAs and linked the U1 interaction to tri-snRNP recruitment, broadening its role to early assembly.","evidence":"CLIP/CRAC sequencing with functional analysis of U1-interaction mutants in yeast","pmids":["23393194"],"confidence":"Medium","gaps":["Direct vs. indirect U1/U2 contacts not separated","Structural basis of U1 contact unknown"]},{"year":2015,"claim":"Transcriptome-wide depletion studies established that Prp8 contacts at the exon-intron junction trigger tri-snRNP integration and that it is preferentially required for introns with weak 5' splice sites.","evidence":"EM with 5'SS oligonucleotide reconstitution; siRNA knockdown with RNA-seq, iCLIP, and minigene rescue by 5'SS strengthening","pmids":["26385511","26392272"],"confidence":"High","gaps":["Quantitative rules for 5'SS strength dependence incomplete","Tissue-specific splicing targets not yet defined"]},{"year":2018,"claim":"Crosslinking-guided mutagenesis and structural analysis of the RNaseH Switch loop refined how Prp8–U5 snRNA contacts position pre-mRNA and toggle splicing fidelity.","evidence":"Amino acid probing, Switch-loop structural analysis, in vitro/in vivo reporters and lariat sequencing in yeast","pmids":["29487104","28416677"],"confidence":"High","gaps":["How fidelity classes map to human disease alleles incompletely resolved","Dynamics of the two Switch-loop states in vivo unmeasured"]},{"year":2021,"claim":"Identifying Ecdysoneless as a Prp8 chaperone that delivers it to the forming U5 snRNP completed an assembly pathway whose disruption reduces Prp8 levels and splicing fidelity.","evidence":"Drosophila genetics, co-IP/MS proteomics, and splicing analysis","pmids":["33444449"],"confidence":"Medium","gaps":["Mechanism of chaperone handoff unresolved","Human relevance of Ecd–Prp8 axis not tested directly"]},{"year":2024,"claim":"Patient iPSC-derived retinal cells linked the PRPF8/Brr2 axis and a specific RP mutation to defective weak/cryptic 5'SS selection, altered U6 snRNA interaction, and spliceosome/mRNA accumulation, providing a disease mechanism for RP13.","evidence":"Patient iPSC-derived retinal cells with transcriptomics, proteomics, PRPF8–U6 snRNA co-IP, and nuclear speckle/poly(A)+ imaging","pmids":["38605034"],"confidence":"High","gaps":["Why photoreceptors are uniquely vulnerable not fully explained","Causal chain from missplicing to degeneration incomplete"]},{"year":null,"claim":"How PRPF8's well-characterized core splicing functions connect to its reported roles in homology-directed DNA repair, androgen receptor regulation, mitophagy via ULK1 splicing, and cancer-associated FN1 splicing remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["Whether these phenotypes are direct or downstream of global splicing changes is untested","No unifying mechanism links the canonical spliceosome role to these specialized outputs"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0003723","term_label":"RNA binding","supporting_discovery_ids":[3,6,28,30,35,50]},{"term_id":"GO:0140098","term_label":"catalytic activity, acting on RNA","supporting_discovery_ids":[31,43]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[26,30,33,34]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[6,9,22]},{"term_id":"GO:0005198","term_label":"structural molecule activity","supporting_discovery_ids":[5,32]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[0,23]},{"term_id":"GO:0005654","term_label":"nucleoplasm","supporting_discovery_ids":[50]}],"pathway":[{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[0,5,39,50]},{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[6,11,39]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[16,50]}],"complexes":["U5 snRNP","U4/U6.U5 tri-snRNP","spliceosome catalytic core","U12-dependent (minor) spliceosome"],"partners":["BRR2","SNU114","AAR2","PRP28","ECD","U5-40K"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q6P2Q9","full_name":"Pre-mRNA-processing-splicing factor 8","aliases":["220 kDa U5 snRNP-specific protein","PRP8 homolog","Splicing factor Prp8","p220"],"length_aa":2335,"mass_kda":273.6,"function":"Plays a role in pre-mRNA splicing as core component of precatalytic, catalytic and postcatalytic spliceosomal complexes, both of the predominant U2-type spliceosome and the minor U12-type spliceosome (PubMed:10411133, PubMed:11971955, PubMed:28076346, PubMed:28502770, PubMed:28781166, PubMed:29301961, PubMed:29360106, PubMed:29361316, PubMed:30315277, PubMed:30705154, PubMed:30728453). Functions as a scaffold that mediates the ordered assembly of spliceosomal proteins and snRNAs. Required for the assembly of the U4/U6-U5 tri-snRNP complex, a building block of the spliceosome. Functions as a scaffold that positions spliceosomal U2, U5 and U6 snRNAs at splice sites on pre-mRNA substrates, so that splicing can occur. Interacts with both the 5' and the 3' splice site","subcellular_location":"Nucleus; Nucleus speckle","url":"https://www.uniprot.org/uniprotkb/Q6P2Q9/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/PRPF8","classification":"Common Essential","n_dependent_lines":1208,"n_total_lines":1208,"dependency_fraction":1.0},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000174231","cell_line_id":"CID001464","localizations":[{"compartment":"chromatin","grade":3}],"interactors":[{"gene":"CD2BP2","stoichiometry":10.0},{"gene":"EFTUD2","stoichiometry":10.0},{"gene":"EAPP","stoichiometry":10.0},{"gene":"CSE1L","stoichiometry":10.0},{"gene":"SNRPD2","stoichiometry":10.0},{"gene":"SNRNP40","stoichiometry":10.0},{"gene":"DDX23","stoichiometry":10.0},{"gene":"SNRPF","stoichiometry":10.0},{"gene":"USP39","stoichiometry":10.0},{"gene":"AAR2","stoichiometry":10.0}],"url":"https://opencell.sf.czbiohub.org/target/CID001464","total_profiled":1310},"omim":[{"mim_id":"621494","title":"RNA, U6 SMALL NUCLEAR 9; RNU6-9","url":"https://www.omim.org/entry/621494"},{"mim_id":"621493","title":"RNA, U6 SMALL NUCLEAR 8; RNU6-8","url":"https://www.omim.org/entry/621493"},{"mim_id":"621491","title":"RNA, U6 SMALL NUCLEAR 2; RNU6-2","url":"https://www.omim.org/entry/621491"},{"mim_id":"620823","title":"RNA, U4 SMALL NUCLEAR 2; RNU4-2","url":"https://www.omim.org/entry/620823"},{"mim_id":"620822","title":"RNA, U4 SMALL NUCLEAR 1; RNU4-1","url":"https://www.omim.org/entry/620822"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Nucleoplasm","reliability":"Supported"},{"location":"Nuclear speckles","reliability":"Supported"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/PRPF8"},"hgnc":{"alias_symbol":["PRPC8","Prp8","hPrp8","SNRNP220"],"prev_symbol":["RP13"]},"alphafold":{"accession":"Q6P2Q9","domains":[{"cath_id":"-","chopping":"64-131_482-662","consensus_level":"medium","plddt":91.1337,"start":64,"end":662},{"cath_id":"-","chopping":"681-798","consensus_level":"medium","plddt":88.4522,"start":681,"end":798},{"cath_id":"-","chopping":"943-984_1049-1191_1193-1233","consensus_level":"medium","plddt":91.1428,"start":943,"end":1233},{"cath_id":"3.90.1570.40","chopping":"1582-1740","consensus_level":"high","plddt":89.5675,"start":1582,"end":1740},{"cath_id":"3.40.140.10","chopping":"2087-2320","consensus_level":"high","plddt":81.1212,"start":2087,"end":2320}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q6P2Q9","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q6P2Q9-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q6P2Q9-F1-predicted_aligned_error_v6.png","plddt_mean":84.94},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=PRPF8","jax_strain_url":"https://www.jax.org/strain/search?query=PRPF8"},"sequence":{"accession":"Q6P2Q9","fasta_url":"https://rest.uniprot.org/uniprotkb/Q6P2Q9.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q6P2Q9/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q6P2Q9"}},"corpus_meta":[{"pmid":"15840809","id":"PMC_15840809","title":"Prp8 protein: at the heart of the spliceosome.","date":"2005","source":"RNA (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/15840809","citation_count":301,"is_preprint":false},{"pmid":"11468273","id":"PMC_11468273","title":"Mutations in the pre-mRNA splicing factor gene PRPC8 in autosomal dominant retinitis pigmentosa (RP13).","date":"2001","source":"Human molecular genetics","url":"https://pubmed.ncbi.nlm.nih.gov/11468273","citation_count":235,"is_preprint":false},{"pmid":"23354046","id":"PMC_23354046","title":"Crystal structure of Prp8 reveals active site cavity of the spliceosome.","date":"2013","source":"Nature","url":"https://pubmed.ncbi.nlm.nih.gov/23354046","citation_count":188,"is_preprint":false},{"pmid":"7781612","id":"PMC_7781612","title":"Extensive interactions of PRP8 protein with the 5' and 3' splice sites during splicing suggest a role in stabilization of exon alignment by U5 snRNA.","date":"1995","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/7781612","citation_count":149,"is_preprint":false},{"pmid":"9774689","id":"PMC_9774689","title":"The human U5-220kD protein (hPrp8) forms a stable RNA-free complex with several U5-specific proteins, including an RNA unwindase, a homologue of ribosomal elongation factor EF-2, and a novel WD-40 protein.","date":"1998","source":"Molecular and cellular biology","url":"https://pubmed.ncbi.nlm.nih.gov/9774689","citation_count":124,"is_preprint":false},{"pmid":"17317632","id":"PMC_17317632","title":"Structure of a multipartite protein-protein interaction domain in splicing factor prp8 and its link to retinitis pigmentosa.","date":"2007","source":"Molecular cell","url":"https://pubmed.ncbi.nlm.nih.gov/17317632","citation_count":118,"is_preprint":false},{"pmid":"23704370","id":"PMC_23704370","title":"Inhibition of RNA helicase Brr2 by the C-terminal tail of the spliceosomal protein Prp8.","date":"2013","source":"Science (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/23704370","citation_count":118,"is_preprint":false},{"pmid":"24781015","id":"PMC_24781015","title":"PRPF8 defects cause missplicing in myeloid malignancies.","date":"2014","source":"Leukemia","url":"https://pubmed.ncbi.nlm.nih.gov/24781015","citation_count":105,"is_preprint":false},{"pmid":"19098916","id":"PMC_19098916","title":"ATP-dependent unwinding of U4/U6 snRNAs by the Brr2 helicase requires the C terminus of Prp8.","date":"2008","source":"Nature structural & molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/19098916","citation_count":105,"is_preprint":false},{"pmid":"10444595","id":"PMC_10444595","title":"Allele-specific genetic interactions between Prp8 and RNA active site residues suggest a function for Prp8 at the catalytic core of the spliceosome.","date":"1999","source":"Genes & development","url":"https://pubmed.ncbi.nlm.nih.gov/10444595","citation_count":104,"is_preprint":false},{"pmid":"10444596","id":"PMC_10444596","title":"Functional interactions of Prp8 with both splice sites at the spliceosomal catalytic center.","date":"1999","source":"Genes & development","url":"https://pubmed.ncbi.nlm.nih.gov/10444596","citation_count":103,"is_preprint":false},{"pmid":"1996139","id":"PMC_1996139","title":"A suppressor of a yeast splicing mutation (prp8-1) encodes a putative ATP-dependent RNA helicase.","date":"1991","source":"Nature","url":"https://pubmed.ncbi.nlm.nih.gov/1996139","citation_count":101,"is_preprint":false},{"pmid":"10024880","id":"PMC_10024880","title":"Splicing factor Prp8 governs U4/U6 RNA unwinding during activation of the spliceosome.","date":"1999","source":"Molecular cell","url":"https://pubmed.ncbi.nlm.nih.gov/10024880","citation_count":92,"is_preprint":false},{"pmid":"10411133","id":"PMC_10411133","title":"The human Prp8 protein is a component of both U2- and U12-dependent spliceosomes.","date":"1999","source":"RNA (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/10411133","citation_count":86,"is_preprint":false},{"pmid":"17934474","id":"PMC_17934474","title":"prp8 mutations that cause human retinitis pigmentosa lead to a U5 snRNP maturation defect in yeast.","date":"2007","source":"Nature structural & molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/17934474","citation_count":85,"is_preprint":false},{"pmid":"1396567","id":"PMC_1396567","title":"Roles of PRP8 protein in the assembly of splicing complexes.","date":"1992","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/1396567","citation_count":83,"is_preprint":false},{"pmid":"8725222","id":"PMC_8725222","title":"Mutagenesis of the yeast gene PRP8 reveals domains governing the specificity and fidelity of 3' splice site selection.","date":"1996","source":"Genetics","url":"https://pubmed.ncbi.nlm.nih.gov/8725222","citation_count":83,"is_preprint":false},{"pmid":"12714658","id":"PMC_12714658","title":"Mutations in the pre-mRNA splicing-factor genes PRPF3, PRPF8, and PRPF31 in Spanish families with autosomal dominant retinitis pigmentosa.","date":"2003","source":"Investigative ophthalmology & visual science","url":"https://pubmed.ncbi.nlm.nih.gov/12714658","citation_count":82,"is_preprint":false},{"pmid":"10024169","id":"PMC_10024169","title":"The C-terminal region of hPrp8 interacts with the conserved GU dinucleotide at the 5' splice site.","date":"1999","source":"RNA (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/10024169","citation_count":81,"is_preprint":false},{"pmid":"17486100","id":"PMC_17486100","title":"Opposing classes of prp8 alleles modulate the transition between the catalytic steps of pre-mRNA splicing.","date":"2007","source":"Nature structural & molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/17486100","citation_count":81,"is_preprint":false},{"pmid":"26392272","id":"PMC_26392272","title":"Regulation of constitutive and alternative mRNA splicing across the human transcriptome by PRPF8 is determined by 5' splice site strength.","date":"2015","source":"Genome biology","url":"https://pubmed.ncbi.nlm.nih.gov/26392272","citation_count":77,"is_preprint":false},{"pmid":"23727230","id":"PMC_23727230","title":"Structural basis of Brr2-Prp8 interactions and implications for U5 snRNP biogenesis and the spliceosome active site.","date":"2013","source":"Structure (London, England : 1993)","url":"https://pubmed.ncbi.nlm.nih.gov/23727230","citation_count":77,"is_preprint":false},{"pmid":"2479028","id":"PMC_2479028","title":"The mammalian analogue of the yeast PRP8 splicing protein is present in the U4/5/6 small nuclear ribonucleoprotein particle and the spliceosome.","date":"1989","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/2479028","citation_count":73,"is_preprint":false},{"pmid":"10924465","id":"PMC_10924465","title":"Suppressors of a cold-sensitive mutation in yeast U4 RNA define five domains in the splicing factor Prp8 that influence spliceosome activation.","date":"2000","source":"Genetics","url":"https://pubmed.ncbi.nlm.nih.gov/10924465","citation_count":70,"is_preprint":false},{"pmid":"18779563","id":"PMC_18779563","title":"Crystal structure of the beta-finger domain of Prp8 reveals analogy to ribosomal proteins.","date":"2008","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/18779563","citation_count":68,"is_preprint":false},{"pmid":"23124066","id":"PMC_23124066","title":"The Prp8 RNase H-like domain inhibits Brr2-mediated U4/U6 snRNA unwinding by blocking Brr2 loading onto the U4 snRNA.","date":"2012","source":"Genes & development","url":"https://pubmed.ncbi.nlm.nih.gov/23124066","citation_count":68,"is_preprint":false},{"pmid":"8862522","id":"PMC_8862522","title":"A connection between pre-mRNA splicing and the cell cycle in fission yeast: cdc28+ is allelic with prp8+ and encodes an RNA-dependent ATPase/helicase.","date":"1996","source":"Molecular biology of the cell","url":"https://pubmed.ncbi.nlm.nih.gov/8862522","citation_count":67,"is_preprint":false},{"pmid":"21441348","id":"PMC_21441348","title":"Prp8, the pivotal protein of the spliceosomal catalytic center, evolved from a retroelement-encoded reverse transcriptase.","date":"2011","source":"RNA (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/21441348","citation_count":67,"is_preprint":false},{"pmid":"12087126","id":"PMC_12087126","title":"Distinct domains of splicing factor Prp8 mediate different aspects of spliceosome activation.","date":"2002","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/12087126","citation_count":66,"is_preprint":false},{"pmid":"7885825","id":"PMC_7885825","title":"Interaction of the yeast splicing factor PRP8 with substrate RNA during both steps of splicing.","date":"1995","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/7885825","citation_count":62,"is_preprint":false},{"pmid":"18953335","id":"PMC_18953335","title":"Localization of Prp8, Brr2, Snu114 and U4/U6 proteins in the yeast tri-snRNP by electron microscopy.","date":"2008","source":"Nature structural & molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/18953335","citation_count":60,"is_preprint":false},{"pmid":"32690086","id":"PMC_32690086","title":"circRNA-UBAP2 promotes the proliferation and inhibits apoptosis of ovarian cancer though miR-382-5p/PRPF8 axis.","date":"2020","source":"Journal of ovarian research","url":"https://pubmed.ncbi.nlm.nih.gov/32690086","citation_count":57,"is_preprint":false},{"pmid":"18836455","id":"PMC_18836455","title":"Structural elucidation of a PRP8 core domain from the heart of the spliceosome.","date":"2008","source":"Nature structural & molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/18836455","citation_count":57,"is_preprint":false},{"pmid":"2138328","id":"PMC_2138328","title":"Affinity purification of spliceosomes reveals that the precursor RNA processing protein PRP8, a protein in the U5 small nuclear ribonucleoprotein particle, is a component of yeast spliceosomes.","date":"1990","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/2138328","citation_count":56,"is_preprint":false},{"pmid":"2139226","id":"PMC_2139226","title":"A mammalian protein of 220 kDa binds pre-mRNAs in the spliceosome: a potential homologue of the yeast PRP8 protein.","date":"1990","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/2139226","citation_count":56,"is_preprint":false},{"pmid":"17473007","id":"PMC_17473007","title":"Crystal structure of the C-terminal domain of splicing factor Prp8 carrying retinitis pigmentosa mutants.","date":"2007","source":"Protein science : a publication of the Protein Society","url":"https://pubmed.ncbi.nlm.nih.gov/17473007","citation_count":53,"is_preprint":false},{"pmid":"16431982","id":"PMC_16431982","title":"Dissection of Prp8 protein defines multiple interactions with crucial RNA sequences in the catalytic core of the spliceosome.","date":"2006","source":"RNA (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/16431982","citation_count":53,"is_preprint":false},{"pmid":"23714367","id":"PMC_23714367","title":"In vivo mutation of pre-mRNA processing factor 8 (Prpf8) affects transcript splicing, cell survival and myeloid differentiation.","date":"2013","source":"FEBS letters","url":"https://pubmed.ncbi.nlm.nih.gov/23714367","citation_count":52,"is_preprint":false},{"pmid":"14688266","id":"PMC_14688266","title":"Motifs IV and V in the DEAH box splicing factor Prp22 are important for RNA unwinding, and helicase-defective Prp22 mutants are suppressed by Prp8.","date":"2003","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/14688266","citation_count":52,"is_preprint":false},{"pmid":"9250687","id":"PMC_9250687","title":"Trans mRNA splicing in trypanosomes: cloning and analysis of a PRP8-homologous gene from Trypanosoma brucei provides evidence for a U5-analogous RNP.","date":"1997","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/9250687","citation_count":51,"is_preprint":false},{"pmid":"24968230","id":"PMC_24968230","title":"Cytoplasmic viral RNA-dependent RNA polymerase disrupts the intracellular splicing machinery by entering the nucleus and interfering with Prp8.","date":"2014","source":"PLoS pathogens","url":"https://pubmed.ncbi.nlm.nih.gov/24968230","citation_count":51,"is_preprint":false},{"pmid":"7838707","id":"PMC_7838707","title":"The budding yeast U5 snRNP Prp8 is a highly conserved protein which links RNA splicing with cell cycle progression.","date":"1994","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/7838707","citation_count":46,"is_preprint":false},{"pmid":"21283520","id":"PMC_21283520","title":"Temporal and tissue specific regulation of RP-associated splicing factor genes PRPF3, PRPF31 and PRPC8--implications in the pathogenesis of RP.","date":"2011","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/21283520","citation_count":44,"is_preprint":false},{"pmid":"30103670","id":"PMC_30103670","title":"Autosomal dominant retinitis pigmentosa-associated gene PRPF8 is essential for hypoxia-induced mitophagy through regulating ULK1 mRNA splicing.","date":"2018","source":"Autophagy","url":"https://pubmed.ncbi.nlm.nih.gov/30103670","citation_count":43,"is_preprint":false},{"pmid":"22039234","id":"PMC_22039234","title":"Autosomal dominant retinitis pigmentosa with intrafamilial variability and incomplete penetrance in two families carrying mutations in PRPF8.","date":"2011","source":"Investigative ophthalmology & visual science","url":"https://pubmed.ncbi.nlm.nih.gov/22039234","citation_count":41,"is_preprint":false},{"pmid":"10628969","id":"PMC_10628969","title":"Extensive genetic interactions between PRP8 and PRP17/CDC40, two yeast genes involved in pre-mRNA splicing and cell cycle progression.","date":"2000","source":"Genetics","url":"https://pubmed.ncbi.nlm.nih.gov/10628969","citation_count":40,"is_preprint":false},{"pmid":"1945827","id":"PMC_1945827","title":"The yeast PRP8 protein interacts directly with pre-mRNA.","date":"1991","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/1945827","citation_count":38,"is_preprint":false},{"pmid":"16737526","id":"PMC_16737526","title":"The distribution and evolutionary history of the PRP8 intein.","date":"2006","source":"BMC evolutionary biology","url":"https://pubmed.ncbi.nlm.nih.gov/16737526","citation_count":37,"is_preprint":false},{"pmid":"15304322","id":"PMC_15304322","title":"Prp8 intein in fungal pathogens: target for potential antifungal drugs.","date":"2004","source":"FEBS letters","url":"https://pubmed.ncbi.nlm.nih.gov/15304322","citation_count":37,"is_preprint":false},{"pmid":"16540695","id":"PMC_16540695","title":"Assembly of Snu114 into U5 snRNP requires Prp8 and a functional GTPase domain.","date":"2006","source":"RNA (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/16540695","citation_count":36,"is_preprint":false},{"pmid":"31600193","id":"PMC_31600193","title":"Spliceosomal Prp8 intein at the crossroads of protein and RNA splicing.","date":"2019","source":"PLoS biology","url":"https://pubmed.ncbi.nlm.nih.gov/31600193","citation_count":35,"is_preprint":false},{"pmid":"15809009","id":"PMC_15809009","title":"The PRP8 inteins in Cryptococcus are a source of phylogenetic and epidemiological information.","date":"2005","source":"Fungal genetics and biology : FG & B","url":"https://pubmed.ncbi.nlm.nih.gov/15809009","citation_count":34,"is_preprint":false},{"pmid":"23686287","id":"PMC_23686287","title":"A conformational switch in PRP8 mediates metal ion coordination that promotes pre-mRNA exon ligation.","date":"2013","source":"Nature structural & molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/23686287","citation_count":32,"is_preprint":false},{"pmid":"26393790","id":"PMC_26393790","title":"Dynamic Contacts of U2, RES, Cwc25, Prp8 and Prp45 Proteins with the Pre-mRNA Branch-Site and 3' Splice Site during Catalytic Activation and Step 1 Catalysis in Yeast Spliceosomes.","date":"2015","source":"PLoS genetics","url":"https://pubmed.ncbi.nlm.nih.gov/26393790","citation_count":31,"is_preprint":false},{"pmid":"36609600","id":"PMC_36609600","title":"PRPF8 increases the aggressiveness of hepatocellular carcinoma by regulating FAK/AKT pathway via fibronectin 1 splicing.","date":"2023","source":"Experimental & molecular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/36609600","citation_count":28,"is_preprint":false},{"pmid":"16133344","id":"PMC_16133344","title":"Multiple genetic and biochemical interactions of Brr2, Prp8, Prp31, Prp1 and Prp4 kinase suggest a function in the control of the activation of spliceosomes in Schizosaccharomyces pombe.","date":"2005","source":"Current genetics","url":"https://pubmed.ncbi.nlm.nih.gov/16133344","citation_count":27,"is_preprint":false},{"pmid":"33397721","id":"PMC_33397721","title":"Small-molecule inhibitors for the Prp8 intein as antifungal agents.","date":"2021","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/33397721","citation_count":27,"is_preprint":false},{"pmid":"26968627","id":"PMC_26968627","title":"Prp8 retinitis pigmentosa mutants cause defects in the transition between the catalytic steps of splicing.","date":"2016","source":"RNA (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/26968627","citation_count":26,"is_preprint":false},{"pmid":"11565750","id":"PMC_11565750","title":"Evidence for a role of Sky1p-mediated phosphorylation in 3' splice site recognition involving both Prp8 and Prp17/Slu4.","date":"2001","source":"RNA (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/11565750","citation_count":26,"is_preprint":false},{"pmid":"31223062","id":"PMC_31223062","title":"Cisplatin protects mice from challenge of Cryptococcus neoformans by targeting the Prp8 intein.","date":"2019","source":"Emerging microbes & infections","url":"https://pubmed.ncbi.nlm.nih.gov/31223062","citation_count":24,"is_preprint":false},{"pmid":"9547260","id":"PMC_9547260","title":"Cloning and characterization of a human DEAH-box RNA helicase, a functional homolog of fission yeast Cdc28/Prp8.","date":"1998","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/9547260","citation_count":23,"is_preprint":false},{"pmid":"33994920","id":"PMC_33994920","title":"Mutant PRPF8 Causes Widespread Splicing Changes in Spliceosome Components in Retinitis Pigmentosa Patient iPSC-Derived RPE Cells.","date":"2021","source":"Frontiers in neuroscience","url":"https://pubmed.ncbi.nlm.nih.gov/33994920","citation_count":22,"is_preprint":false},{"pmid":"23393194","id":"PMC_23393194","title":"Comprehensive in vivo RNA-binding site analyses reveal a role of Prp8 in spliceosomal assembly.","date":"2013","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/23393194","citation_count":21,"is_preprint":false},{"pmid":"28416677","id":"PMC_28416677","title":"Structural toggle in the RNaseH domain of Prp8 helps balance splicing fidelity and catalytic efficiency.","date":"2017","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/28416677","citation_count":21,"is_preprint":false},{"pmid":"11910553","id":"PMC_11910553","title":"Clinical characterization, linkage analysis, and PRPC8 mutation analysis of a family with autosomal dominant retinitis pigmentosa type 13 (RP13).","date":"2002","source":"Ophthalmic genetics","url":"https://pubmed.ncbi.nlm.nih.gov/11910553","citation_count":20,"is_preprint":false},{"pmid":"8782056","id":"PMC_8782056","title":"A new family linked to the RP13 locus for autosomal dominant retinitis pigmentosa on distal 17p.","date":"1996","source":"Journal of medical genetics","url":"https://pubmed.ncbi.nlm.nih.gov/8782056","citation_count":19,"is_preprint":false},{"pmid":"8571961","id":"PMC_8571961","title":"Map refinement of locus RP13 to human chromosome 17p13.3 in a second family with autosomal dominant retinitis pigmentosa.","date":"1996","source":"American journal of human genetics","url":"https://pubmed.ncbi.nlm.nih.gov/8571961","citation_count":18,"is_preprint":false},{"pmid":"15126168","id":"PMC_15126168","title":"Histologic study of retinitis pigmentosa due to a mutation in the RP13 gene (PRPC8): comparison with rhodopsin Pro23His, Cys110Arg, and Glu181Lys.","date":"2004","source":"American journal of ophthalmology","url":"https://pubmed.ncbi.nlm.nih.gov/15126168","citation_count":17,"is_preprint":false},{"pmid":"24231520","id":"PMC_24231520","title":"An unanticipated early function of DEAD-box ATPase Prp28 during commitment to splicing is modulated by U5 snRNP protein Prp8.","date":"2013","source":"RNA (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/24231520","citation_count":17,"is_preprint":false},{"pmid":"8655640","id":"PMC_8655640","title":"The role of PRP8 protein in nuclear pre-mRNA splicing in yeast.","date":"1995","source":"Journal of cell science. Supplement","url":"https://pubmed.ncbi.nlm.nih.gov/8655640","citation_count":16,"is_preprint":false},{"pmid":"26371515","id":"PMC_26371515","title":"Splicing Factor Prp8 Interacts With NES(AR) and Regulates Androgen Receptor in Prostate Cancer Cells.","date":"2015","source":"Molecular endocrinology (Baltimore, Md.)","url":"https://pubmed.ncbi.nlm.nih.gov/26371515","citation_count":16,"is_preprint":false},{"pmid":"38605034","id":"PMC_38605034","title":"PRPF8-mediated dysregulation of hBrr2 helicase disrupts human spliceosome kinetics and 5´-splice-site selection causing tissue-specific defects.","date":"2024","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/38605034","citation_count":15,"is_preprint":false},{"pmid":"29212152","id":"PMC_29212152","title":"PRPF8 is important for BRCA1-mediated homologous recombination.","date":"2017","source":"Oncotarget","url":"https://pubmed.ncbi.nlm.nih.gov/29212152","citation_count":15,"is_preprint":false},{"pmid":"29487104","id":"PMC_29487104","title":"Prp8 positioning of U5 snRNA is linked to 5' splice site recognition.","date":"2018","source":"RNA (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/29487104","citation_count":15,"is_preprint":false},{"pmid":"17544410","id":"PMC_17544410","title":"Sequence requirements for splicing by the Cne PRP8 intein.","date":"2007","source":"FEBS letters","url":"https://pubmed.ncbi.nlm.nih.gov/17544410","citation_count":15,"is_preprint":false},{"pmid":"29635373","id":"PMC_29635373","title":"Structural dynamics of the N-terminal domain and the Switch loop of Prp8 during spliceosome assembly and activation.","date":"2018","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/29635373","citation_count":14,"is_preprint":false},{"pmid":"33444449","id":"PMC_33444449","title":"Ecd promotes U5 snRNP maturation and Prp8 stability.","date":"2021","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/33444449","citation_count":14,"is_preprint":false},{"pmid":"23665464","id":"PMC_23665464","title":"PRP8 intein in cryptic species of Histoplasma capsulatum: evolution and phylogeny.","date":"2013","source":"Infection, genetics and evolution : journal of molecular epidemiology and evolutionary genetics in infectious diseases","url":"https://pubmed.ncbi.nlm.nih.gov/23665464","citation_count":14,"is_preprint":false},{"pmid":"16544141","id":"PMC_16544141","title":"Protein splicing of PRP8 mini-inteins from species of the genus Penicillium.","date":"2006","source":"Applied microbiology and biotechnology","url":"https://pubmed.ncbi.nlm.nih.gov/16544141","citation_count":14,"is_preprint":false},{"pmid":"28707069","id":"PMC_28707069","title":"Variants in the PRPF8 Gene are Associated with Glaucoma.","date":"2017","source":"Molecular neurobiology","url":"https://pubmed.ncbi.nlm.nih.gov/28707069","citation_count":13,"is_preprint":false},{"pmid":"36359856","id":"PMC_36359856","title":"PRP8-Induced CircMaml2 Facilitates the Healing of the Intestinal Mucosa via Recruiting PTBP1 and Regulating Sec62.","date":"2022","source":"Cells","url":"https://pubmed.ncbi.nlm.nih.gov/36359856","citation_count":13,"is_preprint":false},{"pmid":"29087248","id":"PMC_29087248","title":"Variability in clinical phenotypes of PRPF8-linked autosomal dominant retinitis pigmentosa correlates with differential PRPF8/SNRNP200 interactions.","date":"2017","source":"Ophthalmic genetics","url":"https://pubmed.ncbi.nlm.nih.gov/29087248","citation_count":13,"is_preprint":false},{"pmid":"35124606","id":"PMC_35124606","title":"The role of splicing factor PRPF8 in breast cancer.","date":"2022","source":"Technology and health care : official journal of the European Society for Engineering and Medicine","url":"https://pubmed.ncbi.nlm.nih.gov/35124606","citation_count":12,"is_preprint":false},{"pmid":"35543142","id":"PMC_35543142","title":"Heterozygous variants in PRPF8 are associated with neurodevelopmental disorders.","date":"2022","source":"American journal of medical genetics. Part A","url":"https://pubmed.ncbi.nlm.nih.gov/35543142","citation_count":11,"is_preprint":false},{"pmid":"38049848","id":"PMC_38049848","title":"Altered splicing machinery in lung carcinoids unveils NOVA1, PRPF8 and SRSF10 as novel candidates to understand tumor biology and expand biomarker discovery.","date":"2023","source":"Journal of translational medicine","url":"https://pubmed.ncbi.nlm.nih.gov/38049848","citation_count":11,"is_preprint":false},{"pmid":"18695108","id":"PMC_18695108","title":"Phenotypic expression of a PRPF8 gene mutation in a Large African American family.","date":"2008","source":"Archives of ophthalmology (Chicago, Ill. : 1960)","url":"https://pubmed.ncbi.nlm.nih.gov/18695108","citation_count":11,"is_preprint":false},{"pmid":"38790138","id":"PMC_38790138","title":"Spliceosomic dysregulation in pancreatic cancer uncovers splicing factors PRPF8 and RBMX as novel candidate actionable targets.","date":"2024","source":"Molecular oncology","url":"https://pubmed.ncbi.nlm.nih.gov/38790138","citation_count":11,"is_preprint":false},{"pmid":"23857713","id":"PMC_23857713","title":"The U5 snRNA internal loop 1 is a platform for Brr2, Snu114 and Prp8 protein binding during U5 snRNP assembly.","date":"2013","source":"Journal of cellular biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/23857713","citation_count":11,"is_preprint":false},{"pmid":"18945809","id":"PMC_18945809","title":"Suppressors of the cdc-25.1(gf)-associated intestinal hyperplasia reveal important maternal roles for prp-8 and a subset of splicing factors in C. elegans.","date":"2008","source":"RNA (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/18945809","citation_count":11,"is_preprint":false},{"pmid":"26385511","id":"PMC_26385511","title":"Stable tri-snRNP integration is accompanied by a major structural rearrangement of the spliceosome that is dependent on Prp8 interaction with the 5' splice site.","date":"2015","source":"RNA (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/26385511","citation_count":11,"is_preprint":false},{"pmid":"30360737","id":"PMC_30360737","title":"Mutation Analysis of Pre-mRNA Splicing Genes PRPF31, PRPF8, and SNRNP200 in Chinese Families with Autosomal Dominant Retinitis Pigmentosa.","date":"2018","source":"Current molecular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/30360737","citation_count":10,"is_preprint":false},{"pmid":"30674666","id":"PMC_30674666","title":"Prp8 impacts cryptic but not alternative splicing frequency.","date":"2019","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/30674666","citation_count":10,"is_preprint":false},{"pmid":"37019475","id":"PMC_37019475","title":"Retinitis pigmentosa-associated mutations in mouse Prpf8 cause misexpression of circRNAs and degeneration of cerebellar granule cells.","date":"2023","source":"Life science alliance","url":"https://pubmed.ncbi.nlm.nih.gov/37019475","citation_count":9,"is_preprint":false},{"pmid":"35948057","id":"PMC_35948057","title":"Calcimycin Inhibits Cryptococcus neoformans In Vitro and In Vivo by Targeting the Prp8 Intein Splicing.","date":"2022","source":"ACS infectious diseases","url":"https://pubmed.ncbi.nlm.nih.gov/35948057","citation_count":9,"is_preprint":false},{"pmid":"8436222","id":"PMC_8436222","title":"Detection of a plant protein analogous to the yeast spliceosomal protein, PRP8.","date":"1993","source":"FEBS letters","url":"https://pubmed.ncbi.nlm.nih.gov/8436222","citation_count":9,"is_preprint":false},{"pmid":"20093192","id":"PMC_20093192","title":"Sexual mating of Botrytis cinerea illustrates PRP8 intein HEG activity.","date":"2010","source":"Fungal genetics and biology : FG & B","url":"https://pubmed.ncbi.nlm.nih.gov/20093192","citation_count":9,"is_preprint":false},{"pmid":"23154979","id":"PMC_23154979","title":"Spliceosome activation: U4 is the path, stem I is the goal, and Prp8 is the keeper. Let's cheer for the ATPase Brr2!","date":"2012","source":"Genes & development","url":"https://pubmed.ncbi.nlm.nih.gov/23154979","citation_count":9,"is_preprint":false},{"pmid":"32424050","id":"PMC_32424050","title":"A Drosophila model to study retinitis pigmentosa pathology associated with mutations in the core splicing factor Prp8.","date":"2020","source":"Disease models & mechanisms","url":"https://pubmed.ncbi.nlm.nih.gov/32424050","citation_count":8,"is_preprint":false},{"pmid":"31286362","id":"PMC_31286362","title":"PRP8 Intein in Onygenales: Distribution and Phylogenetic Aspects.","date":"2019","source":"Mycopathologia","url":"https://pubmed.ncbi.nlm.nih.gov/31286362","citation_count":8,"is_preprint":false},{"pmid":"10828595","id":"PMC_10828595","title":"Expression map of human chromosome region 17p13.3, spanning the RP13 dominant retinitis pigmentosa locus, the Miller-Dieker lissencephaly syndrome (MDLS) region, and a putative tumour suppressor locus.","date":"2000","source":"Cytogenetics and cell genetics","url":"https://pubmed.ncbi.nlm.nih.gov/10828595","citation_count":8,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":45767,"output_tokens":11490,"usd":0.154826,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":22505,"output_tokens":6042,"usd":0.131788,"stage2_stop_reason":"end_turn"},"total_usd":0.286614,"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\": 1989,\n      \"finding\": \"The mammalian homolog of yeast PRP8 (hPrp8/p220) is a component of the U4/U5/U6 snRNP complex and of purified spliceosomes, as demonstrated by gradient fractionation and immunoprecipitation with anti-PRP8 antibodies in HeLa nuclear extracts.\",\n      \"method\": \"Gradient fractionation, immunoprecipitation, Western blot\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal immunoprecipitation and gradient fractionation, single lab, two orthogonal methods\",\n      \"pmids\": [\"2479028\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1990,\n      \"finding\": \"PRP8 protein maintains stable association with the spliceosome throughout both steps of the splicing reaction and is present in post-splicing complexes containing the excised intron.\",\n      \"method\": \"Affinity purification of spliceosomes, immunoprecipitation with anti-PRP8 antibodies\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — affinity purification and immunological detection, single lab\",\n      \"pmids\": [\"2138328\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1990,\n      \"finding\": \"A mammalian ~220 kDa protein (p220) is UV-crosslinked to pre-mRNAs under splicing conditions; anti-yeast PRP8 antibodies recognize and immunoprecipitate this UV-crosslinked protein, establishing structural conservation and direct pre-mRNA contact.\",\n      \"method\": \"UV-crosslinking, immunoprecipitation, Western blot with anti-PRP8 antisera\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — UV-crosslinking plus immunoprecipitation, single lab, two orthogonal methods\",\n      \"pmids\": [\"2139226\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1991,\n      \"finding\": \"Yeast PRP8 protein directly contacts pre-mRNA in an ATP-dependent, splicing-specific manner; PRP8 only crosslinks to RNA substrates competent for step 1 of splicing, not to mutant substrates with 5' splice site or branchpoint mutations.\",\n      \"method\": \"UV-crosslinking combined with immunoprecipitation, in vitro splicing assay\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — UV-crosslinking with mutant substrates as controls, single lab\",\n      \"pmids\": [\"1945827\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1991,\n      \"finding\": \"A suppressor of prp8-1 (spp81) encodes a putative ATP-dependent RNA helicase (later identified as Brr2), establishing a genetic interaction between PRP8 and an RNA helicase in the splicing pathway.\",\n      \"method\": \"Extragenic suppressor screen, DNA sequencing, sequence homology analysis\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic suppressor screen with sequence characterization, single lab\",\n      \"pmids\": [\"1996139\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1992,\n      \"finding\": \"Functional PRP8 is required for stable formation of U4/U6.U5 triple snRNP complexes and for assembly of triple snRNPs into spliceosomes; depletion of PRP8 also causes dramatic decline in U4, U5, and U6 snRNA levels.\",\n      \"method\": \"Genetic depletion in vivo, temperature-sensitive inactivation in protoplasts, antibody inhibition in vitro, sedimentation analysis\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — three orthogonal loss-of-function approaches (genetic depletion, ts inactivation, antibody inhibition) in single study\",\n      \"pmids\": [\"1396567\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1995,\n      \"finding\": \"PRP8 protein directly contacts at least eight exon nucleotides at the 5' splice site prior to step 1 of splicing, and at least 13 exon nucleotides plus part of the polypyrimidine tract at the 3' splice site after step 1; these interactions are not sequence-specific. The data support a model where PRP8 stabilizes U5 snRNA interactions with both exons for alignment at the spliceosome active site.\",\n      \"method\": \"4-thiouridine UV-crosslinking with site-specific labeling, analysis of mutant and duplicated splice sites\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — systematic site-specific crosslinking with mutant controls, multiple splice-site positions tested\",\n      \"pmids\": [\"7781612\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1995,\n      \"finding\": \"PRP8 protein can be UV-crosslinked to pre-mRNA in PRP2-depleted spliceosomes stalled before step 1, interacts with substrate RNA fragments at the 5' splice site region and the branchpoint–3' splice site region, with the latter interaction established only after step 1 of splicing.\",\n      \"method\": \"UV-crosslinking combined with PRP8-specific immunoprecipitation and RNase T1 treatment\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — immunoprecipitation of crosslinked RNA fragments with enzymatic mapping, single lab\",\n      \"pmids\": [\"7885825\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1996,\n      \"finding\": \"Mutagenesis of yeast PRP8 identifies two separable functional domains: one governing specificity of 3' splice site selection (uridine tract recognition) and one governing fidelity of 3' splice site utilization (PyAG motif recognition), implicating Prp8p in functional roles at the spliceosome active site during the second catalytic step.\",\n      \"method\": \"Random mutagenesis, genetic selection, allele characterization, in vivo splicing assays\",\n      \"journal\": \"Genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — systematic mutagenesis with functional in vivo assays, single lab\",\n      \"pmids\": [\"8725222\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"Human U5-220 kDa protein (hPrp8) forms a stable, RNA-free complex with the U5-116 kDa EF-2 homologue (Snu114), the 200 kDa RNA unwindase (Brr2), and a novel WD-40 repeat protein (U5-40 kDa); the 220 kDa protein binds simultaneously to the 40 kDa and 116 kDa proteins and probably also to the 200 kDa protein.\",\n      \"method\": \"Chaotropic salt dissociation of U5 snRNP, sedimentation analysis, cDNA cloning, biochemical binding analysis\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — biochemical reconstitution with sedimentation and binding analyses, single lab\",\n      \"pmids\": [\"9774689\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"The C-terminal region of hPrp8 (residues 1894–1898) forms a UV-induced crosslink with the conserved GU dinucleotide at the 5' splice site within spliceosomal complex B, mapping the functional interaction domain.\",\n      \"method\": \"UV-crosslinking, immunoprecipitation, proteolytic mapping, size comparison of crosslinked peptides\",\n      \"journal\": \"RNA (New York, N.Y.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — UV-crosslinking with systematic proteolytic mapping to identify contact residues, single lab\",\n      \"pmids\": [\"10024169\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"Yeast Prp8 mutants can suppress mutations at position 2 of the 5' GU, all positions of the 3' YAG, and position A51 in the U6 ACAGAG motif, implying that Prp8 participates in a tertiary interaction between U6 snRNA and both splice site ends and plays a functional role at the active site of the spliceosome.\",\n      \"method\": \"Genetic suppressor screen, allele-specific suppression analysis, in vivo splicing assays\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — allele-specific genetic epistasis with multiple splice-site targets, replicated across two concurrent independent studies (PMID 10444595 and 10444596)\",\n      \"pmids\": [\"10444595\", \"10444596\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"Mutagenesis of the yeast Prp8 region corresponding to the 5'SS:hPrp8 crosslink identifies alleles that suppress both 5' and 3' splice site mutations, placing Prp8 functional interactions with both splice sites at the later stage of splicing affecting the second catalytic step.\",\n      \"method\": \"Site-directed and random mutagenesis, in vivo suppression assays, combined analysis with U1 suppressor snRNA\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal genetic analyses in single study, corroborated by independent concurrent study\",\n      \"pmids\": [\"10444596\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"A novel Prp8 mutation (prp8-201) suppresses the growth defect of cold-sensitive U4-cs1, which blocks U4/U6 unwinding; wild-type Prp8 triggers U4/U6 RNA unwinding only after correct 5' splice site recognition by the U6 ACAGA box, indicating Prp8 governs the timing of spliceosome activation.\",\n      \"method\": \"Genetic suppressor analysis, in vitro splicing assay with cold-sensitive U4-cs1 block, spliceosomal complex analysis\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic suppressor combined with in vitro splicing and complex analysis, single lab\",\n      \"pmids\": [\"10024880\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"Human Prp8 (hPrp8p) is a core component of both the major U2-dependent and the minor U12-dependent spliceosomes, the first non-Sm factor shown to be common to both spliceosomes.\",\n      \"method\": \"Immunoprecipitation with anti-hPrp8 antibodies, Northern blot analysis of snRNAs\",\n      \"journal\": \"RNA (New York, N.Y.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — immunoprecipitation with snRNA analysis, single lab\",\n      \"pmids\": [\"10411133\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"Large-scale suppressor screen identifies five distinct regions (a–e) of Prp8 that control spliceosome activation; two regions contact U1 snRNP (two-hybrid interaction), another mediates indirect contact; allosteric changes in Prp8 are proposed to initiate activation by disrupting U1 snRNP contacts with tri-snRNP and coordinating Brr2 and Prp24 activities.\",\n      \"method\": \"Large-scale genetic suppressor screen, yeast two-hybrid, analysis of genetic interactions\",\n      \"journal\": \"Genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — large-scale suppressor screen plus yeast two-hybrid, single lab\",\n      \"pmids\": [\"10924465\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"Seven different missense mutations in PRPC8 (PRPF8) clustered within a 14-codon stretch at the C-terminus cause autosomal dominant retinitis pigmentosa (RP13), establishing that mutations in this ubiquitous splicing factor cause retinal-specific degeneration.\",\n      \"method\": \"Positional cloning, direct sequencing, cosegregation analysis in multiple families\",\n      \"journal\": \"Human molecular genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — cosegregation demonstrated across multiple independent families, multiple independent mutations identified\",\n      \"pmids\": [\"11468273\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"Deletion of SKY1 (SRPK-family kinase) is synthetically lethal with specific prp8 alleles in a domain implicated in 3'AG recognition fidelity, and sky1 deletion suppresses 3'AG mutations, suggesting that 3' splice site AG recognition by Prp8 is subject to phosphorylation regulation.\",\n      \"method\": \"Genetic synthetic lethality analysis, ACT1-CUP1 splicing reporter assay, yeast genetics\",\n      \"journal\": \"RNA (New York, N.Y.)\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — genetic interaction data, single lab, no direct biochemical evidence for phosphorylation of Prp8\",\n      \"pmids\": [\"11565750\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Distinct domains of Prp8 mediate different aspects of spliceosome activation: regions a, d, and e show allele-specific genetic interactions with Prp28, Brr2 (Prp44), and U6 RNA respectively, revealing that Prp8 coordinates multiple processes including U1 snRNP release and U4/U6 unwinding.\",\n      \"method\": \"Allele-specific genetic interaction analysis, yeast genetics\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — systematic allele-specific genetic epistasis across multiple factors, single lab\",\n      \"pmids\": [\"12087126\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"A missense mutation in PRP8 (R1753K) suppresses multiple helicase-deficient prp22 mutations, suggesting Prp8 stabilizes an RNA-protein or RNA-RNA interaction in the spliceosome that must be disrupted by Prp22's helicase activity for mRNA release.\",\n      \"method\": \"Extragenic suppressor screen, in vitro splicing assay, mRNA release assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — suppressor genetics combined with in vitro splicing assay, single lab\",\n      \"pmids\": [\"14688266\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Assembly of Snu114 into the U5 snRNP requires a functional GTPase domain and Prp8; GTPase domain mutants of Snu114 fail to interact with Prp8 or U5 snRNA and cannot assemble U5 snRNPs, whereas C-terminal truncation mutants assemble spliceosomes but block U4 snRNP release.\",\n      \"method\": \"snRNP and spliceosome assembly analysis in SNU114 mutant extracts, immunoprecipitation\",\n      \"journal\": \"RNA (New York, N.Y.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct assembly analysis with defined mutants, single lab\",\n      \"pmids\": [\"16540695\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Transposon-based dissection of Prp8 establishes that catalytic core RNAs (U5, U6 snRNAs, pre-mRNA) make multiple contacts in the central region of Prp8, identifying a discrete highly conserved region as a candidate splicing cofactor.\",\n      \"method\": \"Transposon-based insertion mutagenesis, RNA-protein crosslinking, functional analysis\",\n      \"journal\": \"RNA (New York, N.Y.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — systematic transposon insertion with crosslinking analysis, single lab\",\n      \"pmids\": [\"16431982\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"The C-terminal domain of yeast Prp8p adopts a Jab1/MPN-like fold (pseudoenzyme with impaired metal-binding site) that serves as a protein-protein interaction platform; RP13-linked mutations in the C-terminal appendix weaken interactions with Brr2 and Snu114.\",\n      \"method\": \"X-ray crystallography, targeted yeast two-hybrid analysis, mutagenesis\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure combined with functional mutagenesis and yeast two-hybrid, single study with multiple orthogonal methods\",\n      \"pmids\": [\"17317632\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"RP13 mutations in Prp8 in yeast cause nuclear accumulation of a precursor U5 snRNP that lacks Brr2, and Prp8 contains a nuclear localization signal required for efficient nuclear import of the precursor U5 snRNP; Brr2 joins U5 snRNP in the nucleus, and RP mutations disrupt the Prp8–Brr2 interaction required for this step.\",\n      \"method\": \"Yeast genetics, immunofluorescence, subcellular fractionation, co-immunoprecipitation\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (genetics, localization, fractionation, co-IP) establishing a biogenesis pathway, single study\",\n      \"pmids\": [\"17934474\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Opposing classes of prp8 alleles modulate the equilibrium between the first and second catalytic steps of splicing: one class suppresses first-step defects (analogous to ribosomal 'ram' mutants) and opposes previously described second-step suppressors; this transition is linked to U6 snRNA and Prp16 ATPase activity.\",\n      \"method\": \"Genetic characterization of prp8 alleles, genetic interaction epistasis with U6 and prp16 mutations, in vivo splicing assays\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — systematic genetic analysis with epistasis, single lab\",\n      \"pmids\": [\"17486100\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"The crystal structure of the beta-finger domain of Prp8 (residues 1822–2095) reveals a beta-hairpin finger protruding from the protein; mutations throughout the beta-finger alter the equilibrium between first and second catalytic steps, while mutations at its base affect U4/U6 unwinding-mediated spliceosome activation.\",\n      \"method\": \"X-ray crystallography, mutagenesis, in vivo functional analysis\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — crystal structure combined with systematic mutagenesis and functional assays, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"18779563\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"A fragment from the C-terminus of Prp8 activates Brr2-dependent U4/U6 snRNA dissociation in vitro; notably, fragments carrying RP-associated prp8 alleles do not stimulate U4/U6 unwinding activity. The same fragment also inhibits Brr2 U4/U6-dependent ATPase activity.\",\n      \"method\": \"In vitro U4/U6 unwinding assay, ATPase assay, recombinant protein fragments\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro reconstitution assay with defined protein fragments and RP mutant variants, multiple activities measured\",\n      \"pmids\": [\"19098916\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Electron microscopy of the yeast tri-snRNP localizes Brr2 to a head domain, while Prp8 and Snu114 occupy a central position; the head (containing Brr2) and arm (containing U4/U6 snRNP) adopt variable relative positions, suggesting conformational dynamics relevant to spliceosome activation.\",\n      \"method\": \"EM projection structure of genetically tagged tri-snRNP proteins\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — EM with genetic tagging for localization, single lab\",\n      \"pmids\": [\"18953335\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"The 1.85 Å crystal structure of PRP8 domain IV reveals a bipartite structure with an RNase H fold linked to a five-helix assembly; RNA binding studies and analysis of mutant yeast alleles in context of this structure show domain IV forms a surface interacting directly with RNA structures at the spliceosome catalytic core.\",\n      \"method\": \"X-ray crystallography (1.85 Å), RNA binding studies, mutant allele analysis\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — high-resolution crystal structure combined with RNA binding studies and mutant analysis, multiple orthogonal methods\",\n      \"pmids\": [\"18836455\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Bioinformatic and structural analysis reveals that Prp8's central conserved domain is related to the catalytic domain of reverse transcriptases (most similar to prokaryotic retroelement RTs), followed by a region analogous to maturase/X domains and a C-terminal RNaseH-like fold, suggesting Prp8 evolved by acquiring nucleic acid-binding domains from inactivated retroelements.\",\n      \"method\": \"Sequence analysis, structural homology analysis\",\n      \"journal\": \"RNA (New York, N.Y.)\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 4 / Moderate — computational/structural prediction without direct biochemical validation of proposed evolutionary origin\",\n      \"pmids\": [\"21441348\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"The RNase H (RH) domain of Prp8 binds U4/U6 snRNA through the single-stranded regions preceding U4/U6 stem I; Brr2 also recognizes this same single-stranded U4 region and translocates along U4 to unwind stem I first; the Prp8 RH domain inhibits U4/U6 unwinding by blocking Brr2 loading onto U4 snRNA.\",\n      \"method\": \"RNA binding assays, cross-linking coupled with mass spectrometry, in vitro U4/U6 unwinding inhibition assay\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro binding and unwinding assays combined with crosslinking-mass spectrometry mapping of contact residues, multiple orthogonal methods\",\n      \"pmids\": [\"23124066\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"The RNase H domain of PRP8 undergoes a conformational switch between the two steps of splicing, and this switch unmasks a metal-binding site involved in the second catalytic step (exon ligation); PRP8 is a metalloprotein that promotes exon ligation within the spliceosome.\",\n      \"method\": \"Biochemical assays, metal binding studies, analysis of prp8 alleles promoting first vs. second step\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — direct metal binding demonstration combined with conformational analysis and allele-based functional validation\",\n      \"pmids\": [\"23686287\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Crystal structure of yeast Prp8 (residues 885–2413) in complex with Aar2 reveals tightly associated domains resembling a bacterial group II intron reverse transcriptase and a type II restriction endonuclease; a large cavity formed by the reverse transcriptase thumb, endonuclease-like, and RNaseH-like domains accommodates splice-site suppressors and the intron branch-point crosslink, establishing this cavity as the active site of the spliceosome.\",\n      \"method\": \"X-ray crystallography of Prp8–Aar2 complex, mapping of suppressor mutations and crosslinks onto structure\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure at high resolution combined with mapping of functionally validated genetic suppressors, landmark structural study\",\n      \"pmids\": [\"23354046\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Structural and biochemical analyses show that the Prp8 Jab1/MPN domain binds exclusively to the N-terminal helicase cassette of Brr2 and stimulates Brr2 activity; RP-associated mutations in the Jab1/MPN domain map to the Brr2 interface; Aar2 and Brr2 are mutually exclusive binders of the Jab1/MPN domain, explaining the cytoplasm-to-nucleus switch during U5 snRNP maturation.\",\n      \"method\": \"X-ray crystallography of Brr2–Prp8 Jab1/MPN domain complex, mutagenesis, biochemical activity assays\",\n      \"journal\": \"Structure (London, England : 1993)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure combined with mutagenesis and functional biochemical assays, multiple orthogonal methods\",\n      \"pmids\": [\"23727230\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Prp8 inserts its C-terminal tail into Brr2's RNA-binding tunnel, thereby competitively blocking Brr2's RNA-binding, ATPase, and U4/U6 unwinding activities; RP-linked Prp8 mutations that map to this tail cause inefficient Brr2 repression as their primary recognizable phenotype.\",\n      \"method\": \"X-ray crystallography, biochemical helicase assays (ATPase, unwinding), mutagenesis\",\n      \"journal\": \"Science (New York, N.Y.)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure plus multiple in vitro biochemical activity assays with RP mutant variants, multiple orthogonal methods in single rigorous study\",\n      \"pmids\": [\"23704370\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"In vivo CLIP/CRAC analysis reveals that Prp8 contacts U1 and U2 snRNAs in addition to its known U5, U6, and pre-mRNA contacts; disruption of Prp8–U1 snRNA interaction reduces tri-snRNP levels in the spliceosome, revealing a role for Prp8 in spliceosomal assembly through U1 snRNA interaction.\",\n      \"method\": \"CLIP/CRAC with next-generation sequencing, in vivo crosslinking, functional analysis of U1 interaction mutants\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — comprehensive in vivo crosslinking combined with functional tri-snRNP assembly analysis, single lab\",\n      \"pmids\": [\"23393194\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Prp8 contacts with nucleotides surrounding the branch-site are enhanced during step 1 catalysis (in C complex), as revealed by UV-induced crosslinking of purified yeast B(act) and C spliceosomes on site-specifically labeled pre-mRNA.\",\n      \"method\": \"UV-induced crosslinking of purified spliceosomal complexes with site-specifically labeled pre-mRNA\",\n      \"journal\": \"PLoS genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — purified spliceosome crosslinking with defined labeling, single lab\",\n      \"pmids\": [\"26393790\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Picornaviral 3D polymerase (RdRp) enters the nucleus and associates with the C-terminal Jab1/MPN domain of Prp8, interfering with the second catalytic step of splicing and causing accumulation of lariat splicing intermediates; this disrupts pre-mRNA splicing of endogenous transcripts involved in cell growth and differentiation.\",\n      \"method\": \"Co-immunoprecipitation, domain mapping, in vitro splicing assay, nuclear fractionation, identification of endogenous trapped pre-mRNAs\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-immunoprecipitation with domain mapping and in vitro splicing assay, single lab\",\n      \"pmids\": [\"24968230\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"PRPF8 deficiency (knockdown or hemizygous deletion) in K562 and CD34+ bone marrow cells causes missplicing defects and increased proliferative capacity; yeast models with homologous PRPF8 mutations abrogate the second-step block in splicing, suggesting the mutations impair proofreading function.\",\n      \"method\": \"Knockdown experiments, whole-RNA deep sequencing, yeast complementation/mutagenesis\",\n      \"journal\": \"Leukemia\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function with transcriptomic readout combined with yeast functional validation, single lab\",\n      \"pmids\": [\"24781015\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"PRPF8 depletion preferentially impairs splicing of introns with weak 5' splice sites across the human transcriptome, leading to mitotic arrest; iCLIP shows PRPF8 depletion decreases RNP complex formation at most splice sites, and experimental enhancement of 5' splice site strength overcomes the effects of PRPF8 depletion.\",\n      \"method\": \"siRNA knockdown, RNA-seq, iCLIP, minigene splicing assays\",\n      \"journal\": \"Genome biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — iCLIP plus RNA-seq plus minigene functional rescue, multiple orthogonal methods establishing mechanism\",\n      \"pmids\": [\"26392272\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Stable tri-snRNP integration into the exon-defined complex requires interaction between Prp8 and nucleotides at the exon–intron junction (5' splice site), and this is the key trigger for the major structural rearrangement from 37S to 45S B-like complex, independent of B-specific proteins or hPrp31 phosphorylation.\",\n      \"method\": \"Electron microscopy, affinity purification, addition of 5'ss RNA oligonucleotides to cross-exon complexes\",\n      \"journal\": \"RNA (New York, N.Y.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — EM structural analysis combined with functional reconstitution using defined RNA oligonucleotides, single lab\",\n      \"pmids\": [\"26385511\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Prp8 interacts physically with the androgen receptor (AR) via the AR nuclear export signal (NES); co-immunoprecipitation and deletion mutagenesis demonstrate Prp8-AR interaction, and Prp8 knockdown induces nuclear accumulation of AR and increases its polyubiquitination, modulating AR transcriptional activity.\",\n      \"method\": \"Co-immunoprecipitation, deletion mutagenesis, shRNA knockdown, rapamycin export assay, luciferase reporter assay\",\n      \"journal\": \"Molecular endocrinology (Baltimore, Md.)\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single co-IP with knockdown, single lab, unclear mechanistic specificity for splicing vs. direct AR regulation\",\n      \"pmids\": [\"26371515\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"A subset of RP-linked Prp8 mutations (mapping to the hinge linking the Jab1-MPN tail to the globular domain) cause defects in the transition between the first and second catalytic steps of splicing, in addition to spliceosome activation defects; genetic analyses link Snu114 GTP/GDP occupancy to Prp8-dependent regulation of Brr2.\",\n      \"method\": \"Yeast genetics, in vivo and in vitro splicing reporter assays, genetic interaction analysis with SNU114 mutants\",\n      \"journal\": \"RNA (New York, N.Y.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — combination of in vitro and in vivo splicing assays with genetic epistasis, single lab\",\n      \"pmids\": [\"26968627\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"The RNaseH domain of Prp8 contains a 17-aa extension (Switch loop) that can adopt two mutually exclusive structures; prp8 alleles mapping to this extension fall into two functional classes—those promoting error-prone/efficient splicing versus hyperaccurate/inefficient splicing—and error-prone alleles suppress a prp2 mutant deficient at promoting the first catalytic step.\",\n      \"method\": \"Systematic mutagenesis, in vitro and in vivo reporter assays, lariat sequencing for genome-wide splice site analysis, genetic epistasis\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — multiple orthogonal approaches including structural rationale, in vitro assays, in vivo reporters, and genome-wide lariat sequencing\",\n      \"pmids\": [\"28416677\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"PRPF8 depletion causes a specific defect in homology-directed repair (HDR) and single-strand annealing (SSA), which require BRCA1; PRPF8 depletion reduces end resection (measured as chromatin-bound RPA), BRCA1 foci, and histone acetylation marks associated with BRCA1-mediated HR.\",\n      \"method\": \"siRNA knockdown, DR-GFP/SA-GFP HR reporter assays, immunofluorescence (BRCA1, RPA, histone acetylation foci)\",\n      \"journal\": \"Oncotarget\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function with multiple functional readouts for HR, single lab\",\n      \"pmids\": [\"29212152\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Mutagenesis of Prp8 residues identified by amino acid probing to be positioned near U5 snRNA reveals their role in 5' splice site recognition; genetic interactions with Isy1 and Snu114 further support that Prp8–U5 snRNA interactions help position the pre-mRNA into the active site.\",\n      \"method\": \"Amino acid probing strategy (chemical crosslinking), site-directed mutagenesis, genetic interaction analysis\",\n      \"journal\": \"RNA (New York, N.Y.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — novel crosslinking approach combined with mutagenesis and genetic interactions, single lab\",\n      \"pmids\": [\"29487104\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"PRPF8 knockdown impairs hypoxia-induced mitophagy by causing aberrant mRNA splicing of ULK1, which initiates autophagy; the RP-associated PRPF8 mutant R2310K is defective in regulating mitophagy.\",\n      \"method\": \"RNAi screen with mt-Keima fluorescent reporter, PRPF8 knockdown, RT-PCR of ULK1 splicing, mitophagy assays\",\n      \"journal\": \"Autophagy\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — RNAi screen with defined reporter, splicing verification, and functional mitophagy assay, single lab\",\n      \"pmids\": [\"30103670\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Two prp-8 alleles in C. elegans identified by genetic screen alter cryptic splice site usage frequency; complementary genetic and structural analyses in yeast implicate these alleles in stability of the spliceosome catalytic core; high-throughput mRNA sequencing shows overall alternative splicing patterns are relatively unchanged despite effects on cryptic splicing.\",\n      \"method\": \"C. elegans genetic screen, RNA-seq (high-throughput mRNA sequencing), yeast structural/genetic analysis\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic screen combined with genome-wide RNA-seq and cross-species analysis, single lab\",\n      \"pmids\": [\"30674666\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"The evolutionarily conserved protein Ecdysoneless (Ecd) chaperones Prp8 delivery to the forming U5 snRNP in the cytoplasm; Ecd deficiency leads to reduced Prp8 protein levels and compromised U5 snRNP biogenesis, causing loss of splicing fidelity; SmD3 was identified as a novel interaction partner of Ecd.\",\n      \"method\": \"Drosophila genetics, proteomic approaches (co-immunoprecipitation/mass spectrometry), Western blot, splicing analysis\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetics combined with proteomics and functional splicing assays, single lab\",\n      \"pmids\": [\"33444449\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"PRPF8 silencing in hepatocellular carcinoma cells modulates fibronectin (FN1) splicing by promoting exclusion of exon 40.2, which reduces FAK/AKT phosphorylation and blunts stress fiber formation, thereby decreasing invasive capacity; CLIPseq analysis shows PRPF8 binds preferentially to exons of protein-coding genes.\",\n      \"method\": \"siRNA knockdown, RNA-seq, CLIPseq, Western blot (FAK/AKT phosphorylation), invasion assay, xenograft tumor growth\",\n      \"journal\": \"Experimental & molecular medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function combined with CLIPseq and defined signaling pathway readout, single lab\",\n      \"pmids\": [\"36609600\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"The PRPF8/Brr2 regulatory axis controls 5' splice site (5'SS) selection; the heterozygous PRPF8 c.6926 A>C (p.H2309P) RP mutation impairs alternative splicing and weak/suboptimal 5'SS selection, enhances cryptic splicing in ciliary/retinal-specific transcripts, alters PRPF8 interaction with U6 snRNA, and causes accumulation of active spliceosomes and poly(A)+ mRNAs at nuclear periphery splicing clusters in photoreceptors.\",\n      \"method\": \"Patient iPSC-derived retinal cells, transcriptomics, proteomics, co-immunoprecipitation (PRPF8-U6 snRNA), immunofluorescence of nuclear speckles/poly(A)+ mRNA\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — comprehensive multi-omics in patient-derived disease-relevant cells combined with molecular and cell biological validation using multiple orthogonal methods\",\n      \"pmids\": [\"38605034\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"PRPF8 (hPrp8/Prp8) is the central scaffolding protein of the spliceosome catalytic core, occupying a large cavity formed by its reverse-transcriptase-like, RNaseH, and Jab1/MPN domains that directly contacts both splice sites, the branch point, and U2, U5, and U6 snRNAs throughout both catalytic steps; it regulates spliceosome activation by controlling the timing of U4/U6 unwinding through both stimulatory and inhibitory interactions with the Brr2 helicase (inserting its C-terminal tail into Brr2's RNA-binding tunnel to reversibly block its activity), coordinates the activities of Prp28 and Snu114 GTPase, undergoes a conformational switch in its RNaseH domain between the two catalytic steps that unmasks a metal-binding site required for exon ligation, and its C-terminal Jab1/MPN domain acts as a protein-protein interaction platform whose disruption by RP13-associated mutations weakens Brr2/Snu114 binding, impairs U5 snRNP biogenesis and nuclear import, and causes defects in 5' splice site selection and the first-to-second step transition underlying retinal degeneration.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"PRPF8 (hPrp8/Prp8) is the central scaffolding protein of the spliceosome catalytic core, an essential, RNA-free component of the U5 snRNP and U4/U5/U6 tri-snRNP that remains stably associated with the spliceosome throughout both transesterification steps and into post-splicing intron complexes [#0, #1, #9]. It is a core factor of both the major U2-dependent and minor U12-dependent spliceosomes [#14], and makes direct, largely non-sequence-specific contacts with the pre-mRNA at the 5' splice site, branch point, and 3' splice site as well as with U5 and U6 snRNAs, positioning the reactive exons and intron termini within the active site [#3, #6, #11, #36]. Crystallographic and biochemical work showed that its central region is built from reverse-transcriptase-like, endonuclease-like, and RNaseH-like domains that together form a large cavity accommodating the splice sites and branch-point crosslink — the catalytic cavity of the spliceosome [#28, #32] — while its C-terminal Jab1/MPN domain is a catalytically dead pseudoenzyme fold serving as a protein-protein interaction platform [#22]. PRPF8 governs the timing of spliceosome activation by regulating the Brr2 helicase that unwinds U4/U6: its RNaseH domain binds the single-stranded U4 region and blocks Brr2 loading, its C-terminal tail inserts into Brr2's RNA-binding tunnel to competitively repress its ATPase and unwinding activities, and its Jab1/MPN domain binds the Brr2 helicase cassette and stimulates activity, coupling 5' splice site recognition to U4/U6 unwinding [#13, #26, #30, #33, #34]. A conformational switch in its RNaseH domain between the two catalytic steps unmasks a metal-binding site required for exon ligation, establishing PRPF8 as a metalloprotein that controls the first-to-second step transition [#31, #43]. PRPF8 also coordinates U5 snRNP biogenesis: it carries a nuclear localization signal required for import of a Brr2-free precursor U5 snRNP, with Aar2 and Brr2 binding the Jab1/MPN domain mutually exclusively to drive the cytoplasm-to-nucleus maturation switch [#23, #33], and its delivery to the nascent U5 snRNP is chaperoned by Ecdysoneless [#48]. Transcriptome-wide, PRPF8 is required for efficient splicing of introns with weak 5' splice sites and for fidelity of 5' splice site selection [#39, #50]. Heterozygous missense mutations clustered at the C-terminus cause autosomal dominant retinitis pigmentosa (RP13) by weakening Brr2/Snu114 binding, impairing U5 snRNP biogenesis and Brr2 repression, and disrupting 5' splice site selection and the first-to-second step transition in photoreceptors [#16, #22, #26, #34, #50].\",\n  \"teleology\": [\n    {\n      \"year\": 1990,\n      \"claim\": \"Establishing that the conserved ~220 kDa Prp8 protein is a stable spliceosome component that directly touches pre-mRNA defined it as a core catalytic-machinery factor rather than a transient assembly factor.\",\n      \"evidence\": \"Gradient fractionation, immunoprecipitation, and UV-crosslinking with anti-PRP8 antisera in HeLa extracts; stable association across both splicing steps\",\n      \"pmids\": [\"2479028\", \"2138328\", \"2139226\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Crosslink sites on pre-mRNA not mapped at nucleotide resolution\", \"No structural basis for RNA contact\"]\n    },\n    {\n      \"year\": 1991,\n      \"claim\": \"Demonstrating ATP-dependent, splicing-competence-restricted crosslinking of Prp8 to pre-mRNA showed its contacts are functionally coupled to the catalytic reaction, not nonspecific binding.\",\n      \"evidence\": \"UV-crosslinking with 5'SS and branchpoint mutant substrates in yeast in vitro splicing\",\n      \"pmids\": [\"1945827\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Which Prp8 domain contacts RNA was unknown\", \"Did not distinguish direct catalytic role from scaffolding\"]\n    },\n    {\n      \"year\": 1992,\n      \"claim\": \"Showing Prp8 is required for stable tri-snRNP formation and snRNA stability established its essential role in spliceosome assembly upstream of catalysis.\",\n      \"evidence\": \"Genetic depletion, ts inactivation, and antibody inhibition with sedimentation analysis in yeast\",\n      \"pmids\": [\"1396567\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism of tri-snRNP stabilization unresolved\", \"snRNA decline could be indirect\"]\n    },\n    {\n      \"year\": 1995,\n      \"claim\": \"Mapping Prp8 contacts to exon nucleotides at both splice sites established it as an aligner that positions exons against U5 snRNA at the active site through both catalytic steps.\",\n      \"evidence\": \"Site-specific 4-thiouridine UV-crosslinking with mutant and duplicated splice sites\",\n      \"pmids\": [\"7781612\", \"7885825\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No structural model of the alignment\", \"Branchpoint-3'SS interaction timing only inferred\"]\n    },\n    {\n      \"year\": 1999,\n      \"claim\": \"Genetic suppressor and crosslinking studies placed Prp8 at the heart of splice-site tertiary interactions and identified it as a controller of the timing of U4/U6 unwinding-driven activation.\",\n      \"evidence\": \"Allele-specific suppression of 5'GU, 3'YAG, and U6 ACAGAG mutations; prp8-201 suppression of U4-cs1 unwinding block; mapping of the 5'SS crosslink to hPrp8 C-terminus\",\n      \"pmids\": [\"10444595\", \"10444596\", \"10024880\", \"10024169\", \"10411133\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular mechanism linking Prp8 to unwinding timing not yet biochemical\", \"Allosteric model untested structurally\"]\n    },\n    {\n      \"year\": 1998,\n      \"claim\": \"Reconstituting an RNA-free Prp8/Snu114/Brr2/U5-40K core complex defined Prp8 as the central hub binding its key protein partners directly.\",\n      \"evidence\": \"Chaotropic dissociation of U5 snRNP with sedimentation and binding analysis\",\n      \"pmids\": [\"9774689\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Binding interfaces not mapped\", \"Stoichiometry and direct vs. bridged contacts uncertain\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Allele-specific genetic dissection assigned distinct Prp8 regions to interactions with Prp28, Brr2, U6 RNA and U1 release, showing Prp8 coordinates multiple activation steps from a single scaffold.\",\n      \"evidence\": \"Large-scale suppressor screens, yeast two-hybrid, and allele-specific epistasis with multiple splicing factors\",\n      \"pmids\": [\"10924465\", \"12087126\", \"16431982\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Genetic interactions did not establish direct physical contacts\", \"No structural assignment of the mapped regions\"]\n    },\n    {\n      \"year\": 2001,\n      \"claim\": \"Identification of clustered C-terminal missense mutations causing autosomal dominant retinitis pigmentosa connected this ubiquitous splicing factor to a tissue-specific degenerative disease.\",\n      \"evidence\": \"Positional cloning and cosegregation across multiple RP13 families\",\n      \"pmids\": [\"11468273\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism of retinal specificity unexplained\", \"Functional consequence of mutations not yet defined\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Crystal structures of the RNaseH-fold domain IV, the beta-finger, and the Jab1/MPN domain provided the physical framework explaining how Prp8 contacts catalytic-core RNAs and partners and how RP mutations act.\",\n      \"evidence\": \"X-ray crystallography of domain IV, beta-finger, and the Jab1/MPN-like C-terminal domain combined with RNA-binding, mutagenesis, and yeast two-hybrid\",\n      \"pmids\": [\"18836455\", \"18779563\", \"17317632\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Full-length architecture still missing\", \"Catalytic role vs. scaffolding role of metal site not yet shown\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Linking opposing prp8 allele classes and the beta-finger to the first-vs-second step equilibrium established Prp8 as an active regulator of the catalytic step transition.\",\n      \"evidence\": \"Genetic epistasis of prp8 alleles with U6 and Prp16 ATPase mutations plus beta-finger mutagenesis\",\n      \"pmids\": [\"17486100\", \"18779563\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural basis of the conformational switch not yet resolved\", \"How Prp8 senses step transition unclear\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"In vitro reconstitution showed Prp8 C-terminal fragments both stimulate and inhibit Brr2-driven U4/U6 unwinding, and that RP mutants fail to stimulate — directly tying disease alleles to Brr2 regulation.\",\n      \"evidence\": \"In vitro U4/U6 unwinding and ATPase assays with recombinant Prp8 fragments and RP variants; EM of tri-snRNP\",\n      \"pmids\": [\"19098916\", \"18953335\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Atomic basis of stimulation vs. inhibition not yet structural\", \"In vivo balance of the two activities unquantified\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Landmark crystal structures of Prp8 in complex with Aar2 and Brr2 revealed the RT/endonuclease/RNaseH cavity as the spliceosome active site and defined the Jab1/MPN–Brr2 interface, mechanistically explaining U5 maturation and Brr2 control.\",\n      \"evidence\": \"X-ray crystallography of Prp8–Aar2 and Brr2–Jab1/MPN complexes with mapping of suppressors and RP mutations; C-terminal tail insertion into the Brr2 RNA tunnel\",\n      \"pmids\": [\"23354046\", \"23727230\", \"23704370\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Conformational states during catalysis inferred, not captured\", \"Dynamics of tail insertion/release in the assembled spliceosome unresolved\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Demonstrating that the RNaseH domain switches conformation to unmask a metal-binding site, and binds single-stranded U4 to block Brr2 loading, defined Prp8 as a metalloprotein that gates both exon ligation and activation timing.\",\n      \"evidence\": \"Metal-binding and conformational assays with step-specific alleles; RNA binding plus crosslinking-MS and in vitro unwinding inhibition\",\n      \"pmids\": [\"23686287\", \"23124066\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Identity of catalytic vs. structural metal not fully defined\", \"Whether the metal directly participates in chemistry untested\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"In vivo CLIP/CRAC extended Prp8 RNA contacts to U1 and U2 snRNAs and linked the U1 interaction to tri-snRNP recruitment, broadening its role to early assembly.\",\n      \"evidence\": \"CLIP/CRAC sequencing with functional analysis of U1-interaction mutants in yeast\",\n      \"pmids\": [\"23393194\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct vs. indirect U1/U2 contacts not separated\", \"Structural basis of U1 contact unknown\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Transcriptome-wide depletion studies established that Prp8 contacts at the exon-intron junction trigger tri-snRNP integration and that it is preferentially required for introns with weak 5' splice sites.\",\n      \"evidence\": \"EM with 5'SS oligonucleotide reconstitution; siRNA knockdown with RNA-seq, iCLIP, and minigene rescue by 5'SS strengthening\",\n      \"pmids\": [\"26385511\", \"26392272\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Quantitative rules for 5'SS strength dependence incomplete\", \"Tissue-specific splicing targets not yet defined\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Crosslinking-guided mutagenesis and structural analysis of the RNaseH Switch loop refined how Prp8–U5 snRNA contacts position pre-mRNA and toggle splicing fidelity.\",\n      \"evidence\": \"Amino acid probing, Switch-loop structural analysis, in vitro/in vivo reporters and lariat sequencing in yeast\",\n      \"pmids\": [\"29487104\", \"28416677\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How fidelity classes map to human disease alleles incompletely resolved\", \"Dynamics of the two Switch-loop states in vivo unmeasured\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Identifying Ecdysoneless as a Prp8 chaperone that delivers it to the forming U5 snRNP completed an assembly pathway whose disruption reduces Prp8 levels and splicing fidelity.\",\n      \"evidence\": \"Drosophila genetics, co-IP/MS proteomics, and splicing analysis\",\n      \"pmids\": [\"33444449\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism of chaperone handoff unresolved\", \"Human relevance of Ecd–Prp8 axis not tested directly\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Patient iPSC-derived retinal cells linked the PRPF8/Brr2 axis and a specific RP mutation to defective weak/cryptic 5'SS selection, altered U6 snRNA interaction, and spliceosome/mRNA accumulation, providing a disease mechanism for RP13.\",\n      \"evidence\": \"Patient iPSC-derived retinal cells with transcriptomics, proteomics, PRPF8–U6 snRNA co-IP, and nuclear speckle/poly(A)+ imaging\",\n      \"pmids\": [\"38605034\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Why photoreceptors are uniquely vulnerable not fully explained\", \"Causal chain from missplicing to degeneration incomplete\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How PRPF8's well-characterized core splicing functions connect to its reported roles in homology-directed DNA repair, androgen receptor regulation, mitophagy via ULK1 splicing, and cancer-associated FN1 splicing remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether these phenotypes are direct or downstream of global splicing changes is untested\", \"No unifying mechanism links the canonical spliceosome role to these specialized outputs\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0003723\", \"supporting_discovery_ids\": [3, 6, 28, 30, 35, 50]},\n      {\"term_id\": \"GO:0140098\", \"supporting_discovery_ids\": [31, 43]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [26, 30, 33, 34]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [6, 9, 22]},\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [5, 32]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [0, 23]},\n      {\"term_id\": \"GO:0005654\", \"supporting_discovery_ids\": [50]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [0, 5, 39, 50]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [6, 11, 39]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [16, 50]}\n    ],\n    \"complexes\": [\n      \"U5 snRNP\",\n      \"U4/U6.U5 tri-snRNP\",\n      \"spliceosome catalytic core\",\n      \"U12-dependent (minor) spliceosome\"\n    ],\n    \"partners\": [\n      \"BRR2\",\n      \"SNU114\",\n      \"AAR2\",\n      \"PRP28\",\n      \"ECD\",\n      \"U5-40K\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":8,"faith_total":8,"faith_pct":100.0}}