Affinage

U2AF2

Splicing factor U2AF 65 kDa subunit · UniProt P26368

Length
475 aa
Mass
53.5 kDa
Annotated
2026-04-28
100 papers in source corpus 43 papers cited in narrative 43 extracted findings

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

U2AF2 (U2AF65) is an essential pre-mRNA splicing factor that recognizes polypyrimidine tracts at 3′ splice sites to nucleate early spliceosome assembly. Its tandem RRM1–RRM2 domains bind pyrimidine-rich RNA through a conformational selection mechanism regulated by an autoinhibitory intrinsically disordered inter-RRM linker that proofreads against weak Py tracts, while the RS domain directly contacts the branch point sequence to promote U2 snRNA–branch point base pairing (PMID:8781232, PMID:32188783, PMID:24734879). U2AF2 cooperatively assembles with SF1 (via its C-terminal UHM domain) and U2AF1 at 3′ splice sites, subsequently exchanging SF1 for SF3B1 during U2 snRNP recruitment; it also recruits UAP56 and the Prp19 complex, couples splicing to transcription by binding the phosphorylated RNA Pol II CTD, and associates with chromatin at gene bodies in an H3K36me3-dependent manner to enhance exon selection accuracy (PMID:9512519, PMID:35780835, PMID:9242493, PMID:21536736, PMID:40315850). U2AF2 binding is competitively modulated by hnRNP C, PTB, MBNL1, and HNRNPA1 to regulate alternative splicing outcomes, and its activity is post-translationally tuned by JMJD6-mediated lysine hydroxylation, SIRT4-mediated deacetylation at K413, and ubiquitin-dependent turnover counteracted by OTUB2 (PMID:23374342, PMID:16982681, PMID:19574390, PMID:39495216, PMID:30662561).

Mechanistic history

Synthesis pass · year-by-year structured walk · 19 steps
  1. 1992 High

    Establishing that U2AF65 is recruited to the 3′ splice site through cross-exon communication with U1 snRNP addressed how distant splice sites are paired during early spliceosome recognition.

    Evidence UV cross-linking and copurification showing U1 snRNP at the 5′ splice site enhances U2AF65 binding to the 3′ splice site

    PMID:1285125

    Open questions at the time
    • Molecular contacts mediating U1 snRNP–U2AF65 cross-exon communication not resolved
    • Whether bridging is SR-protein-dependent was not addressed
  2. 1995 High

    Demonstrating that U2AF65 directly contacts the branch point at the earliest (E complex) stage of spliceosome assembly established its role as the initial branch-point-positioning factor before ATP-dependent steps.

    Evidence Site-specific photochemical cross-linking at the branch point nucleotide in E complex

    PMID:7493318

    Open questions at the time
    • How U2AF65 is subsequently displaced from the branch point during A complex formation was unknown
  3. 1996 High

    Showing that the U2AF65 RS domain directly contacts the branch point and is required for U2 snRNA–branch point base pairing revealed a non-RNA-binding domain performing an essential RNA-contact function in splicing catalysis.

    Evidence UV cross-linking with RS domain mutants and in vitro splicing reconstitution

    PMID:8781232

    Open questions at the time
    • Structural basis of RS domain–branch point interaction not determined
    • Whether RS domain phosphorylation modulates this contact was not tested
  4. 1997 High

    Identifying UAP56 as a U2AF65-recruited DEAD-box ATPase required for stable U2 snRNP–branch point interaction established the enzymatic link between U2AF65 binding and U2 snRNP commitment, while localization studies showed the RS domain targets U2AF65 to nuclear speckles and active splicing sites.

    Evidence Affinity purification and functional complementation in splicing extracts (UAP56); immunofluorescence with domain-deletion mutants (localization)

    PMID:9166400 PMID:9242493

    Open questions at the time
    • The U2AF65 surface that contacts UAP56 was not structurally defined
    • Whether speckle localization is functionally required for splicing was not tested
  5. 1998 High

    Demonstrating cooperative SF1–U2AF65 binding to the branch point–Py tract RNA, mediated by the U2AF65 UHM domain, established the molecular basis for combinatorial 3′ splice site recognition.

    Evidence In vitro cooperative RNA binding assays with U2AF65 deletion mutants

    PMID:9512519

    Open questions at the time
    • The structural basis of SF1–UHM recognition was not yet resolved
    • How the SF1–U2AF65 complex is disrupted during spliceosome progression was unknown
  6. 1999 High

    Reconstitution experiments resolved a longstanding question about U2AF subunit requirements by showing U2AF65 alone suffices for constitutive splicing but U2AF35 is additionally needed for weak Py tracts and enhancer-dependent substrates.

    Evidence U2AF depletion/reconstitution with recombinant subunits and UV cross-linking

    PMID:10567551 PMID:11421359

    Open questions at the time
    • Mechanism by which U2AF35 contributes to enhancer-dependent splicing was unclear
  7. 2001 High

    The crystal structure of the U2AF heterodimer interface revealed a novel tryptophan-mediated tongue-in-groove interaction essential for dimerization and RNA binding, providing the first atomic view of U2AF assembly.

    Evidence 2.2 Å X-ray crystallography with mutagenesis validation

    PMID:11551507

    Open questions at the time
    • Structure did not include the RNA-bound state
    • U2AF35 RRM was disordered in the free state — folding upon U2AF65 binding was shown separately by NMR (PMID:12297299)
  8. 2003 High

    The NMR structure of the U2AF65 UHM–SF1 complex established the UHM–ULM recognition paradigm — a conserved tryptophan insertion into a hydrophobic RRM pocket — that governs multiple splicing factor interactions.

    Evidence NMR structure determination of UHM–ULM complex

    PMID:12718882

    Open questions at the time
    • Whether UHM–ULM interactions are regulated by post-translational modifications was unknown
  9. 2005 High

    Thermodynamic analysis of SF3B1 multi-ULM binding to U2AF65 UHM, showing weaker affinity than SF1, explained why ATP hydrolysis is needed to drive the SF1-to-SF3B1 exchange during spliceosome progression.

    Evidence Quantitative fluorescence and CD binding measurements with purified proteins

    PMID:16376933

    Open questions at the time
    • The ATPase that drives the exchange was not identified
    • Structural basis of SF3B1 ULM trajectory on U2AF65 UHM was resolved only later (PMID:35780835)
  10. 2006 High

    Crystal structures of U2AF65 RRM1–RRM2 bound to polyuridine RNA revealed the hydrogen-bonding network for pyrimidine recognition, while PTB was shown to directly displace U2AF65 from Py tracts to repress alternative exons, establishing competitive displacement as a splicing regulatory mechanism.

    Evidence 2.5 Å X-ray crystallography with SPR and mutagenesis (RNA recognition); UV cross-linking and RNAi (PTB competition)

    PMID:16818232 PMID:16982681

    Open questions at the time
    • How U2AF65 accommodates non-uridine nucleotides at central positions was unresolved
    • Whether PTB competition is a general or exon-specific mechanism was not systematically addressed
  11. 2009 High

    JMJD6-catalyzed lysine hydroxylation of U2AF65 was identified as a novel post-translational modification that modulates alternative splicing, and MBNL1 was shown to compete with U2AF65 via RNA secondary structure, expanding the repertoire of U2AF65 regulatory mechanisms.

    Evidence Mass spectrometry, in vitro hydroxylation assay, reporter splicing assays (JMJD6); in vitro binding competition with stem-loop RNA (MBNL1)

    PMID:19470458 PMID:19574390

    Open questions at the time
    • Which specific lysine residues are hydroxylated and their individual functional consequences were not fully mapped
    • Genome-wide scope of MBNL1–U2AF65 competition was unknown
  12. 2011 High

    Demonstrating that U2AF65 binds the phosphorylated RNA Pol II CTD to recruit itself and the Prp19 complex established a direct molecular link between transcription elongation and co-transcriptional splicing activation.

    Evidence Biochemical purification, phospho-CTD binding assays, splicing complementation

    PMID:21536736

    Open questions at the time
    • Which CTD phosphorylation marks (Ser2 vs Ser5) are recognized was not fully resolved
    • Whether CTD binding and Py-tract binding occur simultaneously was unknown
  13. 2013 High

    Genome-wide iCLIP showed hnRNP C competes with U2AF65 at Alu-derived cryptic splice sites, establishing that widespread competition safeguards the transcriptome from aberrant exonization; additionally, U2AF65 was shown to inhibit splicing at weak alternative exons, revealing a dual activator/repressor role.

    Evidence Quantitative iCLIP, minigene assays, hnRNP C knockdown (competition); overexpression and cross-linking with SMN/Fas substrates (inhibitory role)

    PMID:23374342 PMID:26216990

    Open questions at the time
    • Structural basis of hnRNP C–U2AF65 competition was not resolved
    • The inhibitory mechanism at alternative exons lacked structural explanation
  14. 2014 High

    NMR/SAXS analysis of free multidomain U2AF65 revealed a dynamic conformational ensemble dominated by transient interdomain contacts, with RNA selecting a minor preformed open conformation — establishing conformational selection as the RNA recognition mechanism.

    Evidence NMR paramagnetic relaxation enhancement and SAXS of full-length U2AF65

    PMID:24734879

    Open questions at the time
    • How the conformational ensemble is modulated by U2AF1 binding was unknown
    • Role of post-translational modifications in shifting the ensemble was not addressed
  15. 2018 High

    Systematic comparison of in vitro and in vivo U2AF2 binding by iCLIP revealed that trans-acting RBPs extensively reshape U2AF2 occupancy in cells, enhancing its recruitment to authentic 3′ splice sites and clearing it from intron bodies; separately, HNRNPA1 was shown to redirect U2AF2 to decoy sites.

    Evidence In vitro iCLIP, mathematical modeling, machine learning, minigene validation; HNRNPA1 overexpression with iCLIP

    PMID:29643205 PMID:29650551

    Open questions at the time
    • Identity of the full complement of RBPs that enhance or repress U2AF2 binding was incomplete
    • Whether HNRNPA1-mediated redirection occurs in normal physiology or only upon overexpression was not clear
  16. 2020 High

    The intrinsically disordered inter-RRM linker was shown to autoinhibit U2AF2 binding to weak Py tracts, acting as a fidelity checkpoint, while single-molecule FRET demonstrated that U2AF1 modulates the open–closed RRM conformational equilibrium and MDS-associated U2AF1 mutations perturb this balance.

    Evidence NMR + iCLIP + linker mutagenesis + splicing assays (autoinhibition); single-molecule FRET with structure-guided mutagenesis (conformational dynamics)

    PMID:32188783 PMID:32343311

    Open questions at the time
    • Whether linker autoinhibition is modulated by post-translational modifications was not tested
    • How MDS-associated U2AF1 mutations alter splice site selection genome-wide through U2AF2 conformational changes was not fully resolved
  17. 2020 High

    Crystal structures of cancer-associated U2AF2 mutants (N196K and G301D) showed opposing structural effects on inter-RRM conformation and RNA affinity, providing a structural rationale for how somatic mutations alter splicing in cancer.

    Evidence X-ray crystallography, RNA binding affinity assays, minigene splicing assays

    PMID:33020180

    Open questions at the time
    • Genome-wide splicing consequences of these mutations in patient cells were not characterized
    • Whether these mutations alter interactions with U2AF1 or SF1 was not tested
  18. 2022 High

    Structural studies resolved how U2AF2 accommodates non-uridine nucleotides at the central Py-tract position and how SF3B1 ULM binds the U2AF2 UHM with a distinctive trajectory, with genome-wide analysis showing U2AF2–SF3B1 and U2AF2–SF1 co-regulate distinct splice site subsets.

    Evidence Multiple crystal structures with mutagenesis, eCLIP validation (nucleotide accommodation); 1.8 Å crystal structure, ITC, Co-IP, genome-wide splicing analysis (SF3B1 ULM)

    PMID:35524551 PMID:35780835

    Open questions at the time
    • Whether SF3B1 cancer mutations alter ULM–UHM binding to U2AF2 was not tested
    • Structural basis of the full ternary SF1–U2AF2–RNA vs SF3B1–U2AF2–RNA complexes remains unresolved
  19. 2025 High

    CUT&RUN profiling revealed that U2AF2 associates with chromatin throughout gene bodies in an H3K36me3-dependent, nascent-transcript-independent manner, establishing a chromatin-level mechanism for co-transcriptional exon selection fidelity.

    Evidence CUT&RUN with RNase A treatment, H3K36me3 dependency analysis, U2AF2 knockdown + RNA-seq

    PMID:40315850

    Open questions at the time
    • The reader that bridges H3K36me3 and U2AF2 chromatin recruitment is unknown
    • Whether chromatin-bound U2AF2 directly contacts nascent RNA or acts through an intermediate is not resolved
    • Generality across cell types not established

Open questions

Synthesis pass · forward-looking unresolved questions
  • Key unresolved questions include the structural basis of the full early spliceosomal complex containing U2AF2–SF1–U2AF1–RNA on chromatin, the identity of the H3K36me3 reader that recruits U2AF2 to chromatin, the interplay between multiple post-translational modifications (hydroxylation, acetylation, ubiquitination) in tuning U2AF2 splice site selectivity, and how disease-associated mutations in U2AF2 and its partners alter splicing networks in patient tissues.
  • No integrative structural model of the early 3′ splice site recognition complex on chromatin
  • Crosstalk among hydroxylation, acetylation, and ubiquitination of U2AF2 not studied
  • Genome-wide consequences of U2AF2 cancer mutations in patient-derived cells are lacking

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0003723 RNA binding 5 GO:0060090 molecular adaptor activity 3
Localization
GO:0005634 nucleus 2 GO:0005654 nucleoplasm 1 GO:0005694 chromosome 1
Pathway
R-HSA-8953854 Metabolism of RNA 12 R-HSA-74160 Gene expression (Transcription) 2
Partners
HNRNPCJMJD6OTUB2RBM39SF1SF3B1U2AF1UAP56
Complex memberships
SF1–U2AF2–U2AF1 early spliceosomal complexU2AF heterodimer (U2AF2–U2AF1)

Evidence

Reading pass · 43 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
1996 U2AF65 binds the polypyrimidine tract via its RNA binding domain, and its RS domain directly contacts the branch point to promote U2 snRNA–branch point base pairing, even in the absence of other splicing factors. RS domain mutants that lose branch point contact also lose the ability to support splicing. UV cross-linking, RS domain mutagenesis, in vitro splicing assays Science High 8781232
1997 U2AF65 recruits the DEAD-box ATPase UAP56 to the pre-mRNA branchpoint region, and UAP56 is required for stable U2 snRNP–branchpoint interaction; a novel region of U2AF65 (distinct from the RNA-binding and RS domains) mediates UAP56 recruitment. Affinity purification, functional complementation in splicing extracts, co-immunoprecipitation Genes & Development High 9242493
1992 U2AF65 binding to the 3' splice site is facilitated by U1 snRNP bound to an upstream 5' splice site, establishing an exon-bridging model in which U1 snRNP and U2AF65 communicate across the exon to promote splice site selection. UV cross-linking, copurification, antibody-mediated identification of p61 as U2AF65 Genes & Development High 1285125
1998 mBBP/SF1 binds branchpoint sequences and interacts with U2AF65 via the third RBD (UHM domain) of U2AF65; this interaction promotes cooperative binding to a BPS–polypyrimidine tract RNA, enhancing 3' splice site recognition. In vitro binding assays, cooperative RNA binding with deletion mutants of U2AF65 Genes & Development High 9512519
2001 Crystal structure of the core U2AF heterodimer (U2AF35 central domain + U2AF65 proline-rich region) at 2.2 Å reveals a novel 'tongue-in-groove' tryptophan-based protein–protein interaction; the interacting tryptophan residues are essential for U2AF dimerization and RNA binding by the heterodimer. X-ray crystallography, biochemical mutagenesis Cell High 11551507
2003 NMR structure of the C-terminal UHM domain (RRM3) of U2AF65 in complex with an SF1 N-terminal peptide shows SF1 inserts a conserved tryptophan into a hydrophobic pocket on the helical face of the RRM, establishing a paradigm for UHM–ULM protein recognition. NMR structure determination Molecular Cell High 12718882
2006 X-ray structure of U2AF65 RNA binding domain (RRM1-RRM2) bound to a poly-uridine polypyrimidine tract at 2.5 Å; specific hydrogen bonds between U2AF65 and uracil bases explain polyuridine recognition, with flexible side chains and water molecules enabling adaptation to variant Py tracts. Site-directed mutants confirmed energetic importance of conserved residues. X-ray crystallography, surface plasmon resonance with site-directed mutants Molecular Cell High 16818232
2009 Jmjd6 (an Fe(II)- and 2-oxoglutarate-dependent dioxygenase) catalyzes lysyl-5-hydroxylation of U2AF65, and Jmjd6 alters alternative RNA splicing of a subset of endogenous and reporter genes, implicating the hydroxylation of U2AF65 in splicing regulation. Mass spectrometry identification of hydroxylation, biochemical enzyme assay, reporter gene splicing assays, siRNA knockdown Science High 19574390
1997 U2AF65 is diffusely distributed in the nucleoplasm with additional concentration in nuclear speckles; the RS domain is specifically required for redistribution of U2AF65 to sites of active splicing (shown by adenovirus infection and transient expression of deletion mutants), while other domains are dispensable for speckle targeting. Monoclonal antibody immunofluorescence, transient expression of deletion mutants, adenovirus infection Journal of Cell Biology High 9166400
1998 WT1 (+KTS isoform) directly interacts with U2AF65 in vitro and in vivo, co-localizes with splicing factors in nuclear speckles, and can be incorporated into spliceosomes, indicating WT1 plays a role in pre-mRNA splicing via U2AF65 interaction. GST pulldown, co-immunoprecipitation, immunofluorescence co-localization, spliceosome fractionation Genes & Development High 9784496
2011 U2AF65 directly binds the phosphorylated CTD of RNA polymerase II; this interaction recruits U2AF65 and the Prp19 complex (PRP19C) to the pre-mRNA, providing a mechanism for CTD-dependent splicing activation during co-transcriptional splicing. Biochemical purification, in vitro binding assays with phospho-CTD, co-immunoprecipitation, in vitro splicing complementation Genes & Development High 21536736
2013 hnRNP C directly competes with U2AF65 for binding at Alu-element-derived cryptic splice sites genome-wide; loss of hnRNP C allows U2AF65 to bind Alu polypyrimidine tracts, leading to widespread aberrant exon inclusion from Alu elements. Quantitative iCLIP, minigene splicing assays, siRNA knockdown of hnRNP C Cell High 23374342
2009 MBNL1 competes with U2AF65 for mutually exclusive binding to the 3' end of intron 4 of cTNT pre-mRNA: MBNL1 binds a stem-loop structure while U2AF65 requires the same region in single-stranded form; when U2AF65 is blocked, U2 snRNP cannot be recruited and the downstream exon is skipped. In vitro binding competition assays, RNA structure analysis, mutations that stabilize stem-loop PNAS High 19470458
1995 Site-specific photochemical cross-linking showed that U2AF65 contacts the branch point at the earliest ATP-independent stage of spliceosome formation (E complex), positioning it to direct subsequent U2 snRNP recruitment to the upstream branchpoint. Site-specific UV cross-linking at branch point RNA High 7493318
2005 SF3b155 contains multiple U2AF65-binding ULM sites (at least five tryptophan-containing sites recognized by the U2AF65 UHM); thermodynamic analysis shows the SF1/U2AF65 complex is stabilized by 3.3 kcal/mol more than the SF3b155/U2AF65 complex, consistent with the need for ATP hydrolysis to drive exchange during spliceosome assembly. Intrinsic tryptophan fluorescence, circular dichroism, thermodynamic binding measurements Journal of Molecular Biology High 16376933
2006 SPSP phosphorylation of SF1 by KIS kinase (which bears a UHM domain required for interaction with SF1) increases binding of SF1 to U2AF65 and enhances formation of the ternary SF1–U2AF65–RNA complex. In vitro kinase assay, co-immunoprecipitation, RNA-binding assays FEBS Journal High 16420481
2012 NMR and SAXS structure of the SF1 N-terminal helix-hairpin domain in complex with the U2AF65 UHM reveals a secondary hydrophobic interface that locks the orientation of the two subunits; mutagenesis shows the helix-hairpin is essential for cooperative ternary SF1–U2AF65–RNA complex formation; tandem serine phosphorylation of SF1 SPSP motif rigidifies the linker and slightly enhances RNA binding. NMR, SAXS, mutagenesis, ternary complex binding assays Nucleic Acids Research High 23175611
2019 OTUB2 deubiquitinase directly binds U2AF2 and removes ubiquitin from it, preventing its proteasomal degradation and stabilizing U2AF2 protein levels in NSCLC cells. Co-immunoprecipitation, mass spectrometry, ubiquitination assay, xenograft model Theranostics Medium 30662561
2016 CD82 suppresses U2AF2 activity by inducing its ubiquitination, leading to proteasomal degradation of U2AF2, which reduces U2AF2-mediated CD44 variant exon splicing and inhibits melanoma metastasis. Ubiquitination assays, in vitro and in vivo invasion/metastasis assays, siRNA knockdown Oncogene Medium 27041584
2014 Combined NMR and SAXS analysis of multidomain U2AF65 reveals a highly anisotropic conformational space dominated by transient electrostatic interdomain contacts; the small subpopulation with a preformed RNA-bound domain arrangement is selected by RNA via conformational selection. NMR paramagnetic relaxation enhancement, SAXS Journal of the American Chemical Society High 24734879
2018 In vitro iCLIP experiments show that U2AF2 binds RNA autonomously based on polypyrimidine tract strength, but trans-acting RBPs (including FUBP1, CELF6, and PCBP1) extensively modulate U2AF2 binding in vivo—enhancing recruitment to 3' splice sites and clearing from intron bodies—thereby altering splicing outcomes. In vitro iCLIP, mathematical modeling of in vitro vs in vivo binding, machine learning, minigene splicing validation Genome Research High 29643205
2020 NMR-based structural analysis combined with iCLIP shows that the intrinsically disordered linker region between U2AF2's RRM1 and RRM2 mediates autoinhibitory intramolecular interactions that reduce nonproductive binding to weak Py-tract RNAs, thereby proofreading U2AF2 binding toward stronger Py-tracts at bona fide 3' splice sites. Linker mutations that impair autoinhibition cause promiscuous binding and splicing errors. NMR, iCLIP, RNA binding assays, linker mutagenesis, splicing assays PNAS High 32188783
2020 Single-molecule FRET shows that U2AF1 stabilizes a closed (high-FRET) conformation of the tandem U2AF2 RRMs in the absence of RNA; binding to a strong uridine-rich splice site switches U2AF2 to an open conformation, while weak Py tracts yield a mixture of open and closed states. The MDS-associated U2AF1 S34F mutation modulates this conformational equilibrium. Single-molecule FRET, structure-guided mutagenesis Nucleic Acids Research High 32343311
2022 Crystal structures of U2AF2 bound to cytidine, guanosine, or adenosine at the central Py-tract position show that local RNA flexibility accommodates different nucleotides while the polypeptide backbone remains unchanged; mutational and eCLIP analyses confirm U2AF2 tolerates nucleotide substitutions at the central position. X-ray crystallography, molecular dynamics, RNA binding affinity measurements, eCLIP Nucleic Acids Research High 35524551
2020 Crystal structures of cancer-associated U2AF2 mutants (N196K and G301D) bound to polypyrimidine RNA show that N196K stabilizes an open inter-RRM conformation (increased RNA affinity), while G301D creates unfavorable proximity to RNA phosphodiester (decreased RNA affinity); both mutants alter splicing of endogenous transcripts. X-ray crystallography, RNA binding affinity assays, minigene splicing assays Journal of Biological Chemistry High 33020180
2021 A small-molecule compound (NSC-194308) enhances U2AF2 RNA binding by bridging the two tandem RRMs via hydrophobic and electrostatic interactions (identified by computational docking and structure-guided mutagenesis), stalls spliceosome assembly at the U2AF2 function stage, and inhibits splicing of representative substrates. Chemical screen, RNA binding assays, in vitro splicing, computational docking, structure-guided mutagenesis Cell Chemical Biology High 33689684
2016 Crystal and NMR structures of RBM39-UHM bound to U2AF65-ULM, confirmed by Co-IP from human cell extracts and ITC, establish common and discriminating recognition elements among UHM–ULM splicing factor interactions. X-ray crystallography, NMR, ITC, co-immunoprecipitation Acta Crystallographica Section D High 27050129
2022 Crystal structure of the U2AF2 UHM bound to a SF3B1 ULM site at 1.8 Å reveals a distinctive ULM trajectory across the U2AF2 UHM surface; mutagenesis at the interface disrupts co-immunoprecipitation and alters splicing of endogenous transcripts; genome-wide analysis shows U2AF2–SF3B1 and U2AF2–SF1 co-regulate distinct splice sites. X-ray crystallography, ITC, co-immunoprecipitation, splicing assays, genome-wide analysis Journal of Biological Chemistry High 35780835
2013 U2AF65 promotes its own binding only at weaker polypyrimidine tracts; U2AF65 inhibits splicing of flanking introns of alternative exons in both three-exon and two-exon contexts, demonstrating a splicing inhibitory function that contributes to alternative exon skipping in SMN and Fas pre-mRNAs. Overexpression, minigene splicing assays, splice site mutagenesis, UV cross-linking PNAS Medium 26216990
2011 U2AF65 directly and specifically binds expanded CAG RNA via its RRM3 domain, and forms an RNA/protein complex with the NXF1 nuclear export receptor; U2AF65 acts as an adaptor linking expanded CAG RNA to NXF1 for nuclear export, as demonstrated in Drosophila U2AF50 (ortholog) knockdown and mammalian cell studies. Co-immunoprecipitation, domain mapping (RRM3 deletion), Drosophila genetic knockdown, transgenic mouse analysis Human Molecular Genetics Medium 21725067
2013 U2AF65 stabilizes TRF1 protein by directly interacting with it and interfering with Fbx4 (an E3 ubiquitin ligase for TRF1), thereby inhibiting TRF1 ubiquitin-dependent proteolysis. U2AF65 knockdown reduces TRF1 stability while U2AF65 overexpression extends TRF1 half-life. Co-immunoprecipitation, GST pulldown, siRNA knockdown, ubiquitination assay, protein stability assay Biochemical and Biophysical Research Communications Medium 24389012
2001 Splicing activators promote binding of both U2AF65 and U2AF35 to weak 3' splice sites; U2AF35 is specifically required for maximum enhancer-dependent splicing and acts after U2AF65 has bound the polypyrimidine tract, demonstrating substrate-specific requirement for U2AF35. UV cross-linking, in vitro splicing assays, recombinant protein complementation RNA High 11421359
1999 Recombinant U2AF65 alone is sufficient for splicing of constitutively spliced pre-mRNAs in U2AF-depleted extracts, but U2AF35 and its interaction with U2AF65 are additionally required for substrates with weak Py tracts and exonic splicing enhancers; U2AF35's function is not to change U2AF65 cross-linking to the Py tract. Chromatographic depletion of U2AF, recombinant complementation, UV cross-linking Molecular and Cellular Biology High 10567551
2006 PTB directly displaces U2AF65 from the polypyrimidine tract upstream of the beta-tropomyosin exon 6B 3' splice site, preventing E complex assembly and U2 snRNA–branch point pairing, thereby repressing splicing. UV cross-linking, immunoprecipitation, PTB knockdown by RNAi, psoralen cross-linking, in vitro splicing assays Molecular and Cellular Biology High 16982681
2019 U2AF65 RS domain drives liquid-liquid phase separation amplified by intronic RNA with repeated pyrimidine tracts; U2AF65 cooperatively binds the multi-ULM domain of SF3b155 together with CAPERα; knockdown of U2AF65 or CAPERα increases inclusion of cassette exons preceded by repeated pyrimidine-rich motifs. Phase separation assay, co-immunoprecipitation, siRNA knockdown, splicing assays EMBO Reports Medium 31271494
2013 MBNL1 activates insulin receptor exon 11 inclusion by binding a downstream intronic enhancer, which enhances binding of U2AF65 to the polypyrimidine tract of the upstream intron and promotes spliceosome A complex formation; 5' splice site is required for MBNL1-activated upstream intron removal but not for U2AF65 recruitment. In vitro splicing assay, UV cross-linking of U2AF65, spliceosomal complex analysis Nucleic Acids Research High 24185704
2024 SIRT4, upon TGF-β stimulation and nuclear translocation, deacetylates U2AF2 at K413 in the nucleus, facilitating U2AF2-mediated alternative splicing of CCN2 pre-mRNA and promoting CCN2 protein expression and renal fibrosis. Co-immunoprecipitation, deacetylation assay, siRNA knockdown, knockout mice, mass spectrometry eLife Medium 39495216
2025 CUT&RUN profiling reveals U2AF2 binds chromatin throughout gene bodies of intron-containing genes in a histone H3K36me3-dependent but nascent-transcript-independent manner; chromatin-bound U2AF2 preferentially associates with exons that are skipped upon U2AF2 knockdown, indicating that chromatin association enhances exon selection accuracy during co-transcriptional splicing. CUT&RUN chromatin profiling, RNase A treatment, H3K36me3 dependency analysis, U2AF2 knockdown + RNA-seq Molecular Cell High 40315850
2017 JMJD6 and U2AF65 co-regulate a large number of alternative splicing events; JMJD6's enzymatic hydroxylation activity is required for a subset of co-regulated splicing events; JMJD6-mediated lysine hydroxylation of U2AF65 accounts at least partially for their co-regulated alternative splicing, demonstrated in jmjd6 knockout mice. RASL-Seq, jmjd6 knockout mice, enzymatic activity mutants, RNA-dependent interaction analysis Nucleic Acids Research High 27899633
2002 The U2AF35 RRM is unstructured in solution but its tertiary structure is induced upon binding to U2AF65 (specifically the N-terminal proline-rich region of U2AF65), demonstrating an induced folding mechanism for heterodimer assembly. NMR spectroscopy of free and U2AF65-bound U2AF35 RRM FEBS Letters High 12297299
2021 AKT3-phosphorylated IWS1 regulates H3K36me3 deposition to control cell cycle-dependent inclusion of U2AF2 exon 2; loss of exon 2 produces an RS-domain-deficient U2AF65 that fails to process CDCA5 pre-mRNA, reducing Sororin protein and causing G2/M arrest. RNA-seq in phospho-IWS1-deficient cells, RT-PCR, splicing reporter assays, xenograft models Nature Communications Medium 34330897
2021 U2AF2 binds the polypyrimidine tract of IL7R intron 5 and competes with PTBP1; additionally, U2AF2 binds an ectopic polypyrimidine tract following an exonic branch point sequence within IL7R exon 6, recruiting U2 snRNP components to the exon and thereby repressing exon 6 inclusion. RNA pull-down, siRNA knockdown, minigene splicing assays, evolutionary conservation analysis RNA Medium 33568552
2018 HNRNPA1 overexpression shifts U2AF2 binding from bona fide 3' splice sites to decoy binding sites including Alu-derived intronic sequences for alternative cassette exons, as shown by iCLIP; HNRNPA1 also directly interacts with the U2AF heterodimer to modulate its RNA binding. iCLIP (individual-nucleotide resolution CLIP), HNRNPA1 overexpression Genome Research Medium 29650551

Source papers

Stage 0 corpus · 100 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2013 Direct competition between hnRNP C and U2AF65 protects the transcriptome from the exonization of Alu elements. Cell 377 23374342
2009 Jmjd6 catalyses lysyl-hydroxylation of U2AF65, a protein associated with RNA splicing. Science (New York, N.Y.) 325 19574390
1996 Interaction of U2AF65 RS region with pre-mRNA branch point and promotion of base pairing with U2 snRNA [corrected]. Science (New York, N.Y.) 248 8781232
1997 U2AF65 recruits a novel human DEAD box protein required for the U2 snRNP-branchpoint interaction. Genes & development 228 9242493
1998 WT1 interacts with the splicing factor U2AF65 in an isoform-dependent manner and can be incorporated into spliceosomes. Genes & development 216 9784496
1998 A cooperative interaction between U2AF65 and mBBP/SF1 facilitates branchpoint region recognition. Genes & development 213 9512519
1992 U1 snRNP targets an essential splicing factor, U2AF65, to the 3' splice site by a network of interactions spanning the exon. Genes & development 192 1285125
2003 Structural basis for the molecular recognition between human splicing factors U2AF65 and SF1/mBBP. Molecular cell 184 12718882
2001 A novel peptide recognition mode revealed by the X-ray structure of a core U2AF35/U2AF65 heterodimer. Cell 184 11551507
2005 Steroid hormone receptor coactivation and alternative RNA splicing by U2AF65-related proteins CAPERalpha and CAPERbeta. Molecular cell 168 15694343
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