Affinage

UPF3A

Regulator of nonsense transcripts 3A · UniProt Q9H1J1

Length
476 aa
Mass
54.7 kDa
Annotated
2026-06-10
26 papers in source corpus 13 papers cited in narrative 13 extracted findings
Cross-family judge vs UniProt: Affinage preferred faithfulness: 6/6 claims corpus-supported (100%)

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

UPF3A is a paralog of UPF3B that acts as a context-dependent modulator of nonsense-mediated mRNA decay (NMD), a nuclear-and-cytoplasmic surveillance pathway in which it bridges the exon-junction complex (EJC) to the core UPF machinery (PMID:11113196, PMID:16601204). UPF3A engages UPF2 through shared structural elements—an RRM-like domain, a NONA/paraspeckle-like (NOPS-L) domain, and an extended α-helical region that also support RNA/ribosome binding and RNA-induced oligomerization—and binds UPF2 with ~10-fold higher affinity than UPF3B, competing for the same interface (PMID:35640974). Because UPF3A is intrinsically a weaker NMD activator than UPF3B, this competition allows UPF3A to behave as a potent NMD inhibitor that stabilizes hundreds of transcripts when UPF3B is present; conditional loss of UPF3A in mice causes hyperactive NMD and defects in embryogenesis and gametogenesis (PMID:27040500). When UPF3B is absent, however, UPF3A switches to an activator, driving NMD through its conserved 'mid' domain independently of EJC binding, and the two paralogs are functionally redundant such that only co-depletion strongly blocks NMD (PMID:35451102, PMID:35451084). This activity is held in balance by a post-transcriptional switch in which UPF3B promotes destabilization of UPF3A protein, so that UPF3A levels rise to partially compensate only when UPF3B is lost (PMID:19503078). Beyond canonical NMD, UPF3A is required—together with COMPASS components such as Wdr5—for the genetic compensation response triggered by premature-termination-codon-bearing mRNAs, transcriptionally upregulating homologous genes via enhanced H3K4me3, while a distinct homology-dependent compensation role in zebrafish operates in an H3K4me3-independent manner (PMID:30944473, PMID:37369707).

Mechanistic history

Synthesis pass · year-by-year structured walk · 11 steps
  1. 2001 Medium

    Established UPF3A as a human NMD-pathway component by showing it, like UPF3B, binds the central adaptor UPF2 and shuttles between nucleus and cytoplasm, framing NMD as a process with both compartmental phases.

    Evidence Reciprocal Co-IP of epitope-tagged proteins with domain mapping and immunofluorescence in HeLa cells

    PMID:11113196

    Open questions at the time
    • Did not resolve why UPF3A and UPF3B differ functionally
    • No quantitative comparison of UPF2-binding affinity
  2. 2004 High

    Defined the atomic basis of the UPF2-UPF3 interaction, showing the UPF3 RNP/RRM-like domain docks onto UPF2 via conserved charged residues and that this domain does not itself bind RNA, clarifying the architecture shared by UPF3A.

    Evidence 1.95 Å crystal structure of UPF2 MIF4G–UPF3B RNP complex plus RNA-binding assays

    PMID:15004547

    Open questions at the time
    • Structure was of UPF3B, not UPF3A directly
    • Did not explain functional divergence of the paralogs
  3. 2006 Medium

    Showed UPF3A is intrinsically a weaker NMD inducer than UPF3B and localized the discriminating function to a C-terminal EJC-binding region, separating EJC-dependent NMD induction from EJC-independent translation stimulation.

    Evidence lambdaN/boxB tethering reporter assays with EJC-interaction Co-IP mapping

    PMID:16601204

    Open questions at the time
    • Tethering bypasses normal mRNP recruitment
    • Did not establish physiological consequence of weak UPF3A activity
  4. 2007 High

    Reconstituted the heptameric EJC–UPF1–UPF2–UPF3 complex in vitro and showed UPF2/UPF3 cooperatively stimulate UPF1 ATPase and helicase activity, placing UPF3 mechanistically as an activator of the central NMD enzyme.

    Evidence In vitro reconstitution with recombinant proteins plus ATPase and RNA helicase assays

    PMID:18066079

    Open questions at the time
    • Assays used UPF3B; UPF3A-specific enzymatic stimulation not measured
    • In vitro stoichiometry may not reflect cellular complexes
  5. 2009 High

    Revealed a post-transcriptional regulatory switch in which UPF3B destabilizes UPF3A protein, so UPF3A rises and partially compensates only when UPF3B is lost—while simultaneously impairing NMD through UPF2 competition.

    Evidence Western blotting in UPF3B-mutant patient cells, competitive binding and NMD reporter assays

    PMID:19503078

    Open questions at the time
    • Molecular mechanism of UPF3A destabilization not identified
    • Did not quantify net NMD outcome in normal cells
  6. 2016 High

    Reframed UPF3A as primarily a potent NMD inhibitor whose loss causes hyper-NMD and developmental defects, attributing its repressor role to a weakened EJC-binding domain that lets it displace the stronger activator UPF3B.

    Evidence siRNA/shRNA, conditional knockout mouse, RNA-seq, NMD reporters and Co-IP

    PMID:27040500

    Open questions at the time
    • Did not test UPF3A function in the complete absence of UPF3B
    • Mechanism reconciling inhibition with later activator findings unresolved
  7. 2019 High

    Identified an NMD-independent role for Upf3a in the genetic compensation response, linking PTC-bearing mRNA to COMPASS/Wdr5-mediated H3K4me3 deposition and transcriptional upregulation of homologous genes.

    Evidence Zebrafish knockdown-vs-knockout models, transgenes, H3K4me3 ChIP and genetic epistasis

    PMID:30944473

    Open questions at the time
    • How Upf3a couples PTC mRNA to chromatin modifiers is unknown
    • Conservation of GCR mechanism in mammals not established
  8. 2020 Medium

    Indicated UPF3A influences pathways beyond NMD, with reconstitution altering cholesterol-biosynthesis enzyme levels and the phosphorylation of nuclear gene-expression/splicing regulators.

    Evidence SILAC quantitative proteomics and phosphoproteomics in UPF3A-reconstituted CRC cells

    PMID:32718059

    Open questions at the time
    • Single CRC cell line, single lab
    • Direct versus indirect effects of UPF3A not distinguished
  9. 2022 High

    Provided the quantitative and structural basis for paralog competition: crystal structures of UPF2 bound to either UPF3A or UPF3B showed shared interfaces, UPF3A binds UPF2 ~10-fold tighter, and a disease UPF3B mutation in NOPS-L weakens UPF2 binding ~40-fold.

    Evidence X-ray crystallography, affinity measurements, mutagenesis and RNA/ribosome-binding assays

    PMID:35640974

    Open questions at the time
    • Did not resolve how tighter UPF2 binding maps onto inhibitor-versus-activator behavior
    • In-cell stoichiometry of competition not measured
  10. 2022 High

    Resolved the activator/inhibitor paradox by showing UPF3A strongly activates NMD when UPF3B is deleted and that the EJC-binding domain is dispensable while the conserved 'mid' domain drives activity, with the two paralogs functionally redundant.

    Evidence CRISPR knockout and combinatorial co-depletion, NMD reporters, RNA-seq and domain-mutant complementation across two parallel studies

    PMID:35451084 PMID:35451102

    Open questions at the time
    • Biochemical function of the 'mid' domain undefined
    • Why outcomes differ between cell systems not fully explained
  11. 2023 Medium

    Refined the in vivo NMD picture and the compensation mechanism: UPF3A is largely dispensable for NMD when UPF3B is present in mouse, and in zebrafish leg1 mutants Upf3a drives homology-dependent compensation in an H3K4me3-independent manner.

    Evidence Conditional knockout mice with NMD-target qRT-PCR; zebrafish single/double knockouts with RNA-seq and ULI-NChIP-seq

    PMID:36997282 PMID:37369707

    Open questions at the time
    • Reconciliation of H3K4me3-dependent and -independent compensation modes unresolved
    • Molecular effectors of homology-dependent correction unknown

Open questions

Synthesis pass · forward-looking unresolved questions
  • The molecular mechanism by which the conserved 'mid' domain activates NMD, and how UPF3A links PTC-containing mRNA to transcriptional compensation, remain unresolved.
  • No biochemical activity assigned to the 'mid' domain
  • No identified molecular bridge from UPF3A to COMPASS/chromatin in the GCR
  • Mechanism of UPF3B-dependent UPF3A destabilization unknown

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0060090 molecular adaptor activity 3 GO:0098772 molecular function regulator activity 3 GO:0003723 RNA binding 1
Localization
GO:0005634 nucleus 1 GO:0005829 cytosol 1
Pathway
R-HSA-8953854 Metabolism of RNA 4 R-HSA-74160 Gene expression (Transcription) 2
Partners
EIF4A3MAGOHRBM8AUPF1UPF2UPF3BWDR5
Complex memberships
EJC-UPF surveillance complex (UPF1-UPF2-UPF3-EJC)

Evidence

Reading pass · 13 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2001 Human UPF3A (hUpf3p) was identified as a human orthologue of S. cerevisiae Upf3p; co-immunoprecipitation of epitope-tagged proteins in HeLa cells demonstrated that hUpf2p interacts with both hUpf3p-X (UPF3B) and hUpf3p (UPF3A), and the domains required for these interactions were defined. UPF3A localizes primarily to nuclei and is a shuttling protein, indicating NMD has both nuclear and cytoplasmic components. Co-immunoprecipitation of epitope-tagged proteins in HeLa cells; indirect immunofluorescence for subcellular localization Molecular and cellular biology Medium 11113196
2004 Crystal structure at 1.95 Å of the complex between the interacting domains of human UPF2 (MIF4G domain) and UPF3B (RNP domain) revealed that the protein-protein interface is mediated by highly conserved charged residues and involves the beta-sheet surface of the UPF3B RNP domain. The UPF3b RNP domain does not bind RNA, whereas the UPF2 construct and the complex do. The same RNP domain architecture is shared by UPF3A. X-ray crystallography (1.95 Å resolution); RNA-binding assays Nature structural & molecular biology High 15004547
2006 Using a tethering system, UPF3A was shown to be much less active than UPF3B in inducing NMD and stimulating translation. The C-terminal domain region that discriminates UPF3A from UPF3B in NMD function mediates interaction with EJC components Y14, Magoh, BTZ, and eIF4AIII; this interaction is required for NMD induction. Translation stimulation is independent of EJC interaction and is determined by other regions of the UPF3 proteins. lambdaN/boxB tethering reporter assay; co-immunoprecipitation for EJC interaction mapping RNA (New York, N.Y.) Medium 16601204
2007 Recombinant EJC core was sufficient to reconstitute a stable heptameric complex on RNA with UPF1, UPF2, and UPF3b. EJC proteins MAGOH, Y14, and eIF4AIII provide a composite binding site for UPF3b that bridges to UPF2 and UPF1. In the trimeric UPF complex, UPF2 and UPF3b cooperatively stimulate both ATPase and RNA helicase activities of UPF1. In vitro reconstitution; ATPase assay; RNA helicase assay with recombinant proteins Nature structural & molecular biology High 18066079
2009 UPF3A protein levels are regulated post-transcriptionally: the presence of UPF3B promotes destabilization/reduction of UPF3A levels via a conserved UPF3B-dependent mechanism. When UPF3B is absent (e.g., due to mutations), UPF3A levels rise and partially compensate for NMD, but UPF3A also impairs NMD by competing with UPF3B for binding to UPF2. This UPF3B-dependent destabilization of UPF3A constitutes a post-transcriptional regulatory switch maintaining appropriate NMD levels. Western blotting of UPF3A/UPF3B levels in cells with UPF3B mutations; competitive binding assays; NMD reporter assays Nature structural & molecular biology High 19503078
2016 UPF3A acts primarily as a potent NMD inhibitor, stabilizing hundreds of transcripts, rather than an NMD activator. It acquired repressor activity through impairment of a critical domain (C-terminal EJC-binding domain is weakened relative to UPF3B). Conditional knockout of UPF3A in mice causes 'hyper' NMD and leads to defects in embryogenesis and gametogenesis. UPF3A competes with UPF3B for binding to UPF2, and its NMD-inhibitory function is explained by its weaker NMD activation capacity displacing the stronger activator UPF3B. Loss-of-function in vitro (siRNA/shRNA) and in vivo (conditional knockout mouse); RNA-seq; NMD reporter assays; co-immunoprecipitation Cell High 27040500
2019 In zebrafish, Upf3a (a NMD pathway member) and components of the COMPASS complex including Wdr5 are required for the genetic compensation response (GCR) triggered by mRNAs bearing a premature termination codon (PTC). The GCR is accompanied by enhancement of H3K4me3 at the transcription start site regions of compensatory genes, linking Upf3a to transcriptional upregulation of homologous genes in response to PTC-containing mRNA. Zebrafish knockdown vs. knockout models; transgene analysis; ChIP for H3K4me3; genetic epistasis using upf3a mutants Nature High 30944473
2020 Reconstituted UPF3A expression in KM12 CRC cells (which have a frameshift mutation in UPF3A) caused down-regulation of several enzymes involved in cholesterol biosynthesis and altered phosphorylation of 85 phosphosites in 52 phosphoproteins, predominantly nuclear proteins involved in gene expression regulation and RNA splicing, suggesting UPF3A influences cellular signaling pathways beyond NMD. SILAC-based quantitative proteomics; phosphoproteomics in CRC cell lines with reconstituted UPF3A expression International journal of molecular sciences Medium 32718059
2022 UPF3A and UPF3B share structural homology comprising an RRM-like domain (RRM-L), a NONA/paraspeckle-like domain (NOPS-L), and an extended α-helical domain; these domains are essential for RNA/ribosome-binding, RNA-induced oligomerization, and UPF2 interaction. Crystal structures of UPF2's MIF4GIII domain in complex with UPF3B or UPF3A revealed intimate binding interfaces. UPF3A binds UPF2 with ~10-fold higher affinity than UPF3B. The disease-causing UPF3B mutation Y160D in the NOPS-L domain displaces Y160 from a hydrophobic cleft in UPF2, reducing binding affinity ~40-fold. UPF3A and UPF3B compete for the same UPF2 binding site. X-ray crystallography; binding affinity measurements; mutagenesis; RNA/ribosome binding assays Nucleic acids research High 35640974
2022 In HCT116 cells deleted for UPF3B, UPF3A strongly activates NMD; in cells lacking both UPF3A and UPF3B, NMD is only partially active. Complementation studies show the EJC-binding domain of UPF3 paralogs is dispensable for NMD; instead, the conserved 'mid' domain is consequential for NMD activity. UPF3A can activate NMD independently of EJC binding. CRISPR knockout cell lines; NMD reporter assays; RNA-seq; complementation with domain mutants The EMBO journal High 35451102
2022 Co-depletion of UPF3A and UPF3B (but not single depletion of either) results in marked NMD inhibition and transcriptome-wide upregulation of NMD substrates, demonstrating functional redundancy between UPF3A and UPF3B. Rescue experiments show UPF2-binding or EJC-binding-deficient UPF3B largely retains NMD activity, but deletion of the middle domain combined with other mutations synergistically impairs NMD. siRNA knockdown and CRISPR knockout; RNA-seq; rescue experiments with domain mutants The EMBO journal High 35451084
2023 In zebrafish leg1 deleterious mutants, Upf3a (but not Upf1) is essential for the homology-dependent genetic compensation response (HDGCR) induced by nonsense mutations; this occurs in an H3K4me3-independent manner. Upf3a is also responsible for correcting the expression of hundreds of genes dysregulated in leg1 mutants. Zebrafish single and double knockout mutants; RNA-seq (71 samples); ULI-NChIP-seq for H3K4me3; genetic epistasis Cell discovery High 37369707
2023 In mouse embryonic stem cells, somatic cells, and major organs (liver, spleen, thymus), UPF3A is dispensable for NMD when UPF3B is present; UPF3A may weakly and selectively promote NMD in certain murine organs. UPF3A does not repress NMD in these contexts. Conditional knockout mouse (Upf3a); qRT-PCR and RNA analysis of 33 NMD targets in multiple cell lines and organs Life science alliance Medium 36997282

Source papers

Stage 0 corpus · 26 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2019 PTC-bearing mRNA elicits a genetic compensation response via Upf3a and COMPASS components. Nature 375 30944473
2007 NMD factors UPF2 and UPF3 bridge UPF1 to the exon junction complex and stimulate its RNA helicase activity. Nature structural & molecular biology 274 18066079
2001 Identification and characterization of human orthologues to Saccharomyces cerevisiae Upf2 protein and Upf3 protein (Caenorhabditis elegans SMG-4). Molecular and cellular biology 208 11113196
2004 The structural basis for the interaction between nonsense-mediated mRNA decay factors UPF2 and UPF3. Nature structural & molecular biology 166 15004547
2016 The Antagonistic Gene Paralogs Upf3a and Upf3b Govern Nonsense-Mediated RNA Decay. Cell 133 27040500
2006 Functions of hUpf3a and hUpf3b in nonsense-mediated mRNA decay and translation. RNA (New York, N.Y.) 110 16601204
2009 A UPF3-mediated regulatory switch that maintains RNA surveillance. Nature structural & molecular biology 101 19503078
2011 Nonsense-mediated mRNA decay factors, UPF1 and UPF3, contribute to plant defense. Plant & cell physiology 61 22025558
2011 Drosophila Upf1 and Upf2 loss of function inhibits cell growth and causes animal death in a Upf3-independent manner. RNA (New York, N.Y.) 54 21317294
2008 Upf1 potentially serves as a RING-related E3 ubiquitin ligase via its association with Upf3 in yeast. RNA (New York, N.Y.) 46 18676617
2022 Mammalian UPF3A and UPF3B can activate nonsense-mediated mRNA decay independently of their exon junction complex binding. The EMBO journal 43 35451102
2022 Human UPF3A and UPF3B enable fault-tolerant activation of nonsense-mediated mRNA decay. The EMBO journal 36 35451084
1998 The upf3 protein is a component of the surveillance complex that monitors both translation and mRNA turnover and affects viral propagation. Proceedings of the National Academy of Sciences of the United States of America 35 9671745
2015 The feedback control of UPF3 is crucial for RNA surveillance in plants. Nucleic acids research 22 25820429
2023 Upf3a but not Upf1 mediates the genetic compensation response induced by leg1 deleterious mutations in an H3K4me3-independent manner. Cell discovery 20 37369707
2022 Structures of nonsense-mediated mRNA decay factors UPF3B and UPF3A in complex with UPF2 reveal molecular basis for competitive binding and for neurodevelopmental disorder-causing mutation. Nucleic acids research 17 35640974
2023 Serum methylation of GALNT9, UPF3A, WARS, and LDB2 as noninvasive biomarkers for the early detection of colorectal cancer and advanced adenomas. Clinical epigenetics 14 37794510
2002 Role of RNA surveillance proteins Upf1/CpaR, Upf2 and Upf3 in the translational regulation of yeast CPA1 gene. Current genetics 14 12172963
2020 Functional roles of human Up-frameshift suppressor 3 (UPF3) proteins: From nonsense-mediated mRNA decay to neurodevelopmental disorders. Biochimie 11 33132159
2023 UPF3A is dispensable for nonsense-mediated mRNA decay in mouse pluripotent and somatic cells. Life science alliance 10 36997282
2020 Functions and Clinical Significance of UPF3a Expression in Human Colorectal Cancer. Cancer management and research 10 32606924
2020 (Phospho)proteomic Profiling of Microsatellite Unstable CRC Cells Reveals Alterations in Nuclear Signaling and Cholesterol Metabolism Caused by Frameshift Mutation of NMD Regulator UPF3A. International journal of molecular sciences 5 32718059
2023 Genetic compensation response could exist in colorectal cancer: UPF3A upregulates the oncogenic homologue gene SRSF3 expression corresponding to SRSF6 to promote colorectal cancer metastasis. Journal of gastroenterology and hepatology 4 36807382
2014 An UPF3-based nonsense-mediated decay in Paramecium. Research in microbiology 4 25463387
2023 Spatial expression of the nonsense-mediated mRNA decay factors UPF3A and UPF3B among mouse tissues. Journal of Zhejiang University. Science. B 2 37961809
2026 The nonsense-mediated mRNA decay factor Upf3 negatively regulates bulk autophagy progression in Saccharomyces cerevisiae. Autophagy reports 0 41668711

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