Affinage

NUDT16

U8 snoRNA-decapping enzyme · UniProt Q96DE0

Length
195 aa
Mass
21.3 kDa
Annotated
2026-04-29
15 papers in source corpus 13 papers cited in narrative 13 extracted findings

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

NUDT16 is a multifunctional Nudix hydrolase that integrates nucleotide pool sanitization, RNA decapping, and ADP-ribose processing to maintain genome integrity and regulate the DNA damage response. As a (deoxy)inosine diphosphatase, NUDT16 hydrolyzes IDP and dIDP in the nucleus, and its depletion causes S-phase arrest, single-strand breaks, and chromosome abnormalities, functioning redundantly with ITPA to protect genomic DNA from inosine misincorporation (PMID:20385596, PMID:20081199). NUDT16 also possesses metal-dependent RNA-decapping activity, cleaving m7G and m2,2,7G caps from RNAs including U8 snoRNA, with substrate selectivity governed by the first transcribed nucleotide and cap methylation status (PMID:17567574, PMID:29483298). Critically, NUDT16 functions as a dePARylase that removes ADP-ribosylation from DNA-damage-response proteins—including 53BP1, CtIP, NBS1, SETD3, and HMGA1—thereby preventing their RNF146- or CHFR-mediated ubiquitination and proteasomal degradation, which is essential for their recruitment to double-strand breaks and for homologous recombination repair and replication fork protection (PMID:31911551, PMID:38324469, PMID:39438373, PMID:38272222, PMID:40288645).

Mechanistic history

Synthesis pass · year-by-year structured walk · 10 steps
  1. 2007 High

    The identification of Nudt16 (X29/H29K) as a metal-dependent nuclear RNA decapping enzyme established its first known catalytic activity and revealed how divalent cation identity switches substrate specificity from U8 snoRNA-selective to broad-spectrum decapping.

    Evidence X-ray crystallography of Xenopus X29, active-site mutagenesis, and in vitro decapping assays with human homolog

    PMID:17567574

    Open questions at the time
    • Physiological RNA targets of NUDT16 decapping in human cells not identified
    • In vivo contribution of decapping activity versus other enzymatic activities unknown
  2. 2008 Medium

    Evolutionary analysis demonstrated that Nudt16 RNA-binding and decapping activity is conserved across metazoans, distinguishing it from its catalytically inactive paralog Syndesmos and establishing Nudt16 as an ancient RNA-processing enzyme.

    Evidence In vitro RNA-binding and decapping assay with insect ortholog; phylogenetic reconstruction

    PMID:18820299

    Open questions at the time
    • Functional divergence between Nudt16 and Syndesmos in vivo not tested
    • Cellular RNA substrates across species remain uncharacterized
  3. 2010 High

    Discovery that NUDT16 is primarily a nuclear (deoxy)inosine diphosphatase reframed its principal enzymatic role from RNA decapping to nucleotide pool sanitization, and loss-of-function experiments revealed that this activity is essential for S-phase progression and DNA integrity.

    Evidence Enzymatic screening against 29 substrates with purified protein; siRNA knockdown in HeLa cells with comet assay, cell-cycle analysis, and immunofluorescence; Itpa−/− MEFs with Nudt16 knockdown

    PMID:20081199 PMID:20385596

    Open questions at the time
    • Relative in vivo contributions of inosine diphosphatase versus decapping activity not dissected
    • No in vivo knockout model in a whole organism reported
  4. 2015 High

    The crystal structure of human NUDT16 in complex with its product IMP revealed a dimeric architecture and a positively charged substrate-binding trench, explaining purine selectivity through positions 2 and 6 of the ring and identifying product-release as the rate-limiting step.

    Evidence X-ray crystallography (apo and IMP-bound forms); steady-state kinetics; biophysical binding studies

    PMID:26121039

    Open questions at the time
    • No structure with intact nucleoside diphosphate substrate to capture catalytic geometry
    • Structural basis for discrimination between IDP and other NDP substrates not fully resolved
  5. 2018 Medium

    Systematic analysis of cap analog and capped oligonucleotide substrates refined understanding of NUDT16 decapping selectivity, showing that unmethylated caps and guanine-first transcribed nucleotides are preferred, bridging the nucleotide hydrolase and decapping activities through shared active-site chemistry.

    Evidence In vitro enzymatic screening with purified hNudt16 against dinucleotide cap analogs and capped oligonucleotides

    PMID:29483298

    Open questions at the time
    • Cellular mRNA targets regulated by NUDT16 decapping remain unidentified
    • Relative importance of cap methylation versus first nucleotide identity in vivo unknown
  6. 2019 High

    Structural and biochemical demonstration that NUDT16 processes both free and protein-conjugated ADP-ribose revealed a third major substrate class; engineered active-site mutants that selectively favor protein-conjugated poly(ADP-ribose) provided tools to separate dePARylation from other activities.

    Evidence X-ray crystallography with mono- and di-ADP-ribose complexes; rational mutagenesis (Δ17, F36A, F61S); activity assays on free and conjugated substrates

    PMID:30976021

    Open questions at the time
    • Endogenous protein substrates of dePARylation not yet identified in this study
    • In vivo relevance of dePARylation versus free ADP-ribose hydrolysis not separated
  7. 2020 High

    Identification of 53BP1 as the first endogenous dePARylation substrate of NUDT16 established a direct link to the DNA damage response: NUDT16 removes ADP-ribosylation from 53BP1 to prevent RNF146-mediated ubiquitination, thereby stabilizing 53BP1 and enabling its recruitment to DSBs.

    Evidence Co-IP; catalytically inactive mutant overexpression; ubiquitination assay; IR-induced foci; cell survival after DNA damage

    PMID:31911551

    Open questions at the time
    • PARP responsible for 53BP1 ADP-ribosylation not identified
    • Modification site(s) on 53BP1 not mapped
    • Whether NUDT16 dePARylation of 53BP1 is constitutive or damage-induced not fully resolved
  8. 2021 Medium

    Discovery that NUDT16 mRNA is silenced by small CAG repeat RNAs via RISC-dependent heteroduplex formation provided a disease-relevant mechanism linking polyglutamine expansion disorders to NUDT16-dependent genomic instability.

    Evidence RNA-seq; CAG-CUG heteroduplex formation assay; RISC-mediated silencing in cell and mouse polyglutamine models; pharmacological rescue with DB213

    PMID:33947817

    Open questions at the time
    • Extent of NUDT16 downregulation across different polyglutamine diseases and patient tissues not quantified
    • Contribution of NUDT16 loss versus other targets of sCAG silencing not separated
    • Independent replication in a distinct disease model not yet reported
  9. 2024 High

    Expansion of the dePARylation substrate repertoire to CtIP, NBS1, and SETD3 established NUDT16 as a general stabilizer of homologous recombination factors: in each case, NUDT16 removes ADP-ribosylation to block RNF146- or CHFR-mediated ubiquitin-proteasome degradation, enabling DSB recruitment, HR repair, and replication fork protection.

    Evidence Loss-of-function and catalytic mutant studies; Co-IP; ubiquitination assays; HR repair and DNA fiber assays; epistasis experiments (NUDT16-SETD3 double knockdown); DSB foci analysis

    PMID:38272222 PMID:38324469 PMID:39438373

    Open questions at the time
    • Specific ADP-ribosylation sites on CtIP and NBS1 not mapped
    • Identity of the PARPs that modify each substrate not established
    • Whether NUDT16 dePARylation is selective for specific PAR chain lengths or linkages unknown
  10. 2025 Medium

    Identification of HMGA1 Glu50 as a specific NUDT16 dePARylation site extended the pathway to replication fork protection via FANCD2 recruitment, and demonstrated that NUDT16-HMGA1 axis inhibition sensitizes tumors to chemotherapy and irradiation.

    Evidence Co-IP; dePARylation and ubiquitination assays; DNA fiber assay for fork protection; Glu50 site-directed mutagenesis; cell sensitivity assays

    PMID:40288645

    Open questions at the time
    • Therapeutic window for targeting the NUDT16-HMGA1 axis in vivo not established
    • Whether NUDT16 dePARylation of HMGA1 is CHFR-specific or involves additional E3 ligases unknown
    • Single-lab finding awaiting independent confirmation

Open questions

Synthesis pass · forward-looking unresolved questions
  • A unified understanding of how NUDT16 partitions among its three enzymatic activities (inosine diphosphatase, RNA decapping, dePARylase) in vivo, and which activity predominates under different cellular conditions, remains to be established.
  • No separation-of-function mutant that isolates one activity from the others has been characterized in vivo
  • No whole-organism knockout model has been reported
  • The full complement of dePARylation substrates and the PARPs that generate the modifications are not identified

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0016787 hydrolase activity 6 GO:0140096 catalytic activity, acting on a protein 6 GO:0140098 catalytic activity, acting on RNA 3
Localization
GO:0005634 nucleus 1 GO:0005730 nucleolus 1
Pathway
R-HSA-73894 DNA Repair 5 R-HSA-1430728 Metabolism 3 R-HSA-8953854 Metabolism of RNA 3
Partners
CHFRHMGA1NBNRBBP8RNF146SETD3TP53BP1

Evidence

Reading pass · 13 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2010 NUDT16 is a (deoxy)inosine diphosphatase that hydrolyzes purine nucleoside diphosphates to the corresponding nucleoside monophosphates; highest catalytic efficiency (kcat/Km) was for IDP and dIDP, with moderate activity on (d)ITP. NUDT16 is localized primarily in the nucleus and nucleolus, and its knockdown in HeLa cells caused S-phase cell cycle arrest, reduced proliferation, increased single-strand breaks in nuclear DNA, and elevated inosine in RNA. Recombinant protein enzymatic assay with 29 substrates; siRNA knockdown with cell cycle, comet assay, and RNA analysis; immunofluorescence localization Nucleic acids research High 20385596
2010 NUDT16 and ITPA play a dual protective role by eliminating dIDP/IDP and dITP/ITP from nucleotide pools in mammals. Immortalized Itpa−/− MEFs upregulate Nudt16 to compensate, and siRNA knockdown of Nudt16 in these cells reproduces ITPA-deficient phenotypes (prolonged doubling time, chromosome abnormalities, single-strand breaks in DNA). Knockout mouse embryonic fibroblasts, siRNA knockdown, chromosome aberration assay, comet assay, RT-PCR/Western blot for Nudt16 expression Nucleic acids research High 20081199
2007 X29 (Xenopus Nudt16) and its human homolog H29K (Nudt16) are metal-dependent nuclear decapping proteins that remove m7G and m2,2,7G caps from RNAs; in Mg2+ they specifically decap U8 snoRNA, while Mn2+ or Co2+ broadens substrate specificity to all RNAs. Crystal structure of X29 guided mutagenesis of active-site metal-coordinating residues confirming their essential roles. In vitro decapping assay with purified proteins; X-ray crystal structure; structure-based active site mutagenesis The Journal of biological chemistry High 17567574
2008 Evolutionary conservation of Nudt16 as a nuclear RNA-binding, RNA-decapping enzyme was demonstrated across metazoans; the insect ortholog was shown by in vitro analysis to bind RNA and hydrolyze the m7G cap from the 5' end of RNAs, indicating functional conservation. The closely related paralog Syndesmos (arising from a tetrapod gene duplication) retains RNA-binding but is inactive for decapping. In vitro RNA-binding and decapping assay with purified insect Nudt16 ortholog; phylogenetic analysis Nucleic acids research Medium 18820299
2015 Crystal structure of human NUDT16 in apo-form and in complex with its product inosine monophosphate (IMP) revealed a dimeric structure forming a positively charged trench for substrate binding. Structural and enzymatic analyses showed that substituents at positions 2 and 6 of the purine ring determine substrate selectivity, and IMP product inhibition regulates the catalytic cycle primarily at the product-release step. X-ray crystallography; enzymatic kinetics; biophysical binding studies PloS one High 26121039
2018 Human Nudt16 hydrolyzes dinucleotide cap analogs and short capped oligonucleotides; substrates with guanine as the first transcribed nucleotide are more susceptible than adenine-containing counterparts, and unmethylated caps (GpppG, ApppG) are hydrolyzed more efficiently than m7GpppG caps. In vitro enzymatic screening assay with purified hNudt16 against panel of dinucleotide cap analogs and capped oligonucleotides RNA (New York, N.Y.) Medium 29483298
2019 Human NudT16 (HsNudT16) acts as a phosphodiesterase that processes free and protein-conjugated ADP-ribose into phosphoribose. X-ray crystal structures of HsNudT16 in complex with monomeric and dimeric ADP-ribose identified the active site; rational design of mutants (Δ17, F36A, F61S) widening the active site reduced activity on free ADP-ribose but improved catalytic efficiency for protein-conjugated poly(ADP-ribose). X-ray crystallography with ADP-ribose complexes; rational mutagenesis; enzymatic activity assays on free and protein-conjugated ADP-ribose Scientific reports High 30976021
2020 NUDT16 regulates 53BP1 stability by removing ADP-ribosylation from 53BP1. ADP-ribosylated 53BP1 is recognized by the PAR-binding E3 ubiquitin ligase RNF146, leading to its polyubiquitination and degradation. In response to DNA damage, NUDT16 catalytic activity is required for 53BP1 de-ADP-ribosylation, protein stability, and recruitment to DSBs; a catalytically inactive NUDT16 mutant enhanced 53BP1 Tudor domain binding to TIRR and impaired H4K20me2 recognition. Co-IP; catalytically inactive mutant overexpression; ubiquitination assay; IR-induced foci analysis; cell survival assay Cancer research High 31911551
2021 Small CAG (sCAG) RNAs generated from expanded CAG transcripts hybridize with CUG-containing NUDT16 mRNA to form a CAG-CUG RNA heteroduplex, triggering RISC-dependent silencing of NUDT16, which leads to misincorporation of damaging nucleotides into DNA, DNA damage, and apoptosis in polyglutamine disease models. RNA-seq; RNA heteroduplex formation assay; RISC-mediated silencing in cell and mouse models; pharmacological rescue with DB213 Proceedings of the National Academy of Sciences of the United States of America Medium 33947817
2024 NUDT16 hydrolase activity removes ADP-ribosylation from CtIP, preventing its RNF146-mediated ubiquitination and degradation; ADP-ribosylation of CtIP is required for its interaction with NUDT16, its localization at DSBs, and homologous recombination (HR) repair. NUDT16 itself can also be ADP-ribosylated, and ADP-ribosylated NUDT16 is critical for CtIP stability and recruitment to DSBs. NUDT16 loss-of-function; catalytically inactive NUDT16 mutant; Co-IP; ubiquitination assay; HR repair assay; DSB recruitment foci Nucleic acids research High 38324469
2024 NUDT16 removes ADP-ribosylation from NBS1, preventing RNF146-mediated ubiquitination and degradation of NBS1. Loss of NUDT16 reduces NBS1 protein levels; ADP-ribosylation of NBS1 is required for its localization at DSBs and HR repair. The NUDT16–NBS1 interaction is regulated in response to DNA damage. NUDT16 knockdown; Co-IP; ubiquitination assay; DSB foci analysis; HR repair assay Molecular and cellular biochemistry Medium 39438373
2024 NUDT16 acts as a dePARylase for SETD3: it removes PARylation from SETD3, preventing CHFR E3-ligase-mediated ubiquitination and degradation. SETD3 associates with BRCA2 and promotes its recruitment to stalled replication forks and DSBs; depletion of SETD3 in NUDT16-deficient cells did not further sensitize cells to IR, placing NUDT16-SETD3 in the same pathway for replication stress resolution and HR repair. NUDT16 depletion; Co-IP; ubiquitination assay; replication fork protection assay; epistasis by double knockdown; IR sensitivity The Journal of biological chemistry Medium 38272222
2025 NUDT16 removes PARylation from HMGA1 at Glu50, suppressing CHFR-mediated ubiquitin-proteasome degradation of HMGA1, thereby stabilizing HMGA1 protein. HMGA1 in turn recruits FANCD2 to promote DNA replication and protect stalled replication forks from degradation. NUDT16-HMGA1 inhibition enhances tumor sensitivity to chemotherapy and irradiation. Co-IP; dePARylation assay; ubiquitination assay; replication fork protection (DNA fiber assay); site-directed mutation at Glu50; cell sensitivity assay The Journal of biological chemistry Medium 40288645

Source papers

Stage 0 corpus · 15 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2010 NUDT16 and ITPA play a dual protective role in maintaining chromosome stability and cell growth by eliminating dIDP/IDP and dITP/ITP from nucleotide pools in mammals. Nucleic acids research 55 20081199
2010 NUDT16 is a (deoxy)inosine diphosphatase, and its deficiency induces accumulation of single-strand breaks in nuclear DNA and growth arrest. Nucleic acids research 52 20385596
2008 Evolutionary conservation supports ancient origin for Nudt16, a nuclear-localized, RNA-binding, RNA-decapping enzyme. Nucleic acids research 34 18820299
2020 Nudix Hydrolase NUDT16 Regulates 53BP1 Protein by Reversing 53BP1 ADP-Ribosylation. Cancer research 33 31911551
2021 CAG RNAs induce DNA damage and apoptosis by silencing NUDT16 expression in polyglutamine degeneration. Proceedings of the National Academy of Sciences of the United States of America 26 33947817
2007 Metal determines efficiency and substrate specificity of the nuclear NUDIX decapping proteins X29 and H29K (Nudt16). The Journal of biological chemistry 26 17567574
2015 Structural Basis for the Specificity of Human NUDT16 and Its Regulation by Inosine Monophosphate. PloS one 22 26121039
2019 Structural analyses of NudT16-ADP-ribose complexes direct rational design of mutants with improved processing of poly(ADP-ribosyl)ated proteins. Scientific reports 17 30976021
2018 Hydrolytic activity of human Nudt16 enzyme on dinucleotide cap analogs and short capped oligonucleotides. RNA (New York, N.Y.) 17 29483298
2024 NUDT16 regulates CtIP PARylation to dictate homologous recombination repair. Nucleic acids research 10 38324469
2022 Transcriptomic profile investigations highlight a putative role for NUDT16 in sepsis. Journal of cellular and molecular medicine 6 35174610
2024 The dePARylase NUDT16 promotes radiation resistance of cancer cells by blocking SETD3 for degradation via reversing its ADP-ribosylation. The Journal of biological chemistry 4 38272222
2025 NUDT16 enhances the resistance of cancer cells to DNA-damaging agents by regulating replication fork stability via reversing HMGA1 ADP-ribosylation. The Journal of biological chemistry 3 40288645
2024 NBS1 dePARylation by NUDT16 is critical for DNA double-strand break repair. Molecular and cellular biochemistry 2 39438373
2008 Crystallization and crystallographic analysis of human NUDT16. Acta crystallographica. Section F, Structural biology and crystallization communications 1 18607096