NAE1

NAE1 (APP-BP1) is the regulatory subunit of the heterodimeric NEDD8-activating E1 enzyme, partnering with the catalytic subunit UBA3 to initiate the neddylation cascade that activates cullin-RING ubiquitin ligases and drives cell cycle progression through the S-M checkpoint (PMID:14690597, PMID:10722740). The NAE1-UBA3 heterodimer adenylates NEDD8 in a pseudo-ordered ATP-leading mechanism, forms a NEDD8-adenylate/UBA3-thioester intermediate, and transfers NEDD8 to the E2 enzyme UBE2M/Ubc12; selectivity for NEDD8 over ubiquitin is enforced both by a conserved arginine selectivity gate in the E1 and by Ala72 of NEDD8 (PMID:14690597, PMID:12740388). Within this complex NAE1 acts principally as a scaffold/regulatory subunit—UBA3 carries the catalytic activity while NAE1 accelerates the activation reaction (PMID:29973603). Through this neddylation activity NAE1 controls a broad range of physiological processes: CD8+ T cell activation, proliferation, survival, and mitochondrial function downstream of NFATc1/TCR signaling (PMID:40030035); meiotic recombination and double-strand break repair during spermatogenesis (PMID:40083933); neuromuscular junction formation via the rapsyn E3 neddylation ligase, which stabilizes acetylcholine receptors (PMID:40659529); and cardiac hypertrophy, where K238 crotonylation of NAE1 promotes neddylation and stabilization of gelsolin to drive cytoskeletal remodeling (PMID:39229723). NAE1 abundance is itself regulated by TRIP12-mediated polyubiquitination and proteasomal degradation of the NAE1 monomer (PMID:18627766). Beyond NEDD8, NAE1-UBA3 also functions as the E1 for the URM1 urmylation pathway (PMID:42056084). NAE1 was originally identified through direct binding to the cytoplasmic domain of the amyloid precursor protein (APP), and functions in neuronal apoptosis signaling and Aβ processing through interactions with APP, ASPP2, and Presenilin-1 (PMID:8626687, PMID:14557245, PMID:12694406, PMID:17286867). Bi-allelic loss-of-function variants in NAE1 cause a human disease characterized by impaired neddylation, immune dysfunction, and skeletal transcriptional defects (PMID:36608681).

1. 1996 Medium

   Established the protein now called NAE1 as a direct binding partner of the amyloid precursor protein, linking it to APP biology and noting its homology to the ubiquitin-activating E1 enzyme.

   Evidence Yeast two-hybrid screen, cDNA cloning, and sequence homology analysis

   PMID:8626687

   Open questions at the time

   • Enzymatic activity not yet demonstrated
   • Functional consequence of APP binding undefined
   • Single yeast two-hybrid interaction screen
2. 2000 High

   Defined NAE1 as a UBA3-binding cell cycle regulator, mapping the UBA3 interaction site and placing NAE1 in the neddylation E1/E2 cascade driving the S-M checkpoint.

   Evidence Co-IP, genetic rescue of the ts41 hamster cell cycle mutant, dominant-negative and overexpression experiments in mammalian cells and neurons

   PMID:10722740

   Open questions at the time

   • Catalytic mechanism of NEDD8 activation not resolved
   • Substrate ligases downstream not enumerated
3. 2003 High

   Resolved the structural and kinetic mechanism of NEDD8 activation and the molecular basis of NEDD8-versus-ubiquitin selectivity, establishing how NAE1-UBA3 charges the E2 Ubc12.

   Evidence X-ray crystallography of the APPBP1-UBA3-NEDD8-ATP quaternary complex, in vitro enzyme kinetics with radiolabeled substrates, transthiolation assays, and substrate mutagenesis

   PMID:12740388 PMID:14690597

   Open questions at the time

   • Did not assign relative catalytic contributions of NAE1 versus UBA3 subunits
   • Regulation of the enzyme in cells not addressed
4. 2003 High

   Connected NAE1 neddylation activity to neuronal apoptosis signaling downstream of APP and identified ASPP2 as a negative regulator that suppresses Cullin-1 neddylation.

   Evidence Co-IP, lipid raft fractionation, dominant-negative Ubc12 epistasis, and siRNA in primary neurons (APP/C31 binding); reciprocal endogenous Co-IP and ts41/neddylation/apoptosis assays (ASPP2)

   PMID:12694406 PMID:14557245

   Open questions at the time

   • Mechanism by which ASPP2 inhibits the E1 not defined
   • Physiological relevance of neuronal apoptosis pathway in vivo unresolved
5. 2006 Medium

   Confirmed via the Drosophila ortholog that APP-family proteins antagonistically regulate the NEDD8 conjugation pathway and that NAE1 loss triggers apoptosis, generalizing the APP-neddylation link.

   Evidence Genetic loss-of-function, yeast two-hybrid/Co-IP, NEDD8 conjugation assays, and overexpression epistasis in Drosophila imaginal discs

   PMID:16628230

   Open questions at the time

   • Ortholog data may not fully translate to mammalian context
   • Molecular mechanism of APPL antagonism unresolved
6. 2007 Medium

   Implicated NAE1 in Aβ processing by showing it associates with Presenilin-1/γ-secretase and negatively regulates Aβ42 production.

   Evidence Co-IP from rat brain, glycerol gradient fractionation, siRNA knockdown, and overexpression in primary neurons

   PMID:17286867

   Open questions at the time

   • Whether regulation of Aβ requires neddylation catalytic activity unclear
   • Single-lab study without in vivo confirmation
7. 2008 Medium

   Identified TRIP12 as an E3 ligase that selectively degrades the NAE1 monomer, revealing post-translational control of cellular neddylation capacity.

   Evidence Yeast two-hybrid, Co-IP, in vitro ubiquitination assay, and siRNA knockdown with CUL1 neddylation readout

   PMID:18627766

   Open questions at the time

   • Physiological conditions triggering NAE1 turnover not defined
   • In vivo relevance not established
8. 2018 Medium

   Determined that catalytic activity resides in UBA3 while NAE1 serves as a scaffold/regulatory subunit accelerating the reaction, clarifying the division of labor within the E1 heterodimer.

   Evidence Quantitative FRET, computational electrostatic analysis, and in vitro reconstitution with individual subunits

   PMID:29973603

   Open questions at the time

   • Single-lab in vitro result without independent replication
   • Structural basis of NAE1's rate acceleration not detailed
9. 2023 Medium

   Established NAE1 as a human disease gene, showing bi-allelic loss-of-function impairs neddylation with immune and skeletal transcriptional consequences.

   Evidence Patient fibroblasts with bi-allelic variants, proteasomal stress assay, NF-κB translocation assay, lymphocyte stimulation, and transcriptomics

   PMID:36608681

   Open questions at the time

   • Single cohort study
   • Mechanistic link from neddylation loss to RUNX2/SOX9 downregulation not resolved
10. 2024 High

    Revealed that crotonylation of NAE1 at K238 directs neddylation toward gelsolin, defining a PTM-controlled substrate-stabilization mechanism in cardiac hypertrophy.

    Evidence Quantitative crotonylomics, NAE1 K238R/K238Q knock-in mice, Co-IP/MS, in vitro neddylation, and actin-severing assays

    PMID:39229723

    Open questions at the time

    • Crotonyltransferase/decrotonylase enzymes acting on K238 not identified
    • Whether K238 crotonylation governs other substrates unknown
11. 2025 High

    Demonstrated NAE1-mediated neddylation is essential across diverse physiological contexts—CD8+ T cell antitumor function, meiotic recombination, and neuromuscular junction formation—each via distinct downstream effectors.

    Evidence Conditional/germ-cell/muscle-specific Nae1 knockouts, NFATc1 binding assays, single-cell transcriptomics, ubiquitinomics, rapsyn-C366A epistasis, and AChR clustering/NMJ morphology assays; gastric cancer TFR1 neddylation and ferroptosis assays

    PMID:40030035 PMID:40083933 PMID:40659529 PMID:41135076

    Open questions at the time

    • Full substrate repertoire in each tissue incompletely mapped
    • TFR1 ferroptosis study is single-lab Medium confidence
12. 2026 High

    Expanded NAE1's enzymatic scope beyond NEDD8 by showing NAE1-UBA3 also serves as the E1 for the URM1 urmylation pathway under normal and oxidative stress conditions.

    Evidence Activity-based URM1 probe, proteomic characterization, cell-based validation, and pharmacological inhibition with pevonedistat

    PMID:42056084

    Open questions at the time

    • Relative cellular flux through NEDD8 versus URM1 pathways unquantified
    • Determinants of NAE1 substrate selection between modifiers undefined

• How NAE1 substrate/effector specificity is tuned across tissues and between the NEDD8 and URM1 pathways, and how its post-translational regulation integrates with disease, remains open.
• No unified model linking PTM state, partner availability, and substrate choice
• Mechanism routing NAE1 between neddylation and urmylation unknown