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Comprehensive Research Report: DCAF10 (DDB1- and CUL4-associated factor 10)

Gene Identity and Protein Overview

DCAF10 (also known as WDR32) is a human gene encoding a DDB1- and CUL4-associated factor 10 protein (UniProt ID: Q5QP82). The protein belongs to the WD repeat DCAF10 family and contains characteristic WD40 repeat domains that fold into a β-propeller structure (kremer2026cul4addb1dcaf10isan pages 1-2). DCAF10 is evolutionarily conserved, with homologs identified across mammalian species (mistry2020expressionprofilingof pages 1-2).

Primary Molecular Function: Substrate Receptor in CRL4 E3 Ubiquitin Ligase Complexes

DCAF10 functions as a substrate receptor (specificity factor) for Cullin 4-RING E3 ubiquitin ligase (CRL4) complexes, comprising CUL4A (or CUL4B), DDB1, DCAF10, and the catalytic subunit RBX1 (kremer2026cul4addb1dcaf10isan pages 1-2, zemke2023adenoviruse1abinding pages 1-2, zemke2023adenoviruse1abinding pages 5-7). As a substrate receptor, DCAF10 recruits specific target proteins to the E3 ligase complex, facilitating their ubiquitination and subsequent proteasomal degradation (kremer2026cul4addb1dcaf10isan pages 1-2, kremer2026cul4addb1dcaf10isan pages 6-8). The interaction between DCAF10 and the CRL4 scaffold requires DDB1 as an adaptor protein; DCAF10 binds DDB1 through conserved WDxR motifs within its WD40 repeats (zemke2023adenoviruse1abinding pages 5-7).

Substrate Recognition Mechanism: The Ac-Gly/N-Degron Pathway

Recent breakthrough research by Kremer et al. (2026) identified DCAF10 as an N-recognin for proteins bearing N-terminally acetylated glycine (Ac-Gly) residues, particularly those that normally undergo N-myristoylation (kremer2026cul4addb1dcaf10isan pages 1-2). This discovery defines a novel N-degron pathway termed the "Ac-Gly/MO (myristoylation-omitted) N-degron pathway" (kremer2026cul4addb1dcaf10isan pages 1-2, kremer2026cul4addb1dcaf10isan pages 8-9).

Structural Basis of Recognition:
AlphaFold 3 predictions combined with biochemical validation demonstrate that DCAF10 recognizes Ac-Gly through a deep pocket within its β-propeller structure (kremer2026cul4addb1dcaf10isan pages 2-3, kremer2026cul4addb1dcaf10isan pages 3-4). The acetyl group of the N-terminal glycine is critical for high-affinity binding; specific DCAF10 residues including Phe172, Asn215, Lys257, Ile475, and Glu477 interact with the Ac-Gly moiety and are highly conserved across species (kremer2026cul4addb1dcaf10isan pages 3-4). Additional sequence features influence binding affinity: hydrophobic residues at positions 3-4 of the substrate strengthen interaction, while acidic residues at positions 5-7 weaken binding (kremer2026cul4addb1dcaf10isan pages 2-3, kremer2026cul4addb1dcaf10isan pages 3-4).

Substrate Specificity:
The recognition motif is not simply Ac-Gly but includes downstream residues, with a preference for serine at position 6 and lysine at position 7 (kremer2026cul4addb1dcaf10isan pages 2-3). This selectivity distinguishes proteins targeted by DCAF10 from those that are robustly acetylated but not degraded, such as THOC7, which contains acidic residues (Asp6-Asp7-Glu8) that repel DCAF10's negatively charged pocket environment (kremer2026cul4addb1dcaf10isan pages 2-3, kremer2026cul4addb1dcaf10isan pages 3-4).

Identified Protein Substrates

Table: This table summarizes the best-supported DCAF10 substrates and substrate-like targets, highlighting how DCAF10 recognizes them, what post-translational states are required, and the biological contexts in which these interactions matter. It is useful for separating direct DCAF10 substrates from cases, such as IRF3, where the evidence currently supports indirect regulation.

Src Family Kinases (SFKs)

The most extensively characterized DCAF10 substrates are Src family kinases—Lyn, Fyn, and Src—which normally require N-myristoylation for membrane localization and proper function (kremer2026cul4addb1dcaf10isan pages 1-2, kremer2026cul4addb1dcaf10isan pages 6-8, kremer2026cul4addb1dcaf10isan pages 8-9). When N-myristoylation by NMT1/2 is impaired or absent, these proteins can be alternatively N-terminally acetylated by NatA, generating an Ac-Gly N-terminus that DCAF10 recognizes (kremer2026cul4addb1dcaf10isan pages 1-2, kremer2026cul4addb1dcaf10isan pages 5-6, kremer2026cul4addb1dcaf10isan pages 4-5).

Experimental Evidence:
- Peptide pull-down assays using mass spectrometry identified DCAF10 as the primary binder to acetylated Lyn, Fyn, and Src N-terminal peptides (kremer2026cul4addb1dcaf10isan pages 2-3).
- In vitro ubiquitination assays with reconstituted CUL4A-DDB1-DCAF10 complexes directly ubiquitinated immunoprecipitated Lyn, Fyn, and Src proteins (kremer2026cul4addb1dcaf10isan pages 6-8).
- DCAF10 knockdown increased Lyn, Fyn, and Src protein levels, particularly when NMT1/2 was depleted, without affecting mRNA levels (kremer2026cul4addb1dcaf10isan pages 5-6, kremer2026cul4addb1dcaf10isan pages 4-5).
- Cycloheximide chase experiments showed accelerated degradation of SFKs upon NMT1/2 depletion, which was reversed by proteasome inhibitor MG132 (kremer2026cul4addb1dcaf10isan pages 4-5).

Functional Context:
DCAF10-mediated degradation acts complementarily to the CUL2-elonginB/C-ZYG11B/ZER1 Gly/N-degron pathway, which recognizes free (non-acetylated) N-terminal glycine (kremer2026cul4addb1dcaf10isan pages 6-8, kremer2026cul4addb1dcaf10isan pages 8-9). This dual surveillance system ensures proteostasis control over SFKs that fail proper N-myristoylation. Mutation of the N-terminal glycine to alanine (G2A, which can be acetylated) or proline (G2P, which resists acetylation) confirmed that both glycine identity and acetylation are required for DCAF10-dependent degradation (kremer2026cul4addb1dcaf10isan pages 8-9).

RUVBL1/2 AAA+ ATPases

RUVBL1 and RUVBL2 (also known as Pontin and Reptin) are essential AAA+ ATPases that function as subunits of multiple protein complexes, including HSP90 co-chaperones and chromatin-modifying complexes (zemke2023adenoviruse1abinding pages 1-2, zemke2023adenoviruse1abinding pages 7-11). These proteins are required for assembly of multiprotein machines involved in DNA repair, transcriptional responses to interferon, and metabolic stress responses (zemke2023adenoviruse1abinding pages 7-11).

Evidence for DCAF10 Targeting:
Co-immunoprecipitation studies identified RUVBL1/2 as DCAF10-interacting proteins (zemke2023adenoviruse1abinding pages 7-11). In the context of adenovirus infection, viral protein E1A binds DCAF10 and promotes assembly of a functional CRL4 E3 ligase complex that targets RUVBL1/2 for degradation (zemke2023adenoviruse1abinding pages 1-2, zemke2023adenoviruse1abinding pages 5-7, zemke2023adenoviruse1abinding pages 7-11). Depletion of RUVBL1/2 prevented IRF3 stabilization induced by inhibition of EP300/CREBBP acetyltransferases, positioning RUVBL1/2 as critical mediators of innate immune responses that DCAF10 can negatively regulate (zemke2023adenoviruse1abinding pages 7-11).

ALOX15B (Arachidonate 15-Lipoxygenase)

In KRAS-mutant pancreatic ductal adenocarcinoma (PDAC), DCAF10 mediates degradation of ALOX15B, a lipoxygenase enzyme involved in ferroptosis induction (li2025krasabhd17calox15baxispromotes pages 6-8).

Mechanistic Details:
ALOX15B degradation by DCAF10 requires prior depalmitoylation by the enzyme ABHD17C (li2025krasabhd17calox15baxispromotes pages 6-8). When ALOX15B is S-palmitoylated at Cys106, it localizes to the membrane and is protected from degradation. ABHD17C-mediated depalmitoylation promotes ALOX15B translocation to the cytoplasm and enhances its interaction with DCAF10 (li2025krasabhd17calox15baxispromotes pages 6-8). AlphaFold 3 analysis predicted that the N-terminal region of ALOX15B (residues N183-R221) docks with the central WD40 repeat region of DCAF10 (residues E188-S318), and this interaction was confirmed experimentally with a binding affinity (KD) of 6.96 nM (li2025krasabhd17calox15baxispromotes pages 6-8). The CUL4/DDB1/DCAF10 complex ubiquitinates ALOX15B at lysine 175 (K175), promoting its proteasomal degradation (li2025krasabhd17calox15baxispromotes pages 6-8).

IRF3 (Indirect Regulation)

Interferon regulatory factor 3 (IRF3) is stabilized when DCAF10 is depleted, leading to increased expression of interferon-stimulated genes (ISGs) (zemke2023adenoviruse1abinding pages 2-5, zemke2023adenoviruse1abinding pages 7-11). However, IRF3 was not detected in direct DCAF10 immunoprecipitates, suggesting indirect regulation (zemke2023adenoviruse1abinding pages 5-7). Current evidence indicates that DCAF10 controls IRF3 levels indirectly through degradation of RUVBL1/2, which are required for IRF3 stabilization (zemke2023adenoviruse1abinding pages 7-11).

Subcellular Localization

DCAF10 functions in both nuclear and cytoplasmic compartments as part of the ubiquitin-proteasome system (zemke2023adenoviruse1abinding pages 1-2, zemke2023adenoviruse1abinding pages 5-7). The protein interacts with nuclear proteins including transcription factors and chromatin-associated proteins, while also regulating cytoplasmic substrates such as Src family kinases (kremer2026cul4addb1dcaf10isan pages 1-2, zemke2023adenoviruse1abinding pages 1-2). DCAF10's ability to assemble with CUL4A/B-DDB1 allows it to function wherever these scaffold proteins localize, providing flexibility to access substrates in different cellular compartments (zemke2023adenoviruse1abinding pages 5-7).

Biological Processes and Signaling Pathways

Table: This table summarizes the main biological processes and pathways currently linked to human DCAF10, emphasizing its role as a CRL4 substrate receptor in proteostasis, signaling, immunity, reproduction, viral infection, and pancreatic cancer.

N-Degron Pathway and Protein Quality Control

DCAF10 defines a novel branch of the N-degron pathway that monitors N-terminal co-translational modifications (kremer2026cul4addb1dcaf10isan pages 1-2). When proteins fail to receive their proper N-terminal modification—particularly myristoylation—they expose alternative N-termini that serve as degradation signals (degrons). The Ac-Gly/MO pathway mediated by DCAF10 complements the established Gly/N-degron pathway (mediated by ZYG11B/ZER1), creating redundant quality control over N-terminally misprocessed proteins (kremer2026cul4addb1dcaf10isan pages 6-8, kremer2026cul4addb1dcaf10isan pages 8-9).

Innate Immunity and Viral Pathogenesis

DCAF10 plays a complex role in innate immune regulation. Under normal conditions or during viral infection, DCAF10-mediated degradation of RUVBL1/2 suppresses IRF3 accumulation and limits expression of interferon-stimulated genes (ISGs) (zemke2023adenoviruse1abinding pages 2-5, zemke2023adenoviruse1abinding pages 7-11). Human adenovirus exploits this mechanism: the viral E1A protein binds DCAF10 and promotes assembly of an active CRL4 complex that degrades RUVBL1/2, thereby blunting antiviral responses and facilitating viral replication (zemke2023adenoviruse1abinding pages 1-2, zemke2023adenoviruse1abinding pages 5-7, zemke2023adenoviruse1abinding pages 7-11).

Cell Signaling via Src Family Kinase Regulation

Src family kinases are critical signaling molecules implicated in cancer progression and cellular transformation (kremer2026cul4addb1dcaf10isan pages 1-2). By degrading improperly N-myristoylated SFKs, DCAF10 may serve as a tumor suppressor mechanism that limits accumulation of delocalized, potentially oncogenic kinases (kremer2026cul4addb1dcaf10isan pages 6-8, kremer2026cul4addb1dcaf10isan pages 8-9). Conversely, dysregulation of DCAF10 could contribute to altered SFK signaling in disease contexts.

Testicular Development and Spermatogenesis

Expression profiling studies demonstrate remarkably high DCAF10 expression in testis compared to other tissues. Analysis of mouse and human RNA-seq datasets revealed that approximately 74-83% of DCAF subfamily genes, including DCAF10, are predominantly or specifically expressed in testis (mistry2020expressionprofilingof pages 5-9, mistry2020expressionprofilingof pages 2-4). DCAF10 shows highest transcript levels in testis with very low expression in other tissues, suggesting a specialized role in spermatogenesis (mistry2020expressionprofilingof pages 5-9). During testicular development, DCAF10 expression is upregulated during post-natal stages from neonatal day 20 through adulthood, coinciding with active spermatogenesis (mistry2020expressionprofilingof pages 5-9). This pattern implicates DCAF10 in germ cell development and suggests that ubiquitin-dependent protein remodeling by DCAF10 may be critical for male reproductive biology (mistry2020expressionprofilingof pages 1-2).

Cancer Biology: Pancreatic Cancer and Ferroptosis Evasion

In KRAS-mutant pancreatic ductal adenocarcinoma, the KRAS/ERK1 signaling axis promotes ABHD17C-mediated depalmitoylation of ALOX15B, facilitating its DCAF10-dependent degradation (li2025krasabhd17calox15baxispromotes pages 6-8). Since ALOX15B promotes ferroptosis (a form of regulated cell death driven by lipid peroxidation), its degradation allows cancer cells to evade ferroptotic death, promoting tumor progression (li2025krasabhd17calox15baxispromotes pages 6-8). This mechanism highlights DCAF10 as a potential therapeutic target in KRAS-mutant PDAC.

Current Understanding and Recent Developments (2023-2026)

The most significant recent advance in DCAF10 biology is the identification of its role as an N-recognin for N-terminally acetylated glycine residues by Kremer et al. (2026, Nature Communications) (kremer2026cul4addb1dcaf10isan pages 1-2). This work employed peptide pull-downs, quantitative mass spectrometry, AlphaFold 3 structural predictions, and in vitro reconstitution of the CUL4A-DDB1-DCAF10 E3 ligase to demonstrate direct substrate recognition and ubiquitination (kremer2026cul4addb1dcaf10isan pages 2-3, kremer2026cul4addb1dcaf10isan pages 3-4, kremer2026cul4addb1dcaf10isan pages 1-2, kremer2026cul4addb1dcaf10isan pages 6-8, kremer2026cul4addb1dcaf10isan pages 8-9).

Zemke et al. (2023, Journal of Virology) revealed how adenovirus hijacks DCAF10 to manipulate host innate immunity, demonstrating that the viral E1A protein recruits DCAF10 into a CRL4 complex that degrades RUVBL1/2, thereby suppressing interferon responses (zemke2023adenoviruse1abinding pages 1-2, zemke2023adenoviruse1abinding pages 5-7, zemke2023adenoviruse1abinding pages 7-11).

Li et al. (2025, Advanced Science) identified DCAF10's role in KRAS-driven pancreatic cancer, showing that the KRAS/ABHD17C/ALOX15B axis promotes ferroptosis evasion through DCAF10-mediated ALOX15B degradation (li2025krasabhd17calox15baxispromotes pages 6-8).

Expert Perspectives and Analysis

The discovery of DCAF10 as an Ac-Gly N-recognin fundamentally expands our understanding of N-degron pathways. While N-terminal acetylation is generally considered a protective modification that prevents degradation, DCAF10 demonstrates that acetylation can also create context-dependent degrons (kremer2026cul4addb1dcaf10isan pages 1-2). This dual role emphasizes the complexity of N-terminal processing and its integration with proteostasis networks.

The complementary surveillance by DCAF10 (recognizing Ac-Gly) and ZYG11B/ZER1 (recognizing free Gly) ensures robust quality control over proteins that fail N-myristoylation, preventing accumulation of mislocalized proteins that could disrupt cellular functions (kremer2026cul4addb1dcaf10isan pages 6-8, kremer2026cul4addb1dcaf10isan pages 8-9). This redundancy underscores the importance of proper co-translational modification for cellular homeostasis.

The tissue-specific expression pattern of DCAF10, particularly its high abundance in testis, suggests that this protein may have specialized substrates in male germ cells that remain to be identified (mistry2020expressionprofilingof pages 5-9, mistry2020expressionprofilingof pages 2-4, mistry2020expressionprofilingof pages 1-2). Further research is needed to characterize DCAF10's specific role in spermatogenesis and whether defects in DCAF10 contribute to male infertility.

Summary

DCAF10 is a substrate receptor for CUL4A-DDB1 E3 ubiquitin ligase complexes that recognizes N-terminally acetylated glycine degrons, particularly on proteins that normally undergo N-myristoylation. Its primary substrates include Src family kinases (Lyn, Fyn, Src), RUVBL1/2 AAA+ ATPases, and ALOX15B. DCAF10 functions in protein quality control, innate immune regulation, cell signaling, testicular development, and is implicated in viral immune evasion and cancer progression. The protein localizes to both nuclear and cytoplasmic compartments and exhibits particularly high expression in testis. Recent structural and biochemical studies have defined the molecular basis of DCAF10's substrate recognition through a β-propeller pocket that binds Ac-Gly with sequence-specific constraints. These advances position DCAF10 as a critical regulator of proteostasis and a potential therapeutic target in cancer and infectious diseases.

Key Citations:
- Kremer et al., 2026, Nature Communications, DOI: 10.1038/s41467-025-68074-9
- Zemke et al., 2023, Journal of Virology, DOI: 10.1128/jvi.00993-23
- Li et al., 2025, Advanced Science, DOI: 10.1002/advs.202504470
- Mistry et al., 2020, BMC Genomics, DOI: 10.1186/s12864-020-07016-9

References

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11. (li2025krasabhd17calox15baxispromotes pages 6-8): Man Li, Xuexin Yu, Yuanji Liu, Shuqin Ouyang, Long Wu, Xiaohong Chen, Huiqi Yu, Haoming Chen, Senmao Lian, Ziwen Li, Liyun Gong, Libing Song, and Jun Li. Kras/abhd17c/alox15b axis promotes pancreatic cancer progression via ferroptosis evasion. Advanced science, pages e04470, Jun 2025. URL: https://doi.org/10.1002/advs.202504470, doi:10.1002/advs.202504470. This article has 8 citations and is from a peer-reviewed journal.

12. (zemke2023adenoviruse1abinding pages 2-5): Nathan R. Zemke, Emily Hsu, William D. Barshop, Jihui Sha, James A. Wohlschlegel, and Arnold J. Berk. Adenovirus e1a binding to dcaf10 targets proteasomal degradation of ruvbl1/2 aaa+ atpases required for quaternary assembly of multiprotein machines, innate immunity, and responses to metabolic stress. Journal of Virology, Dec 2023. URL: https://doi.org/10.1128/jvi.00993-23, doi:10.1128/jvi.00993-23. This article has 5 citations and is from a domain leading peer-reviewed journal.

13. (kremer2026cul4addb1dcaf10isan pages 5-6): Nora Kremer, Franziska Mueller, Hang Nguyen, Louisa Schulz, Tanja Popp, Elena Artes, Julian Wolters, Michael Renner, Ingrid Vetter, Stefano Maffini, Maria S. Robles, Andrea Musacchio, and Tanja Bange. Cul4a-ddb1-dcaf10 is an n-recognin for n-terminally acetylated src kinases. Nature Communications, Jan 2026. URL: https://doi.org/10.1038/s41467-025-68074-9, doi:10.1038/s41467-025-68074-9. This article has 2 citations and is from a highest quality peer-reviewed journal.

14. (mistry2020expressionprofilingof pages 5-9): Bhavesh V. Mistry, Maha Alanazi, Hanae Fitwi, Olfat Al-Harazi, Mohamed Rajab, Abdullah Altorbag, Falah Almohanna, Dilek Colak, and Abdullah M. Assiri. Expression profiling of wd40 family genes including ddb1- and cul4- associated factor (dcaf) genes in mice and human suggests important regulatory roles in testicular development and spermatogenesis. BMC Genomics, Aug 2020. URL: https://doi.org/10.1186/s12864-020-07016-9, doi:10.1186/s12864-020-07016-9. This article has 32 citations and is from a peer-reviewed journal.

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Artifacts

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Citations

1. mistry2020expressionprofilingof pages 1-2
2. mistry2020expressionprofilingof pages 5-9
3. mistry2020expressionprofilingof pages 2-4
4. https://doi.org/10.1038/s41467-025-68074-9,
5. https://doi.org/10.1186/s12864-020-07016-9,
6. https://doi.org/10.1128/jvi.00993-23,
7. https://doi.org/10.1002/advs.202504470,

DCAF10 as an Ac-Gly N-Recognin: Evidence Assessment and Proteome-Wide Substrate Scope

Summary

DCAF10 (DDB1- and CUL4-Associated Factor 10) has been identified as a bona fide N-recognin that specifically binds N-terminally acetylated glycine (Ac-Gly) degrons and mediates ubiquitin-dependent proteasomal degradation of substrates through the CRL4 (CUL4A–DDB1–DCAF10) E3 ubiquitin ligase complex. This defines a novel branch of the N-degron pathway system — one that monitors the replacement of N-terminal myristoylation by acetylation and triggers degradation of the aberrantly modified protein. The primary evidence comes from a single peer-reviewed study by Kremer et al. (2026; PMID: 41484149), which demonstrated direct Ac-Gly peptide binding by DCAF10 via peptide pull-downs and mass spectrometry, reconstituted in vitro ubiquitination of N-terminally acetylated Src-family kinases (SFKs) by CUL4A–DDB1–DCAF10, and endogenous regulation of Lyn kinase through siRNA knockdown, CRISPR/Cas9 knockout, and degron mutagenesis rescue experiments.

A specific Ac-Gly N-degron recognin molecular function annotation for DCAF10 is justified based on the current peer-reviewed evidence. The data satisfy multiple criteria for N-recognin designation: (i) direct binding to the N-terminal acetylated glycine motif, (ii) reconstituted E3 ligase activity on Ac-Gly-bearing substrates, (iii) endogenous substrate validation with genetic loss-of-function, and (iv) degron mutagenesis demonstrating dependence on the N-terminal glycine residue. This goes well beyond mere CRL4 complex membership or generic substrate-adaptor activity. However, the proteome-wide breadth of this recognition activity remains experimentally uncharacterized. While the authors suggest the mechanism may extend to all ~200+ N-terminally myristoylated human proteins, validated endogenous substrates are currently limited to Src-family kinases (Lyn validated endogenously; Fyn and Src demonstrated in vitro). No proteome-wide degradomics screen has yet been published to define the full substrate repertoire of CRL4–DCAF10.

Prior literature established DCAF10 as a functional CRL4 substrate receptor but did not identify its intrinsic substrate specificity. Adenovirus E1A was shown to hijack DCAF10 to redirect CRL4 activity toward RUVBL1/2 (PMID: 37962355), and OTUD1 was found to stabilize DCAF10 to promote CUL4A-DDB1-mediated MCL1 degradation (PMID: 33898171). Neither study identified the endogenous degron recognized by DCAF10 itself. The Kremer et al. 2026 study thus represents the first identification of DCAF10's intrinsic substrate specificity, and no preprint or non-peer-reviewed evidence providing additional data was identified.

Key Findings

Finding 1: DCAF10 Directly Binds N-Terminally Acetylated Glycine Degrons and Functions as a CRL4 Substrate Receptor

Kremer et al. (2026) used an unbiased peptide pull-down approach coupled with mass spectrometry to identify DCAF10 as the specific cellular factor that binds Ac-Gly peptides derived from Src-family kinase N-termini. The authors report: "Using peptide pull-downs, mass spectrometry, and AlphaFold 3 predictions, we identify DCAF10 as the E3 ligase substrate receptor for alternatively N-terminally acetylated SFKs." The interaction was further supported by AlphaFold 3 structural predictions, which modeled the DCAF10–Ac-Gly interface. Critically, reconstituted CUL4A–DDB1–DCAF10 complexes ubiquitinated N-terminally acetylated SFKs in vitro — as stated: "In vitro, a CUL4A-DDB1-DCAF10 complex ubiquitinates N-terminally acetylated SFKs." — establishing that the complete E3 ligase activity depends on DCAF10 as the substrate-recruiting component. Degron mutagenesis — specifically, mutation of the N-terminal glycine residue — abolished DCAF10 recognition, confirming that the Ac-Gly motif is the minimal degron element required for binding.

This evidence satisfies the classical definition of an N-recognin: a ubiquitin ligase component that directly recognizes an N-terminal degron to initiate ubiquitin-dependent proteolysis. DCAF10 joins a growing family of N-recognins that includes UBR1/UBR2 (Arg/N-degron pathway), MARCHF6/Doa10 (Ac/N-degron pathway for Nt-acetylated Met, Ala, Val, Ser, Thr, Cys), and ZYG11B/ZER1 (Gly/N-degron pathway for unmodified Gly). DCAF10 is unique in recognizing acetylated glycine specifically — a modification state not targeted by ZYG11B/ZER1, which instead recognize unmodified (non-acetylated, non-myristoylated) N-terminal glycine.

Finding 2: Endogenous Lyn Kinase Is a Validated DCAF10 Substrate

The endogenous Src-family kinase Lyn was validated as a physiological DCAF10 substrate through multiple complementary genetic approaches. The study reports: "Combining siRNA-mediated knockdown and CRISPR/Cas9-mediated knockout of endogenous Lyn with inducible Lyn-GFP variants confirms that DCAF10 regulates SFK levels by recognizing an N-terminal acetylated glycine residue." Specifically:

• siRNA-mediated DCAF10 knockdown stabilized endogenous Lyn protein levels, demonstrating that DCAF10 is required for normal Lyn turnover.
• CRISPR/Cas9-mediated knockout of endogenous Lyn combined with inducible Lyn-GFP variants confirmed DCAF10-dependent regulation in a clean genetic background.
• Degron mutagenesis rescue experiments showed that Lyn-GFP variants with mutated N-terminal glycine residues were no longer subject to DCAF10-mediated degradation, establishing that the Ac-Gly degron is both necessary and sufficient for recognition.

In addition to Lyn, Fyn and Src were demonstrated as in vitro substrates of the CRL4–DCAF10 complex, showing that the recognition extends across multiple SFK family members. However, endogenous validation (knockdown/knockout stabilization) was reported only for Lyn.

Finding 3: DCAF10 Monitors the Myristoylation-to-Acetylation Switch as a Protein Quality Control Mechanism

The biological function of the CRL4–DCAF10 pathway is best understood as a protein quality control mechanism that monitors the fidelity of N-terminal myristoylation. Src-family kinases normally require co-translational N-myristoylation at Gly2 for proper membrane targeting and function. When myristoylation fails — due to NMT inhibition, competition for NMT access, or other perturbations — the exposed N-terminal glycine can instead be Nα-acetylated by N-terminal acetyltransferases (NATs). This Ac-Gly modification then serves as a degron recognized by CRL4–DCAF10, triggering ubiquitination and proteasomal degradation of the aberrantly modified protein.

Kremer et al. summarize this model: "Thus, we define a novel N-degron pathway that monitors replacement of myristoylation by acetylation and activates degradation of SFKs upon acetylation. This mechanism may extend to other N-terminally myristoylated proteins beyond SFKs."

This quality control circuit can be represented as:

Normal pathway:
  Met-Gly... → MetAP cleavage → Gly... → NMT myristoylation → Myr-Gly-protein
  → Membrane localization & function ✓

Degradation pathway (myristoylation failure + acetylation):
  Gly... → NAT acetylation → Ac-Gly-protein → CRL4-DCAF10 recognition
  → Ubiquitination → Proteasomal degradation ✗

Gly/N-degron pathway (myristoylation failure, no modification):
  Gly... (unmodified) → CRL2-ZYG11B/ZER1 recognition
  → Ubiquitination → Proteasomal degradation ✗

This model positions CRL4–DCAF10 as complementary to the previously characterized CRL2–ZYG11B/ZER1 Gly/N-degron pathway (PMID: 31273098), which monitors completely unmodified N-terminal glycine. Together, these two pathways provide comprehensive surveillance of N-terminal glycine modification status: ZYG11B/ZER1 targets bare Gly, while DCAF10 targets Ac-Gly. Both systems act as quality control checkpoints for the ~200+ human proteins that undergo N-terminal myristoylation.

Finding 4: Proteome-Wide Substrate Scope Remains Experimentally Undefined Beyond SFKs

While the mechanistic evidence for DCAF10 as an Ac-Gly N-recognin is strong, the proteome-wide breadth of its substrate recognition has not been experimentally defined. Kremer et al. explicitly state that "this mechanism may extend to other N-terminally myristoylated proteins beyond SFKs," but no proteome-wide screen has been published. Key open questions include:

The approximately 200+ human proteins known to be NMT substrates with N-terminal glycine (as documented in chemical proteomic surveys, e.g., PMID: 40887160) represent a large candidate pool. Whether DCAF10 recognizes all of these when they are alternatively acetylated, or whether additional sequence features (positions 2–6, structural context) are required for productive binding, is a critical unanswered question. This parallels the known extended specificity observed for ZYG11B/ZER1, which engage primarily the first four residues of Gly/N-degrons through armadillo repeat cavities (PMID: 34214466).

Finding 5: Prior DCAF10 Literature Confirms CRL4 Complex Membership but Not N-Degron Specificity

Three prior publications reference DCAF10 function, all confirming its role as a CRL4 substrate receptor but none identifying its intrinsic substrate specificity:

Zemke et al. showed that "Human respiratory adenoviruses counter this by assembling a CUL4-based ubiquitin ligase complex that polyubiquitinylates RUVBL1 and 2 inducing their proteasomal degradation" — confirming DCAF10 as a functional CRL4 substrate receptor, but in a viral-redirected context where E1A, not DCAF10's intrinsic specificity, determines substrate selection. Luo et al. demonstrated that "OTUD1 stabilizes DDB1 and CUL4 associated factor 10 (DCAF10) and recruits the cullin 4A (CUL4A)-damage specific DNA binding protein 1 (DDB1) complex to promote myeloid cell leukemia sequence 1 (MCL1) degradation" — again confirming CRL4 association but with OTUD1-dependent substrate targeting.

These studies collectively established DCAF10 as a bona fide component of the CRL4 ubiquitin ligase machinery, providing essential context for the Kremer et al. discovery. The adenovirus study is particularly noteworthy because it demonstrates that viral proteins can exploit DCAF10's substrate-recruiting function — a pattern seen with other DCAFs (e.g., DCAF1/VprBP hijacked by HIV-1 Vpr).

{{figure:dcaf10_evidence_summary.png|caption=Summary of evidence supporting DCAF10 as an Ac-Gly N-recognin and the N-terminal glycine quality control pathway. The diagram illustrates how CRL4-DCAF10 (Ac-Gly) and CRL2-ZYG11B/ZER1 (unmodified Gly) provide complementary surveillance of N-terminal glycine modification status in protein quality control.}}

Mechanistic Model: The N-Terminal Glycine Quality Control Network

The discovery of DCAF10 as an Ac-Gly N-recognin completes a conceptual framework for comprehensive N-terminal glycine quality control in human cells. This network can be understood as a three-branch decision tree:

Branch 1: Productive Myristoylation (Normal)

After methionine aminopeptidase (MetAP) exposes the N-terminal glycine, N-myristoyltransferases (NMT1/NMT2) attach a C14 myristoyl group. This lipid anchor enables membrane association and proper protein function. The myristoylated protein is not recognized by any N-degron pathway — the bulky lipid moiety prevents both acetylation and recognition by Gly/N-degron E3 ligases. Chemical proteomic studies have confirmed that NMT modifies over 200 proteins with N-terminal glycines (PMID: 40887160), and the N-terminal glycine is absolutely required for myristoylation as established by substrate specificity studies (PMID: 11955007).

Branch 2: Alternative Acetylation → CRL4–DCAF10 Degradation (Newly Discovered)

If myristoylation fails (NMT inhibition, substrate competition, or stoichiometric imbalance), the exposed N-terminal glycine can be Nα-acetylated by NATs. The resulting Ac-Gly motif is recognized by DCAF10, which recruits the CUL4A–DDB1 scaffold to ubiquitinate the substrate for proteasomal degradation. This pathway specifically targets alternatively acetylated proteins that should have been myristoylated. NMT inhibition has been shown to cause broad loss of myristoylation across >100 protein targets with subsequent cellular effects including ER stress and apoptosis (PMID: 27267252), suggesting that the DCAF10 quality control pathway may be widely relevant under pharmacological NMT inhibition.

Branch 3: Unmodified Glycine → CRL2–ZYG11B/ZER1 Degradation (Previously Known)

If the N-terminal glycine remains completely unmodified (neither myristoylated nor acetylated), it is recognized by the CRL2 substrate receptors ZYG11B and ZER1 through the Gly/N-degron pathway (PMID: 31273098). Structural studies show that ZYG11B/ZER1 use armadillo repeats to form a deep, narrow cavity that engages the first four residues of the Gly/N-degron, with five conserved hydrogen bonds accommodating the unmodified α-amino group (PMID: 34214466). Notably, ZER1 also has capacity to recognize other small N-terminal residues (Ser, Ala, Thr, Cys) that are normally shielded by Nt-acetylation (PMID: 36496439), connecting the Gly/N-degron pathway to broader Nt-acetylation quality control.

Integration with the Broader N-Degron Pathway Landscape

The N-degron pathway field has identified multiple branches over the past three decades (PMID: 22524314; PMID: 26743630; PMID: 39780575):

DCAF10 fills a previously unrecognized gap: no prior N-recognin was known to target acetylated glycine specifically. The canonical Ac/N-degron pathway (MARCHF6) primarily targets Nt-acetylated Met, Ala, Val, Ser, Thr, and Cys — residues that are commonly Nt-acetylated by NatA, NatB, or NatC. N-terminal glycine acetylation is rare under normal conditions (glycine is preferentially myristoylated), making Ac-Gly a conditional degron that only appears when myristoylation fails. A recent study defined the Ac/N-degron recognition domain in MARCHF6, showing that it uses a specific Ac/N-domain to bind Nt-acetylated substrates like RGS2 and PLIN2 (PMID: 40992841; PMID: 39216628), but DCAF10's specificity for Ac-Gly is mechanistically distinct from MARCHF6's broader Ac/N-degron recognition.

Assessment: Is a Specific N-Degron Recognin Annotation Justified?

Evidence Supporting Annotation

Caveats

• Single primary study: All mechanistic evidence comes from one paper (Kremer et al. 2026). Independent replication would strengthen confidence.
• Structural evidence is computational: The AlphaFold 3 model has not been validated by experimental structural determination (X-ray crystallography or cryo-EM).
• Limited endogenous validation: Only Lyn has been validated as an endogenous substrate through genetic loss-of-function experiments. Fyn and Src evidence is limited to in vitro assays.
• No proteome-wide screen: The full substrate repertoire is unknown.
• Competing acetyltransferase unidentified: The specific NAT complex (likely NatA/NAA10) that acetylates Gly when myristoylation fails has not been explicitly identified.

Recommendation

A specific Ac-Gly N-degron recognin annotation is justified for DCAF10, with the caveat that validated endogenous substrates are currently limited to Src-family kinases. The recommended annotation would read:

DCAF10 functions as a substrate receptor for the CRL4 (CUL4A–DDB1) E3 ubiquitin ligase that specifically recognizes N-terminally acetylated glycine (Ac-Gly) degrons, mediating ubiquitin-dependent proteasomal degradation. Validated endogenous substrate: Lyn kinase. Additional in vitro substrates: Fyn, Src.

An annotation limited to "CRL4 complex membership/substrate adaptor activity" would be insufficient — it would fail to capture the specific N-degron recognition function that distinguishes DCAF10 from other DCAFs. The combination of peptide pull-down, in vitro ubiquitination, endogenous Lyn validation, and degron mutagenesis data collectively meets the evidentiary threshold for a specific molecular function annotation. The evidence is methodologically rigorous, employing multiple orthogonal approaches (biochemistry, proteomics, structural prediction, genetics) with both in vitro and cellular validation and both loss-of-function (siRNA, CRISPR KO) and gain-of-function (inducible Lyn-GFP) approaches.

Evidence Base: Literature Summary

Primary Evidence (Peer-Reviewed)

Kremer et al. (2026) — CUL4A-DDB1-DCAF10 is an N-recognin for N-terminally acetylated Src kinases. PMID: 41484149

This is the foundational study establishing DCAF10 as an Ac-Gly N-recognin. Key experimental results:
- Peptide pull-downs + mass spectrometry: Identified DCAF10 as specific Ac-Gly binder
- AlphaFold 3 modeling: Structural support for the DCAF10–Ac-Gly interaction
- In vitro ubiquitination: CUL4A–DDB1–DCAF10 ubiquitinates Ac-SFKs
- Endogenous validation: DCAF10 KD/KO stabilizes Lyn; degron mutagenesis confirms Ac-Gly requirement
- Model: Novel N-degron pathway monitoring myristoylation-to-acetylation switch

Supporting Evidence (CRL4 Complex Context)

Complementary N-Degron Pathway Literature

Preprint / Non-Peer-Reviewed Evidence

No preprint or non-peer-reviewed evidence providing additional data on DCAF10's Ac-Gly recognin function was identified in our literature search. All substantive evidence comes from the peer-reviewed publication PMID: 41484149.

Limitations and Knowledge Gaps

Limitations of Current Evidence

1. Single-study dependence: All Ac-Gly N-recognin evidence derives from Kremer et al. 2026. No independent replication or preprint corroboration has been identified. While the study is methodologically rigorous, independent validation is the gold standard.

2. Computational structural model: The DCAF10–Ac-Gly binding mode is predicted by AlphaFold 3, not determined experimentally. While AlphaFold predictions are increasingly reliable, a high-resolution co-crystal structure or cryo-EM structure would provide definitive evidence for the molecular recognition mechanism and reveal how DCAF10 discriminates Ac-Gly from myristoylated Gly and unmodified Gly.

3. Narrow endogenous validation: Only Lyn has been rigorously validated as an endogenous DCAF10 substrate using both siRNA knockdown and CRISPR knockout with degron-mutant rescue. The evidence for Fyn and Src is limited to in vitro ubiquitination assays, which do not account for cellular context, competing modifications, or compartmentalization. It remains possible that some SFKs are preferentially handled by other quality control pathways in vivo.

4. Unknown sequence context requirements: Whether DCAF10 recognizes any Ac-Gly-bearing protein or requires additional sequence features (residues 2–6, structural elements) has not been systematically tested. The SFKs share substantial N-terminal sequence similarity (conserved myristoylation motif), so the current data cannot distinguish between Ac-Gly-only recognition and an extended motif requirement. For comparison, ZYG11B/ZER1 engage primarily the first four residues of Gly/N-degrons (PMID: 34214466).

5. Quantitative parameters missing: Binding affinities (Kd values), ubiquitination kinetics (kcat/Km), and cellular degradation half-lives have not been reported in the available literature.

6. Acetyltransferase identity: The specific NAT complex responsible for acetylating N-terminal Gly when myristoylation fails has not been explicitly identified. This is a key step in the proposed quality control pathway.

Key Knowledge Gaps

1. Proteome-wide substrate scope: No Global Protein Stability (GPS) profiling, TMT-based degradomics, or systematic peptide library screen has been performed for DCAF10 substrates. The ~200+ human NMT substrates represent a rich candidate pool, but the actual fraction targeted by DCAF10 is unknown.

2. Non-NMT Ac-Gly substrates: Some proteins may acquire N-terminal glycine through endoproteolytic cleavage (e.g., signal peptide processing, caspase cleavage) rather than MetAP processing. Whether such proteins can generate Ac-Gly degrons recognized by DCAF10 is entirely unexplored.

3. Tissue and developmental specificity: DCAF10 expression patterns in the context of Ac-Gly recognin function have not been characterized. NMT substrate proteins and NMT expression vary across cell types, suggesting tissue-specific quality control requirements.

4. Physiological triggers: Under what physiological or pathological conditions does myristoylation failure occur at sufficient scale to activate the DCAF10 pathway? NMT inhibition is pharmacologically achievable (e.g., IMP-1002; PMID: 34695132), but endogenous triggers (metabolic stress, lipid depletion, NMT downregulation) remain undefined.

5. Cancer relevance: DCAF10 is frequently lost in lung adenocarcinomas (PMID: 28336923). Whether loss of DCAF10 leads to accumulation of Ac-Gly-bearing proteins and contributes to oncogenesis — perhaps through stabilization of aberrantly modified SFKs with altered signaling properties — is an important translational question.

Proposed Follow-Up Experiments

High Priority

1. Proteome-wide degradomics upon NMT inhibition ± DCAF10 knockout: Perform TMT-based quantitative proteomics in DCAF10-KO vs. wild-type cells treated with NMT inhibitors (e.g., IMP-1002). Proteins stabilized specifically in DCAF10-KO cells upon NMT inhibition are candidate substrates. This would define the full substrate repertoire and answer whether Ac-Gly recognition extends broadly beyond SFKs.

2. Systematic Ac-Gly peptide library binding assays: Screen a library of Ac-Gly peptides representing all ~200+ NMT substrates for DCAF10 binding (e.g., fluorescence polarization, isothermal titration calorimetry, or AlphaScreen). This would reveal whether sequence context beyond Gly1 influences recognition affinity and help define the minimal degron motif.

3. Experimental structure determination: Obtain a co-crystal structure or cryo-EM structure of DCAF10 (or the full CUL4A–DDB1–DCAF10 complex) bound to an Ac-Gly peptide. This would validate the AlphaFold 3 prediction and reveal the precise molecular basis for Ac-Gly selectivity over unmodified Gly and myristoylated Gly.

Medium Priority

1. Quantitative binding and kinetic parameters: Measure DCAF10–Ac-Gly binding affinity (Kd by ITC or SPR), and compare with Ac-Ala, Ac-Ser, and other Ac-N-terminal residues to confirm glycine specificity. Measure ubiquitination kinetics (kcat/Km) for multiple substrates.

2. In vivo validation of the full SFK family: Perform DCAF10 KD/KO experiments and assess stabilization of endogenous Fyn, Src, Yes, Hck, Fgr, Blk, and Lck. This would determine whether all SFKs are endogenous DCAF10 substrates or only a subset.

3. NMT inhibitor synergy with DCAF10 status: Test whether DCAF10 loss sensitizes or desensitizes cancer cells to NMT inhibitors. If DCAF10 loss prevents degradation of unmyristoylated SFKs, these proteins might accumulate in an aberrant form with distinct (potentially oncogenic) signaling properties.

Lower Priority / Long-Term

1. Acetyltransferase identification: Determine which NAT complex (likely NatA/NAA10) acetylates Gly when myristoylation fails, and whether NAT knockdown synergizes with or phenocopies DCAF10 loss.

2. Physiological trigger characterization: Identify endogenous conditions (metabolic stress, myristic acid deprivation, NMT expression changes during differentiation) under which myristoylation-to-acetylation switching occurs at physiologically meaningful levels.

3. DCAF10 loss in cancer models: Given DCAF10 genomic loss in lung adenocarcinomas, test whether DCAF10 reconstitution suppresses tumorigenic phenotypes in DCAF10-deficient cancer cell lines, and whether this depends on its Ac-Gly recognin activity.

Conclusion

DCAF10 is a validated Ac-Gly N-recognin that functions within the CRL4 E3 ubiquitin ligase complex to degrade proteins bearing N-terminally acetylated glycine degrons. This discovery, reported by Kremer et al. (2026; PMID: 41484149), defines a new branch of the N-degron pathway system that complements the CRL2–ZYG11B/ZER1 Gly/N-degron pathway for unmodified glycine. Together, these pathways provide comprehensive quality control for N-terminal glycine modification, ensuring that proteins failing to undergo proper N-myristoylation are eliminated regardless of their subsequent modification state.

A specific N-degron recognin molecular function annotation is justified for DCAF10 based on direct Ac-Gly binding, reconstituted E3 ligase activity, endogenous Lyn substrate validation with genetic approaches, and degron mutagenesis. An annotation limited to generic CRL4 complex membership would be insufficient to capture this experimentally demonstrated specificity. However, the annotation should reflect that validated endogenous substrates are currently limited to Src-family kinases, and the proteome-wide breadth of Ac-Gly recognition — while biologically plausible for all ~200+ NMT substrates — awaits experimental characterization.

OpenScientist prompt: DCAF10 Ac-Gly N-recognin specificity

Investigate the evidence that human DCAF10 is a CRL4 substrate receptor recognizing N-terminally acetylated glycine degrons, and assess how broad that substrate-recognition activity is across the proteome.

Focus on:

• direct Ac-Gly binding and CUL4A-DDB1-DCAF10 ubiquitination evidence;
• endogenous substrates, especially Src-family kinases such as Lyn, Fyn, and Src;
• whether Ac-Gly recognition is restricted to normally N-myristoylated proteins or applies more broadly;
• degron mutagenesis, rescue experiments, and proteome-wide candidate testing.

Please assess whether a specific N-degron-recognin molecular-function annotation is justified now, or whether only CRL4 complex membership/substrate-adaptor activity is supported. Include PMIDs and note any very recent/preprint evidence separately from peer-reviewed evidence.