ZDHHC20

ZDHHC20 is an integral membrane protein S-acyltransferase (palmitoyltransferase) that transfers palmitoyl groups from palmitoyl-CoA to cysteine residues on a broad array of substrate proteins, thereby controlling their membrane association, stability, and signaling competence across immunity, DNA damage repair, and oncogenesis. Its catalytic mechanism involves membrane deformation that hydrates the DHHC-motif catalytic cysteine for autoacylation, with a transmembrane hydrophobic cavity that confers selectivity for C16 acyl chains (PMID:31858978, PMID:35563480). Validated substrates include EGFR (C-terminal tail pinning and signal attenuation) (PMID:27153536), IFITM3 (antiviral activity at lysosomes) (PMID:29079573), ORAI1 (lipid raft targeting for T cell Ca²⁺/NFAT signaling) (PMID:34913437), YTHDF3 (protection from chaperone-mediated autophagy, stabilizing MYC mRNA in pancreatic cancer) (PMID:38821916), FASN (competition with ubiquitin-proteasome degradation in hepatocarcinogenesis) (PMID:39696259), CD80 (membrane retention and T cell costimulation) (PMID:38467718), KAP1/TRIM28 (ATM-phosphorylation-dependent palmitoylation enhancing chromatin binding and DNA damage repair) (PMID:41109928), CMPK2 (mitochondrial localization and MAVS-dependent antiviral IFN-I responses) (PMID:42011944), and FGFR3 (RAB11A-scaffolded plasma membrane retention in bladder cancer) (PMID:41586981). For multiple substrates, ZDHHC20-mediated palmitoylation competes with ubiquitination to protect proteins from proteasomal or lysosomal degradation, establishing palmitoylation as a stability switch (PMID:39696259, PMID:38467718, PMID:38821916).

1. 2016 High

   The first substrate and physiological function of ZDHHC20 were established: palmitoylation of EGFR C-terminal cysteines pins its tail to the plasma membrane, restraining signaling—revealing that loss of ZDHHC20-mediated palmitoylation sensitizes cancer cells to EGFR inhibitors.

   Evidence Mass spectrometry, Cys-to-Ala mutagenesis, DHHC20 knockdown, and transformation/migration assays in cancer cell lines

   PMID:27153536

   Open questions at the time

   • Whether EGFR palmitoylation by ZDHHC20 is regulated by upstream signals was not addressed
   • The stoichiometry of palmitoylation on individual EGFR cysteines was not determined
2. 2017 High

   ZDHHC20 was shown to palmitoylate the antiviral effector IFITM3 and uniquely colocalize with it at lysosomes, distinguishing it from other DHHC enzymes and linking its activity to innate antiviral defense.

   Evidence Systematic ZDHHC library screen across 23 family members, ZDHHC KO cell lines, colocalization imaging, and influenza infection assays

   PMID:29079573

   Open questions at the time

   • Whether ZDHHC20 lysosomal localization is dynamically regulated was not resolved
   • The precise palmitoylation sites on IFITM3 modified by ZDHHC20 specifically (versus other ZDHHCs) were not delineated
3. 2019 High

   Reconstitution and simulation revealed that ZDHHC20 deforms the cytoplasmic membrane leaflet to hydrate its DHHC catalytic cysteine, establishing the structural basis for its autoacylation step.

   Evidence Biochemical reconstitution in POPC nanodiscs combined with microsecond all-atom molecular dynamics simulations

   PMID:31858978

   Open questions at the time

   • No high-resolution experimental structure of ZDHHC20 was obtained
   • How membrane deformation couples to the subsequent substrate acylation (transacylation) step was not determined
4. 2021 High

   ZDHHC20-mediated palmitoylation of ORAI1 at Cys143 was shown to be required for lipid-raft targeting, TCR signaling at the immune synapse, sustained Ca²⁺ entry, and IL-2 secretion, establishing ZDHHC20 as a regulator of adaptive immunity.

   Evidence Cys143 mutagenesis, patch-clamp electrophysiology, SOCE measurements, NFATC1 translocation, and IL-2 secretion in Jurkat T cells

   PMID:34913437

   Open questions at the time

   • Whether ZDHHC20 palmitoylates ORAI1 in primary T cells in vivo was not tested
   • Whether other ZDHHCs contribute to ORAI1 palmitoylation was not fully excluded
5. 2022 Medium

   Molecular dynamics and structure-activity studies defined the acyl chain length selectivity mechanism: a hydrophobic cavity in ZDHHC20's transmembrane helices accommodates C16 palmitoyl-CoA, and a specific alanine residue determines isoform-selective inhibitor binding.

   Evidence MD simulations with V185G mutant shifting to C18 selectivity; SAR of acrylamide-based inhibitors with isoform selectivity assays

   PMID:35563480 PMID:36262404

   Open questions at the time

   • The C16 selectivity mechanism was primarily computationally derived and not validated by in vitro reconstitution with different acyl-CoA substrates
   • Inhibitor selectivity was tested against ZDHHC2 but not against the full ZDHHC family
6. 2024 High

   Multiple studies converged to show that ZDHHC20-mediated palmitoylation stabilizes substrates (YTHDF3, FASN, CD80) by competing with ubiquitin-dependent degradation, establishing a general palmitoylation-versus-ubiquitination stability switch operating in cancer and immunity.

   Evidence Palmitoylation site mutagenesis, ubiquitination assays, protein half-life measurements, and in vivo models for YTHDF3/MYC in pancreatic cancer, FASN in hepatocarcinogenesis, and CD80 in T cell costimulation

   PMID:38467718 PMID:38821916 PMID:39696259

   Open questions at the time

   • Whether the palmitoylation-ubiquitination competition involves direct steric occlusion of the same or proximal lysine residues was not resolved
   • The E3 ligases targeting YTHDF3 and CD80 for degradation upon loss of palmitoylation were not identified (only the SNX8-TRIM28 complex was identified for FASN)
7. 2024 Medium

   An in vivo shRNA screen identified ZDHHC20 as essential for pancreatic cancer metastatic outgrowth, functioning through modulation of tumor–NK cell interactions rather than cell-autonomous growth.

   Evidence In vivo shRNA screen with immunocompromised and NK-cell-depleted animal models; chemical genetics substrate profiling

   PMID:38733589

   Open questions at the time

   • The specific ZDHHC20 substrates mediating NK cell evasion were identified by chemical genetics but not individually validated
   • Whether the immune evasion mechanism operates in human patients was not tested
8. 2024 Medium

   ZDHHC20 was shown to activate PI3K-AKT signaling in hepatocellular carcinoma, promoting proliferation that is reversible by PI3K/AKT inhibitors, though the direct palmitoylation substrate was not identified.

   Evidence ZDHHC20 knockdown/overexpression, phospho-PI3K/AKT western blots, pharmacological inhibitors (LY294002, MK2206), xenograft model

   PMID:39309302

   Open questions at the time

   • The palmitoylation substrate linking ZDHHC20 to PI3K-AKT activation was not identified
   • Whether this effect is independent of ZDHHC20's known substrates (e.g., EGFR, FASN) was not tested
9. 2025 High

   ATM-dependent phosphorylation of ZDHHC20 at Ser339 was shown to activate palmitoylation of KAP1/TRIM28, enhancing its chromatin binding and recruitment of DDR factors BRCA1 and 53BP1—revealing ZDHHC20 as a signal-regulated enzyme in the DNA damage response.

   Evidence ZDHHC family screen, acyl-biotin exchange, phospho-site mutagenesis, ZDHHC20 KO mice, chromatin fractionation, DDR factor recruitment and radiosensitivity assays

   PMID:41109928

   Open questions at the time

   • Whether other DNA damage kinases phosphorylate ZDHHC20 at additional sites was not explored
   • The structural basis for how Ser339 phosphorylation enhances enzymatic activity was not determined
10. 2026 High

    ZDHHC20 palmitoylation of CMPK2 at Cys137/153 was shown to maintain its mitochondrial localization and promote MAVS-dependent type I IFN responses, with PPT1 acting as the counteracting depalmitoylase—establishing a writer-eraser pair for antiviral innate immunity.

    Evidence Metabolic chemical library screen, CMPK2 Cys mutagenesis, PPT1 inhibitor DC661, IFN-I measurement, viral replication assays, in vivo diet and inhibitor studies

    PMID:42011944

    Open questions at the time

    • Whether other ZDHHCs also palmitoylate CMPK2 was not fully excluded
    • The structural basis of ZDHHC20-CMPK2 recognition was not resolved
11. 2026 Medium

    RAB11A was identified as a scaffold that recruits ZDHHC20 to palmitoylate FGFR3 for plasma membrane retention in bladder cancer, with SREBP2-driven transcription of RAB11A as an upstream regulatory node.

    Evidence Co-immunoprecipitation, promoter-reporter assays, RAB11A/ZDHHC20 knockdown, xenograft models

    PMID:41586981

    Open questions at the time

    • The palmitoylation site(s) on FGFR3 were not validated by site-directed mutagenesis
    • Whether RAB11A scaffolding is specific to FGFR3 or extends to other ZDHHC20 substrates was not tested
12. 2026 Medium

    MEF2A was identified as a direct transcriptional activator of ZDHHC20, and ZDHHC20 was placed upstream of NF-κB signaling in AML, promoting proliferation, glycolysis, and chemoresistance.

    Evidence ChIP assay, dual-luciferase reporter, MEF2A-KD/ZDHHC20-OE epistasis rescue, NF-κB pathway western blots, xenograft model

    PMID:41882733

    Open questions at the time

    • The palmitoylation substrate connecting ZDHHC20 to NF-κB activation was not identified
    • Whether MEF2A regulation of ZDHHC20 operates in non-AML contexts was not explored

• Key unresolved questions include the high-resolution structure of ZDHHC20, the rules governing substrate recognition across its diverse substrates, whether signal-dependent regulation (as shown for ATM-Ser339 phosphorylation) is a general feature, and the therapeutic window of ZDHHC20 inhibition given its roles in both tumor promotion and immune function.
• No experimental high-resolution structure of ZDHHC20 has been determined
• Substrate recognition determinants beyond the hydrophilic ridge in TM4 remain undefined
• In vivo consequences of systemic ZDHHC20 inhibition on immune homeostasis are unknown