ZDHHC4

ZDHHC4 is an ER-resident DHHC-family palmitoyl acyltransferase that S-palmitoylates a diverse array of substrates to control their stability, trafficking, and downstream signaling across innate immunity, pain sensation, cardiac metabolism, neurodegeneration, and cancer. It palmitoylates MAVS at Cys79 (recruited by CPT1A), stabilizing MAVS by switching ubiquitination from K48- to K63-linked chains to potentiate type I interferon signaling (PMID:38016475); palmitoylates TRPV1 at multiple cysteines to promote its lysosomal degradation and attenuate inflammatory pain (PMID:39528731); and palmitoylates GSK3β at Cys14 to activate EZH2–STAT3 signaling in glioblastoma stem cells (PMID:35606353). ZDHHC4 also palmitoylates CD36 to drive its sarcolemmal localization and fatty acid uptake—a process transcriptionally upregulated by FoxO1 in the diabetic heart and by ATF3 in macrophages (PMID:40357580, PMID:39047111)—and palmitoylates ZEB-2, NFATC4, KAI1, and CCDC50, linking its activity to EMT, nuclear transcription factor trafficking, anti-angiogenesis, and selective autophagy (PMID:41603995, PMID:35150145, PMID:34530889, PMID:41566717).

1. 2015 Medium

   Establishing ZDHHC4 as a functional palmitoyl acyltransferase: a MYTH screen identified ZDHHC4 as a PAT that interacts with and palmitoylates the D2 dopamine receptor, affecting D2R stability and surface expression—the first direct evidence linking ZDHHC4 to substrate palmitoylation.

   Evidence Membrane yeast two-hybrid screen, Co-IP, click chemistry palmitoylation assay, cysteine mutagenesis in heterologous cells

   PMID:26535572

   Open questions at the time

   • Single lab study without independent replication
   • Specific palmitoylated cysteine(s) on D2R not mapped
   • No in vivo validation of D2R palmitoylation by ZDHHC4
2. 2021 Medium

   Extending ZDHHC4 substrates to membrane proteins with distinct biological outputs: ZDHHC4 was shown to palmitoylate KAI1/CD82, controlling its membrane localization and enabling anti-angiogenic LIF induction via Src/p53, and separately to palmitoylate the SARS-CoV-2 spike protein to facilitate viral syncytia formation and entry.

   Evidence Palmitoylation assays, loss-of-function experiments, in vivo angiogenesis models (KAI1); ZDHHC overexpression panel, pseudovirus entry and syncytia assays (spike protein)

   PMID:34528721 PMID:34530889

   Open questions at the time

   • For SARS-CoV-2 spike, ZDHHC4 was not individually dissected from other ZDHHC family members
   • Palmitoylation site(s) on KAI1 not mapped at residue level
   • No structural basis for substrate selectivity
3. 2022 Medium

   Defining ZDHHC4 as a signaling-modulating enzyme through site-specific palmitoylation: ZDHHC4 palmitoylates GSK3β at Cys14 to shift its phosphorylation state and activate EZH2–STAT3 in GBM, and palmitoylates NFATC4 to enable its cytoplasm-to-nucleus trafficking, linking ZDHHC4 to both cancer stemness and lysosomal Ca²⁺ homeostasis in neurodegeneration.

   Evidence Cys14 mutagenesis, phosphorylation analysis, stemness/drug-resistance readouts (GSK3β); subcellular fractionation, Ppt1-KO mice, rescue experiments (NFATC4)

   PMID:35150145 PMID:35606353

   Open questions at the time

   • ZDHHC4 versus ZDHHC8 individual contributions to NFATC4 palmitoylation not fully resolved
   • In vivo GBM model validation of ZDHHC4 dependence lacking
   • Structural basis for Cys14 selectivity on GSK3β unknown
4. 2023 High

   Revealing an adaptor-dependent recruitment mechanism: CPT1A recruits ER-localized ZDHHC4 to palmitoylate MAVS at Cys79, switching ubiquitination from K48- to K63-linked chains and thereby stabilizing MAVS to amplify type I interferon signaling—the first demonstration that ZDHHC4 substrate access can be controlled by an adaptor protein.

   Evidence Reciprocal Co-IP, Cys79 mutagenesis, ubiquitination assays, IFN-I functional readout

   PMID:38016475

   Open questions at the time

   • Whether CPT1A-mediated recruitment is a general mechanism for other ZDHHC4 substrates is untested
   • Structural details of CPT1A–ZDHHC4 interaction unresolved
5. 2024 High

   Demonstrating that ZDHHC4 controls ion channel turnover and pain: ZDHHC4 palmitoylates TRPV1 at four cysteines to promote its lysosomal degradation, counterbalanced by depalmitoylase APT1, establishing a palmitoylation–depalmitoylation cycle that regulates inflammatory pain in vivo.

   Evidence Co-IP, quadruple-cysteine mutagenesis, acyl-biotin exchange, electrophysiology, in vivo pain behavior assays

   PMID:39528731

   Open questions at the time

   • Whether palmitoylation at each individual cysteine contributes equally to TRPV1 degradation is unresolved
   • Mechanism by which palmitoylation targets TRPV1 to lysosomes not defined
6. 2024 High

   Establishing transcriptional regulation of ZDHHC4 as a metabolic control point: FoxO1 transcriptionally upregulates ZDHHC4 in diabetic cardiomyocytes, and ATF3 does so in macrophages, with both converging on CD36 palmitoylation to drive its plasma membrane localization and fatty acid uptake, directly linking ZDHHC4 expression to metabolic disease.

   Evidence ChIP-seq/qPCR, luciferase assays, cardiomyocyte-specific FoxO1-KO mice, pharmacological ZDHHC inhibition (FoxO1); Atf3 overexpression/knockdown, CD36 cysteine mutagenesis, subcellular fractionation (ATF3)

   PMID:39047111 PMID:40357580

   Open questions at the time

   • Whether FoxO1 and ATF3 regulate ZDHHC4 in a tissue-specific or generalizable manner is unknown
   • Relative contribution of ZDHHC4 versus ZDHHC5 to CD36 palmitoylation not individually quantified
7. 2025 Medium

   Confirming ZDHHC4 as a hepatic regulator of CD36-dependent lipid uptake through a parallel pathway: EVA1A deficiency upregulates ZDHHC4/5 while repressing APT1, boosting CD36 palmitoylation and membrane localization to increase fatty acid uptake in hepatocytes.

   Evidence Hepatocyte-specific Eva1a-KO, transcriptomics, palmitoylation and subcellular fractionation assays

   PMID:41306774

   Open questions at the time

   • ZDHHC4 and ZDHHC5 not individually separated in this context
   • Downstream metabolic phenotype not fully dissected for ZDHHC4 alone
8. 2026 High

   Expanding ZDHHC4 to EMT regulation and selective autophagy: ZDHHC4 palmitoylates ZEB-2 at C478 to promote its deubiquitination and stabilization (driving melanoma EMT), and palmitoylates CCDC50 to target MAP2K4 for autophagic degradation; the natural compound lactucopicrin directly binds His72 of ZDHHC4 to enhance its enzymatic activity.

   Evidence Mass spectrometry, Co-IP, C478 mutagenesis, EMT readout (ZEB-2); DARTS binding assay, His72 mutagenesis, in vivo osteoarthritis model (CCDC50/lactucopicrin)

   PMID:41566717 PMID:41603995

   Open questions at the time

   • Whether lactucopicrin-mediated activation of ZDHHC4 is specific or affects other ZDHHCs is not addressed
   • Structural basis of His72 in ZDHHC4 catalytic mechanism not fully elucidated

• Unresolved: the structural basis for ZDHHC4 substrate selectivity, how adaptor recruitment generalizes across substrates, and whether ZDHHC4 has substrates beyond those already identified that explain its broad biological footprint.
• No crystal or cryo-EM structure of ZDHHC4
• No systematic unbiased palmitoyl-proteomics to define the full ZDHHC4 substrate repertoire
• Catalytic mechanism and role of His72 versus the DHHC motif not structurally resolved