Protein palmitoylation is a reversible lipid modification that plays pivotal roles in various physiological processes. Aberrant palmitoylation frequently contributes to cancer development. However, the precise contributions of S-palmitoylating enzymes, known as palmitoyltransferases, to the initiation and progression of hepatocellular carcinoma (HCC) remain poorly understood. The role of zinc finger DHHC-type palmitoyltransferase 20 (ZDHHC20) in HCC tumorigenesis was assessed using diethylnitrosamine (DEN)-induced and DEN plus CCl4 HCC mouse models in ZDHHC20 knockout animals. To delineate the downstream targets and molecular mechanisms of ZDHHC20, the study employed palmitoylation liquid chromatography-mass spectrometry analysis, acyl-biotin exchange assay, co-immunoprecipitation, ubiquitination assays, protein half-life assays, and immunofluorescence microscopy. Genetic ablation of ZDHHC20 markedly suppressed hepatocarcinogenesis triggered by chemical carcinogens in both in vivo HCC mouse models. Palmitoylation profiling identified 97 proteins harboring 123 cysteine residues that undergo ZDHHC20-dependent palmitoylation. Notably, fatty acid synthase (FASN) was palmitoylated by ZDHHC20 at cysteine sites 1471 and 1881. Either genetic deletion or pharmacological blockade of ZDHHC20, along with substitution mutations at the critical FASN cysteines (C1471S/C1881S), accelerated FASN degradation. ZDHHC20-catalyzed palmitoylation of FASN further antagonized its breakdown via the ubiquitin-proteasome system mediated by the E3 ubiquitin ligase complex SNX8-TRIM28. The results underscore the essential function of ZDHHC20 in driving hepatocarcinogenesis and uncover an antagonistic interplay between palmitoylation and ubiquitination that regulates protein stability.
