Abstract: 【Abstract】 Objective To investigate the palmitoyl-proteomic profiles of psoriatic lesions and their role in psoriasis pathogenesis and targeted modification therapy. Methods In March 2024, 5 adult patients with moderate-to-severe plaque psoriasis and 5 healthy controls were recruited from Shanghai Skin Disease Hospital. Psoriatic lesion （PS） samples and normal skin tissue （NN） samples were collected, and label-free quantitative palmitoyl-proteomic analysis was performed using liquid chromatography-tandem mass spectrometry （LC-MS/MS）. Acyl-biotin exchange combined with Western blot analysis was employed to verify the palmitoylation status of key proteins in tissue samples. Gene Ontology （GO） and Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment analyses were conducted to analyze the biological functions of differentially palmitoylated proteins in psoriasis. The Gene Expression Omnibus database analysis showed that the palmitoyltransferase-related genes ZDHHC12 and ZDHHC21 were significantly upregulated in psoriatic lesions, and qPCR was performed to verify their expression in tissue samples. Sixteen 7-week-old specific-pathogen-free C57BL/6 mice were randomly divided into 4 groups: mice in the 2-BP control group and IMQ + 2-BP group were injected intraperitoneally with the broad-spectrum palmitoylation inhibitor 2-bromopalmitate （2-BP） for 8 consecutive days, while those in the IMQ + vehicle group received injections of an equal volume of vehicle; from day 3 after the first injection, psoriasiform skin lesions were induced by topical application of imiquimod （IMQ） on the mouse dorsal skin for 5 consecutive days in the IMQ + 2-BP group and IMQ + vehicle group; mice in the blank control group received no treatment. Changes in skin manifestations were observed during the experiment. On day 9, the mice were sacrificed, and skin histopathological features were assessed. Immunofluorescence staining was performed to determine the expression of filaggrin （Flg） in tissues, and qPCR to determine the mRNA expression of interleukin-22 （IL-22）, C-C motif chemokine ligand （CCL）5, CCL20, and Flg. Total protein palmitoylation levels in the skin were also measured. The two-independent-samples t test was used for comparisons between two groups, one-way analysis of variance for comparisons among multiple groups, and ?ídák's test for post-hoc multiple comparisons. Results A total of 1 453 differentially S-palmitoylation sites were identified in psoriasis lesions by LC-MS/MS. Key proteins in the PS group, including signal transducer and activator of transcription 1 （STAT1）, fatty acid-binding protein 5 （FABP5）, and keratin 16 （KRT16）, showed increased palmitoylation levels. Palmitoylation detection showed that the relative palmitoylation levels of STAT1, FABP5, and KRT16 were significantly higher in the PS group （2.50 ± 1.00, 4.06 ± 1.05, 1.17 ± 0.37, respectively） than in the NN group （0.46 ± 0.17, 0.13 ± 0.06, 0.10 ± 0.08, respectively; all?P?< 0.05）. GO analysis showed that downregulated palmitoylated proteins were mainly involved in cytoskeletal regulation, extracellular matrix interactions, and complement and coagulation cascades; KEGG analysis showed that upregulated palmitoylated proteins were mainly enriched in immune-related pathways, including interferon, NOD-like receptor, retinoic acid-inducible gene Ⅰ-like receptor, IL-17 signaling, and Th17 cell differentiation pathways. qPCR showed that the relative mRNA expression levels of ZDHHC12 and ZDHHC21 were significantly higher in the PS group （1.52 ± 0.13, 2.73 ± 0.94, respectively） than in the healthy control group （0.94 ± 0.22, 0.96 ± 0.54, respectively; both?P?< 0.05）. In the animal experiment, mice in the IMQ + vehicle group developed psoriasiform lesions from day 2 after IMQ application, presenting as pale erythematous patches with a few white scales; on day 4, the erythema deepened in color and the scales thickened, with micaceous desquamation; meanwhile, only mild pale erythematous patches with thin and loose scales were observed in the IMQ + 2-BP group. Compared with the IMQ + vehicle group, the IMQ + 2-BP group showed significantly decreased total protein palmitoylation levels and mRNA expression of inflammatory cytokines IL-22, CCL5, and CCL20, but significantly increased mRNA and protein expression of Flg （all?P?< 0.05）. Conclusion This study systematically characterized the palmitoyl-proteomic landscape of psoriasis, screened out differentially palmitoylated proteins as well as their enriched signaling pathways, and laid preliminary experimental foundations for the mechanistic research and palmitoylation-targeted therapy of psoriasis.

Key words: Psoriasis, Metabolomics, Protein palmitoylation, STAT1 transcription factor, Models, animal, Immunoregulation