Palmitoyl-proteomic profiling of large yellow croaker monocytes/macrophages with or without zDHHC20b knockdown during Pseudomonas plecoglossicida infection

Full MS scans were acquired over m/z 300-1500, with 20 PASEF-MS/MS scans per cycle. The MS/MS scan range was m/z 400-8500, and the isolation window was 7 m/z. Raw DIA data were processed using DIA-NN (v1.8).

Trypsin/P was specified as the cleavage enzyme with up to one missed cleavage allowed. N-terminal methionine excision and carbamidomethylation of cysteine were set as fixed modifications. The false discovery rate (FDR) was controlled at <1%.

Functional enrichment analysis of differentially palmitoylated proteins was performed using Gene Ontology (GO) and KEGG pathway annotations. GO IDs were assigned using eggNOG-mapper, and proteins were classified into biological process, cellular component, and molecular function categories. KEGG annotation was performed by BLAST search against the KEGG database (e-value ≤1e-4). Fisher's exact test identified significantly enriched terms, defined as those with fold enrichment >1.5 and p-value <0.05. The selection criteria for differentially palmitoylated proteins were defined as an absolute log2 (fold change) ≥1 (corresponding to a ≥2-fold alteration in protein abundance), in conjunction with a false discovery rate (FDR)-adjusted p-value <0.05 as the significance threshold.