Multi‑PTM landscape of four lysine acylations at shared residues in oral squamous cell carcinoma

Posttranslational modifications (PTMs) are central regulators of cellular signaling and play critical roles in cancer biology. While individual PTMs have been implicated in oral squamous cell carcinoma (OSCC), the crosstalk and coregulatory patterns among different PTM types in this malignancy remain largely unexplored. Integrated proteomic analysis via liquid chromatography-tandem mass spectrometry (LC–MS/MS) was performed to profile four lysine acylations (2-hydroxyisobutyrylation (Khib), crotonylation (Kcr), acetylation (Kac), and malonylation (Kma)) in OSCC tissues and matched adjacent tissues. Proteins harboring at least one differentially modified peptide or site were defined as differentially modified proteins (DMPs) and subsequently subjected to functional enrichment analysis via Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment. Protein protein interaction (PPI) networks were constructed via STRING and Cytoscape to screen hub modified proteins, and Spearman correlation analysis was applied to evaluate the regulatory relationships among the four PTMs. Eighty proteins were identified as being commonly modified by at least three of the studied PTMs. KEGG pathway analysis, which maps proteins to pre-defined pathway categories, indicated that these DMPs were annotated to pathways related to Salmonella infection, motor proteins, focal adhesion, and proteoglycans in cancer. A key finding was the identification of specific lysine residues that were found to be modified by multiple PTM types across samples. Notably, the modification levels of Khib, Kcr, and Kac at the FLNA-K837 and DCN-K164 site differed between OSCC tissues and adjacent tissues ( P < 0.05). This study describes the co-modification patterns among multiple PTMs in OSCC. These combinatorial modifications may contribute to regulating key metabolic and signaling pathways. These preliminary observations warrant validation in larger cohorts and functional studies. The identified PTM sites may serve as candidates for further investigation as potential biomarkers. These findings identify key PTM site hubs such as FLNA and DCN, whose site‑specific acylation patterns show alterations in OSCC. Further studies are needed to determine whether these modifications have clinical utility. The current findings should be considered hypothesis-generating rather than definitive.