Understanding protein ISGylation, a multifaceted posttranslational modification.

ISG15 represents a key ubiquitin-like modifier induced primarily by interferon signaling. ISG15 is synthesized as a precursor, processed to a mature form, and covalently conjugated to substrates through a dedicated E1-E2-E3 enzymatic cascade involving UBE1L, UBE2L6, and E3 ligases such as HERC5, TRIM25, and ARIH1. This modification is reversed by deISGylases, particularly the highly specific protease USP18, which also negatively regulates interferon signaling. ISGylation impacts diverse molecular and cellular processes, including protein stability and function, protein-protein interaction, autophagy, transcription/translation, DNA damage response, and innate immunity. Advances in chemical biology and mass spectrometry-based proteomics have enabled the characterization of enzymes involved in (de)ISGylation and mapping of ISGylated proteins and sites. Dysregulated ISGylation is implicated in cancer, infection, neurodegenerative disorders, and inflammatory diseases, underscoring its broad pathophysiological relevance.