N-terminal truncation diversifies USP18 function during early myogenesis

Ubiquitin-specific protease 18 (USP18) is primarily studied for its dual role in type-I interferon (IFN-I) signaling. At basal levels, however, USP18 also regulates muscle cell development, independently of IFN-I signaling. USP18 exists as a canonical long isoform (LF) and a short isoform (SF) generated by N-terminal truncation. Endogenous SF accumulates in the nucleus during muscle differentiation, but its cellular functions remain poorly understood. Here, we compared USP18 SF and LF in muscle cells stably overexpressing each isoform. We show that their distinct N-termini are associated with marked differences in protein stability, subcellular localization, and protein-protein interactions. SF displayed a substantially expanded interactome, with increased association with nuclear proteins, chaperones, and regulators of protein homeostasis. Functionally, SF enhanced an early stage of myogenic differentiation and was associated with changes in alternative splicing, including key sarcomere genes. Together, these findings identify the SF USP18 proteoform as a potential regulator of muscle differentiation and highlight the N-terminus as a determinant of non-canonical USP18 functions and proteoform diversity.