A novel stop-loss variant in ISG15 causes C-terminal dysfunction and impairs USP18-mediated negative feedback.

Introduction: ISG15 is a type I interferon (IFN-I)-inducible ubiquitin-like protein that restrains IFN-I signaling. Classical ISG15 deficiency is thought to cause disease mainly by disrupting USP18 stabilization, which leads to sustained IFN-I signaling and interferon-driven autoinflammation. Whether patient-derived C-terminal ISG15 variants can impair IFN-I negative feedback through altered C-terminal functional presentation has remained unclear. Methods: We studied a Chinese boy with early-onset recurrent fever, severe ulcerative skin lesions, and elevated liver enzymes. Whole-exome sequencing was performed to identify ISG15 variants. Patient peripheral blood mononuclear cells (PBMCs) and skin tissue were analyzed for IFN-I pathway activation. Functional effects of the identified variants were assessed in 293T cells, and structural analysis was performed. The patient was treated with the JAK inhibitor baricitinib. Results: Two novel compound heterozygous ISG15 variants were identified: c.234C>A, which produced a truncated ISG15 protein, and c.498A>T, a stop-loss variant predicted to extend the ISG15 C-terminus and associated with markedly impaired ISGylation. Patient PBMCs and skin tissue showed increased p-STAT1 signaling and IFN-I pathway activation, with decreased USP18 expression and altered ISG15-USP18 regulation in PBMCs. In 293T cells, c.498A>T alone did not reduce USP18 abundance, suggesting that this variant contributes to disease through impaired C-terminal functional presentation rather than USP18 loss alone. Structural analysis suggested that the C-terminal extension may alter LRLRGG motif presentation, weakening ISG15-USP18-mediated negative feedback. Baricitinib reduced IFN-I-related markers and was associated with improvement of ulcerative skin lesions and liver enzyme abnormalities. Discussion: This study expands the genetic and mechanistic spectrum of ISG15-associated type I interferonopathy, and supports that JAK-STAT inhibition with baricitinib can effectively control IFN-I-driven inflammation.