Trained immunity is often defined as long-term functional modification of chromatin in cells of the innate immune system. This occurs following a stimulus that alters responsiveness to that or other stimuli upon repeated exposures. Upon exposure to type I interferon (IFN-I), a key antiviral and inflammatory cytokine, cells become hyporesponsive to subsequent IFN-I stimulations, protecting themselves from excessive inflammation. Herein, we show that this form of trained immunity is not chromatin but rather posttranslationally regulated. ISG15 and USP18 are IFN-I-stimulated genes (ISGs) that, as free intracellular molecules, control the cellular response to IFN-I at the IFN-I receptor level. Additionally, ISG15 covalently attaches to proteins in a nonspecific manner, in a process termed ISGylation, and USP18 reverses this modification in a process termed deISGylation. We have studied patients with defective ISGylation. They present a higher incidence of atopy and inflammation. Herein, we demonstrate that basal, IFN-I-independent ISGylation serves to sequester free ISG15 and thereby keeps cells ready for optimal response to IFN-I. Furthermore, in an IFN-I-dependent manner, cells that are devoid of ISGylation or deISGylation exhibit a remarkable paucity of trained immunity compared to WT cells. These results suggest that ISGylation and deISGylation control the response to IFN-I by sequestering or supplying, respectively, free ISG15.
