Interferon-stimulated gene 15kDa (ISG15) is a ubiquitin-like molecule that, in a similar fashion to ubiquitin conjugation, can be covalently linked to cellular proteins. In addition, ISG15 was also found to act as a conjugation-independent signaling molecule. ISG15 and the other components of the ISGylation pathway are induced by type I interferons and play a key role in resistance to infection, in immune regulation and likely in cancer. However, the mechanisms of action of ISG15 secretion and conjugation are largely undefined. In this project, we aim to address a number of major outstanding questions about its mechanisms of action.