The conjugation of ubiquitin, a 76-amino-acid peptide, to a protein substrate provides a tag that either marks the labelled protein for degradation or modulates its function. The process of protein ubiquitylation--which is catalysed by coordinated enzymatic reactions that are mediated by enzymes known as E1, E2 and E3--has an important role in the modulation of immune responses. Importantly, protein ubiquitylation is a reversible process, and removal of ubiquitin molecules is mediated by de-ubiquitylating enzymes: for example, A20, which has been implicated in the regulation of immune responses. In addition, the conjugation of ubiquitin-like molecules, such as ISG15 (interferon-stimulated protein of 15 kDa), to proteins is also involved in immune regulation. This Review covers recent progress in our understanding of protein ubiquitylation in the immune system.