Structural insights into the Urm1-Uba4 pathway and its biological roles

Ubiquitin-related modifier 1 (Urm1) is a unique and evolutionarily conserved member of the ubiquitin-like protein (UBL) family that represents a molecular link between ancestral sulfur carrier proteins (SCPs) and canonical eukaryotic UBLs. Urm1 is required for the thiolation of tRNAs and a non-canonical post-translational modification, called 'urmylation'. Activation of Urm1 by its E1-like enzyme, ubiquitin-like protein activator 4 (Uba4), involves the sequential adenylation, thioesterification, and thiocarboxylation of Urm1's C-terminus. Thereby, Urm1 can provide sulfur for the tRNA modification reaction or catalyze its conjugation to target proteins through a mechanism that is independent of E2-conjugating enzymes and E3 ligases. Recent structural studies have resolved several key intermediates of the fungal Uba4-Urm1 system, shedding light onto its two distinct subdomains and their dynamical interplay. Notably, Urm1 also interacts with several additional up- or downstream partners of the two pathways. Foremost, urmylation couples an UBL-conjugation reaction with the persulfidation of a cysteine residue in the target proteins. This protective oxidative post-translational modification underscores Urm1's central role in redox regulation and cellular stress responses. Here, we aim to summarize the most recent mechanistic insights and structural advances in the eukaryotic Urm1-Uba4 pathway.