The ubiquitin-like modifier FAT10 is up-regulated in many different cell types by IFNγ and TNFα (TNF) and directly targets proteins for proteasomal degradation. FAT10 gets covalently conjugated to its conjugation substrates by the E1 activating enzyme UBA6, the E2 conjugating enzyme USE1, and E3 ligases including Parkin. To date, USE1 was supposed to be the only E2 enzyme for FAT10ylation, and we show here that a knockout of USE1 strongly diminished FAT10 conjugation. Remarkably, under inflammatory conditions in the presence of TNF, FAT10 conjugation appears to be independent of USE1. We report on the identification of additional E2 conjugating enzymes, which were previously not associated with FAT10. We confirm their capacity to be charged with FAT10 onto their active site cysteine, and to rescue FAT10 conjugation in the absence of USE1. This finding strongly widens the field of FAT10 research by pointing to multiple, so far unknown pathways for the conjugation of FAT10, disclosing novel possibilities for pharmacological interventions to regulate FAT10 conjugation under inflammatory conditions and/or viral infections.
© 2023 Schnell et al.