Ubiquitin acetylation inhibits polyubiquitin chain elongation

Ubiquitylation is a versatile post‐translational modification ( PTM ). The diversity of ubiquitylation topologies, which encompasses different chain lengths and linkages, underlies its widespread cellular roles. Here, we show that endogenous ubiquitin is acetylated at lysine ( K )‐6 (AcK6) or K 48. Acetylated ubiquitin does not affect substrate monoubiquitylation, but inhibits K 11‐, K 48‐, and K 63‐linked polyubiquitin chain elongation by several E 2 enzymes in vitro . In cells, A cK6‐mimetic ubiquitin stabilizes the monoubiquitylation of histone H 2 B —which we identify as an endogenous substrate of acetylated ubiquitin—and of artificial ubiquitin fusion degradation substrates. These results characterize a mechanism whereby ubiquitin, itself a PTM , is subject to another PTM to modulate mono‐ and polyubiquitylation, thus adding a new regulatory layer to ubiquitin biology.