UBE2S elongates ubiquitin chains on APC/C substrates to promote mitotic exit

The anaphase-promoting complex (APC/C) ubiquitin ligase is the target of the spindle-assembly checkpoint (SAC), ubiquitylating protein substrates whose degradation regulates progress through mitosis1-3. The identity of the ubiquitin-conjugating (E2) enzymes that work with the APC/C is unclear. In an RNA interference screen for factors that modify release from drug-induced SAC activation, we identify here the E2 enzyme, UBE2S, as an auxillary factor for the APC/C that promotes mitotic exit. UBE2S is dispensable in a normal mitosis, but its depletion prolongs drug-induced mitotic arrest and suppresses mitotic slippage. In vitro, UBE2S elongates ubiquitin chains initiated by the E2 enzymes UBCH10 and UBCH5, enhancing the degradation of APC/C substrates by the proteasome. Indeed, following release from SAC arrest, UBE2S-depleted cells neither degrade crucial APC/C substrates, nor silence this checkpoint, whereas SAC bypass via BUBR1 depletion or Aurora-B inhibition negates the requirement for UBE2S. Thus, UBE2S acts with the APC/C in a two-step mechanism controlling substrate ubiquitylation that is essential for mitotic exit after prolonged SAC activation, providing a new model for APC/C function in human cells.