PKMYT1 has an important role in the timing and fidelity of chromosome segregation

Abstract Mitotic entry is a tightly regulated process controlled by CDK1-Cyclin B1. Its activity is negatively regulated by the WEE family kinases, WEE1, WEE2 and PKMYT1, to prevent premature mitotic entry. While WEE1 has an established role in regulating CDK2 during S-phase, the full scope of PKMYT1’s roles in the cell cycle is surprisingly unexplored. Here, we show that PKMYT1 activity increases during anaphase and we reveal a novel mitotic function for PKMYT1 that is distinct from its family member WEE1. Chemical inhibition of PKMYT1 induces premature anaphase, leading to chromosome segregation errors. These errors, including chromatin bridges and micronuclei, consequently activate the cGAS–STING pathway. We further demonstrate that PKMYT1 contributes to maintaining spindle assembly checkpoint integrity, as its inhibition promotes mitotic slippage in the presence of anti-microtubule drugs. Our findings reveal that PKMYT1 provides an additional regulatory mechanism, acting alongside Cyclin B1 degradation, to control CDK1-Cyclin B1 activity and ensure the fidelity of the metaphase-to-anaphase transition.