Basis of catalytic assembly of the mitotic checkpoint complex

Accurate genome inheritance by daughter cells requires that sister chromatids in the mother attach to microtubules emanating from opposite poles of the mitotic spindle (bi-orientation). A surveillance mechanism named the spindle assembly checkpoint (SAC) monitors the microtubule attachment process, temporarily halting sister chromatid separation and mitotic exit until completion of bi-orientation1. SAC failure results in abnormal chromosome numbers (aneuploidy), a hallmark of many tumours. The HORMA domain protein MAD2 is a subunit of the SAC effector mitotic checkpoint complex (MCC). Structural conversion from open to closed MAD2 is required for MAD2 incorporation in MCC1. In vitro, MAD2 conversion and MCC assembly requires several hours2–4, while the SAC response in cells is established in a few minutes5–7. To address this discrepancy, we reconstituted with purified components a near-complete SAC signalling system and monitored MCC assembly with real-time sensors. Dramatic acceleration of MAD2 conversion and MCC assembly was observed when MPS1 phosphorylated the MAD1:MAD2 complex, triggering its template function in the MAD2 conversion and contributing to the establishment of a physical platform for MCC assembly. Thus, catalytic activation of the SAC depends on regulated protein-protein interactions that accelerate the spontaneous but rate-limiting conversion of MAD2 required for MCC assembly.