Aurora B/C-dependent phosphorylation promotes Rec8 cleavage in mammalian oocytes

To generate haploid gametes, cohesin is removed in a stepwise manner from chromosome arms in meiosis I and the centromere region in meiosis II to segregate chromosomes and sister chromatids, respectively. Meiotic cohesin removal requires cleavage of the meiosis-specific kleisin subunit Rec8 by the protease separase.(1,2) In yeast and C. elegans, Rec8 on chromosome arms has to be phosphorylated to be cleaved in meiosis I,(3-7) whereas Rec8 at the centromere is protected from cleavage by the action of PP2A-B56.(8-10) However, in mammalian meiosis, it is unknown whether Rec8 has to be equally phosphorylated for cleavage, and if so, the identity of the relevant kinase(s). This is due to technical challenges, as Rec8 is poorly conserved, preventing a direct translation of the knowledge gained from model systems such as yeast and C. elegans to mammals. Additionally, there is no turnover of Rec8 after cohesion establishment, preventing phosphomutant analysis of functional Rec8. To address the very basic question of whether Rec8 cleavage requires its phosphorylation in mammals, we adapted a biosensor that detects separase activity to study Rec8 cleavage in single mouse oocytes by live imaging. Crucially, through phosphomutant analysis, we identified phosphorylation sites in Rec8 promoting cleavage. We found that Rec8 cleavage depends on Aurora B/C kinase activities and identified an aminoacid residue that is phosphorylated in vivo. Accordingly, inhibition of Aurora B/C kinases during meiotic maturation impairs endogenous Rec8 phosphorylation and chromosome segregation.

Automated summary

To separate chromosomes and sister chromatids in meiosis, cohesin needs to be removed from chromosome arms and centromere region. While phosphorylation of Rec8 is known to be necessary for its cleavage in yeast and C. elegans, it remains unclear whether this holds true in mammals. This study used live imaging and phosphomutant analysis to show that phosphorylation of Rec8 by Aurora B/C kinases is required for its cleavage in mammalian meiosis.