KEN-dependent degradation of centromere protein F

Introduction Successful completion of chromosome segregation and cytokinesis requires the coordinated activation and inactivation of numerous mitotic regulators. Inactivation is largely brought about by ubiquitindependent proteolysis, with the relevant E3 ubiquitin ligase being the anaphase promoting complex, or cyclosome (APC/C) (Irniger et al., 1995; King et al., 1995; Sudakin et al., 1995). During the early stages of mitosis, the APC/C targets cyclin A and Nek2A for destruction (den Elzen and Pines, 2001; Di Fiore and Pines, 2007; Hames et al., 2001; Pines and Hunter, 1991). Following chromosome alignment, securin and cyclin B1 are then targeted for degradation, thereby triggering anaphase onset and mitotic exit, respectively (Clute and Pines, 1999; Hagting et al., 2002). After anaphase onset, a number of other APC/C substrates are destroyed including the Aurora kinases, Plk1, annilin, geminin, Skp2 and Tome-1 (Peters, 2006). The APC/C is activated by two cofactors, Cdc20 and Cdh1. Destruction of pre-anaphase substrates is Cdc20-dependent. Cdc20 is itself an APC/C substrate that is turned over during mitosis when the spindle checkpoint is active (Nilsson et al., 2008). However, once the spindle checkpoint has become satisfied and anaphase initiated, APC/C activation switches from Cdc20 to Cdh1 (Peters, 2006). This switch occurs because prior to anaphase, Cdh1 is inhibited by Cdk1-dependent phosphorylation; following cyclin B1 destruction, Cdk1 activity declines thereby alleviating inhibition of Cdh1 by promoting its association with the APC/C (Kramer et al., 2000; Visintin et al., 1998; Zachariae et al., 1998). This not only initiates Cdh1-activated APC/C (APC/CCdh1)-mediated degradation but, because Cdc20 is itself a APC/CCdh1 substrate, it also inhibits Cdc20-dependent degradation (Huang et al., 2001). Degradation of Cdc20 substrates is essential for mitotic progression; however, the significance of Cdh1-dependent proteolysis is less clear. Although degradation of Aurora A, Aurora B and Plk1 might facilitate efficient spindle reorganisation and timely cytokinesis (Floyd et al., 2008; Lindon and Pines, 2004), it is striking that, following rescue of placental defects, Cdh1-null mouse embryos develop relatively normally and survive to birth (Garcia-Higuera et al., 2008; Li et al., 2008). Indeed, it was recently shown that despite extensive RNAi-mediated repression of Cdh1, several presumed Cdh1 substrates (including Plk1, Cdc20 and survivin) were degraded normally (Floyd et al., 2008); Aurora A and B were the only substrates stabilised following Cdh1 RNAi. Centromere protein F (Cenp-F) is a kinetochore protein required for chromosome segregation (Bomont et al., 2005; Holt et al., 2005; Laoukili et al., 2005; Yang et al., 2005). Pulse-chase experiments conducted over 10 years ago showed that Cenp-F is degraded at some time during or after mitosis (Liao et al., 1995). However, exactly when Cenp-F is degraded, the underlying mechanism, and the functional significance of its proteolysis are all unresolved issues. Overexpression of Cdh1 suppresses Cenp-F expression (Sorensen Cdc20 is required for the post-anaphase, KEN-dependent degradation of centromere protein F