Regulation of APC/C activators in mitosis and meiosis

Abstract

The anaphase-promoting complex/cyclosome (APC/C) is a multisubunit E3 ubiquitin ligase that triggers the degradation of multiple substrates during mitosis. Cdc20/Fizzy and Cdh1/Fizzy-related activate the APC/C and confer substrate specificity through complex interactions with both the core APC/C and substrate proteins. The regulation of Cdc20 and Cdh1 is critical for proper APC/C activity and occurs in multiple ways: targeted protein degradation, phosphorylation, and direct binding of inhibitory proteins. During the specialized divisions of meiosis, the activity of the APC/C must be modified to achieve proper chromosome segregation. Recent studies show that one way in which APC/C activity is modified is through the use of meiosis-specific APC/C activators. Furthermore, regulation of the APC/C during meiosis is carried out by both mitotic regulators of the APC/C as well as meiosis-specific regulators. Here, we review the regulation of APC/C activators during mitosis and the role and regulation of the APC/C during female meiosis.

Figure 1. Regulation of Cdc20 and Cdh1 during the Cell Cycle

(top) During S and G2 phases, APC/C^Cdc20 is thought to be bound and inhibited by Emi1. Emi1 is degraded in prometaphase, and APC/C^Cdc20 becomes active against early mitotic APC/C substrates like Cyclin A and Nek2A. In prometaphase APC/C^Cdc20 remains inhibited from targeting Cyclin B, Securin, and other substrates by the spindle assembly checkpoint through direct binding of Mad2, Bub3, and BubR1. Upon release of the spindle assembly checkpoint, APC/C^Cdc20 becomes fully active. In late anaphase and G1, Cdc20 is targeted by APC/C^Cdh1 for degradation by the proteasome. It will not be expressed again until S phase. (bottom) Cdh1 is inhibited from associating with the core APC/C in S, G2, and M phases by phosphorylation by cyclin-dependent kinases. In Drosophila, Rca1 is thought to inhibit Cdh1 in G2, and in vertebrates Emi1 is thought to inhibit APC/C^Cdh1 at the G1-S transition. Recent findings suggest that APC/C^Cdh1 must be inhibited from targeting Securin from degradation by Rae1-Nup98 in metaphase. Dephosphorylation of Cdh1 by Cdc14 phophatase as well as a decrease in mitotic Cdk-cyclin activity leads to loss of inhibitory phosphorylation on Cdh1 in late mitosis and allows for full activation of APC/C^Cdh1 in late mitosis and G1. In addition, Cdh1 may be subject to ubiquitin-mediated degradation in late G1 by APC/C^Cdh1 (indicated by dashed arrow) and in S phase by SCF ubiquitin ligase. See text for references.

Figure 2. Structure and regulation of Emi1 and Emi2/Erp1

(A) The Emi1 protein contains an F-box domain, a C-terminal Zn²⁺ binding region (ZBR), and a D-box. The role of the F-box is unclear, but the D-box is the domain through which Emi1 binds the D-box receptor on the core APC/C, whereas the ZBR domain seems to act to inhibit access of substrates to the complex. See text for details and references. Emi1 is a substrate of the E3 ubiquitin ligase SCF^β-TrCP (Guardavaccaro et al. 2003, Margottin-Goguet et al. 2003). Phosphorylation by both Cdk and Plk1 contribute to recognition and ubiquitination of Emi1 by SCF^β-TrCP in late prophase (Margottin-Goguet et al. 2003, Moshe et al. 2004, Hansen et al. 2004). (B) The Emi2/Erp1 protein also contains an F-box domain, a C-terminal Zn²⁺ binding region (ZBR), and a D-box. Upon fertilization, activated calmodulin kinase II (CaMKII) acts as a priming kinase for Plx1 on Emi2. Phosphorylation by Plx1 targets Emi2 for degradation targeted by SCF^β-TrCP. Phosphorylation of Emi2 by p90^RSK, part of the pathway that establishes CSF arrest, is required for Emi2 stability during oocyte maturation. Phosphorylation of Emi2 by Cdc2 (Cdk1)/CycB may disrupt the interaction between Emi2 and APC/C. See text for details and references.

Figure 3. Meiotic Progression in Females

Oocytes are arrested in prophase I to allow for oocyte growth and differentiation before initiation of the meiotic divisions. Then, oocytes arrest again at different times to await egg activation or fertilization. This secondary arrest ensures that the completion of meiosis is properly coordinated with fertilization of the oocyte. In Drosophila, the secondary meiotic arrest takes place in metaphase I. Egg activation releases this arrest. In most vertebrates the secondary meiotic arrest takes place in metaphase II and is called cytostatic factor (CSF) arrest. CSF is released by fertilization. MPF (CycB/Cdc2 (Cdk1)) activity levels rise upon entry into the meiotic divisions. They must drop in between meiosis I and meiosis II to exit meiosis I, but some activity must be maintained to suppress DNA replication in between the two divisions. MPF levels drop again at exit from meiosis II. See text for details and references.

Figure 4. Meiotic APC/C Regulators

(A) In S. cerevisiae Mnd2 inhibits APC/C^Ama1, a meiosis-specific form of the APC/C, beginning in prophase. Inhibition is not released until late meiosis when Mnd2 protein disappears from meiotic cells. Full activation of APC/C^Ama1 may also require APC/C^Cdc20-mediated degradation of cyclins in meiosis I. (B) In S. pombe Mes1 is an inhibitor of APC/C^Slp1/Cdc20 in between the meiotic divisions. Mes1 itself is an APC/C substrate but is able to compete for binding to the APC/C more efficiently than other substrates. (C) In vertebrates regulation of the APC/C during meiosis has been well-studied. In prophase, prior to germinal vesicle breakdown (GVBD), inhibition of APC/C^Cdh1 by Emi1 allows for an accumulation of MPF that is required for entry into the first meiotic division. Emi1 is targeted for degradation by SCF^β-TrCP at GVBD. APC/C^Cdc20 activity is thought to be regulated by the spindle assembly checkpoint in meiosis I for proper homolog alignment and disjunction. Similar to the role of Mes1, Emi2/Erp1 helps maintain low APC/C^Cdc20 activity levels in between the two meiotic divisions. Emi2/Erp1 is also a main component of CSF arrest and prevents APC/C^Cdc20 from being active until fertilization. The spindle assembly checkpoint may play a role is establishing but not in maintaining CSF arrest. At fertilization, a transient increase in free cytosolic Ca²⁺ activates calmodulin kinase II and the phosphatase calcineurin. Calmodulin kinase II phosphorylates Emi2, which ultimately leads to its degradation by SCF^β-TrCP. Calcineurin dephosphorylates Apc3, a core APC/C subunit, and Cdc20/Fzy, both of which may contribute to activation of APC/C^Cdc20 upon fertilization. See text for references and details.