Tung JJ, Hansen DV, Ban KH, Loktev AV, Summers MK, Adler III JR et al. A role for the anaphase-promoting complex inhibitor Emi2/XErp1, a homolog of early mitotic inhibitor 1, in cytostatic factor arrest of Xenopus eggs. Proc Natl Acad Sci USA 102: 4318-4323

Department of Pathology, Stanford University School of Medicine, 300 Pasteur Drive, Stanford, CA 94305, USA.
Proceedings of the National Academy of Sciences (Impact Factor: 9.67). 04/2005; 102(12):4318-23. DOI: 10.1073/pnas.0501108102
Source: PubMed


Unfertilized vertebrate eggs are arrested in metaphase of meiosis II with high cyclin B/Cdc2 activity to prevent parthenogenesis. Until fertilization, exit from metaphase is blocked by an activity called cytostatic factor (CSF), which stabilizes cyclin B by inhibiting the anaphase-promoting complex (APC) ubiquitin ligase. The APC inhibitor early mitotic inhibitor 1 (Emi1) was recently found to be required for maintenance of CSF arrest. We show here that exogenous Emi1 is unstable in CSF-arrested Xenopus eggs and is destroyed by the SCF(betaTrCP) ubiquitin ligase, suggesting that endogenous Emi1, an apparent 44-kDa protein, requires a stabilizing factor. However, anti-Emi1 antibodies crossreact with native Emi2/Erp1/FBXO43, a homolog of Emi1 and conserved APC inhibitor. Emi2 is stable in CSF-arrested eggs, is sufficient to prevent CSF release, and is rapidly degraded in a Polo-like kinase 1-dependent manner in response to calcium-mediated egg activation. These results identify Emi2 as a candidate CSF maintenance protein.

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Available from: David V Hansen, May 08, 2014
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    • "Metaphase II-CSF in vertebrates (reviewed by Masui, 2000; Kishimoto, 2003) is now known to involve the Mos/MEK/MAPK kinase cascade and activity of the Cdc20-activated anaphase promoting complex/cyclosome (APC/C cdc20 ) inhibitor Emi2, also known as Erp1 in Xenopus (Schmidt et al., 2005; Tung et al., 2005; Shoji et al., 2006; reviewed by Wu and Kornbluth, 2008). "
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    ABSTRACT: The fertilising sperm triggers a transient Ca(2+) increase that releases eggs from cell cycle arrest in the vast majority of animal eggs. In vertebrate eggs, Erp1, an APC/C(cdc20) inhibitor, links release from metaphase II arrest with the Ca(2+) transient and its degradation is triggered by the Ca(2+)-induced activation of CaMKII. By contrast, many invertebrate groups have mature eggs that arrest at metaphase I, and these species do not possess the CaMKII target Erp1 in their genomes. As a consequence, it is unknown exactly how cell cycle arrest at metaphase I is achieved and how the fertilisation Ca(2+) transient overcomes the arrest in the vast majority of animal species. Using live-cell imaging with a novel cyclin reporter to study cell cycle arrest and its release in urochordate ascidians, the closest living invertebrate group to the vertebrates, we have identified a new signalling pathway for cell cycle resumption in which CaMKII plays no part. Instead, we find that the Ca(2+)-activated phosphatase calcineurin (CN) is required for egg activation. Moreover, we demonstrate that parthenogenetic activation of metaphase I-arrested eggs by MEK inhibition, independent of a Ca(2+) increase, requires the activity of a second egg phosphatase: PP2A. Furthermore, PP2A activity, together with CN, is required for normal egg activation during fertilisation. As ascidians are a sister group of the vertebrates, we discuss these findings in relation to cell cycle arrest and egg activation in chordates.
    Full-text · Article · Nov 2013 · Development
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    • "Extracts prepared from these eggs are called CSF-arrested extracts. At fertilization, a transient increase in cytoplasmic calcium triggers APC/C activation by degrading the APC/C inhibitor Erp1/Emi2 and activating calcineurin (Liu and Maller, 2005; Rauh et al, 2005; Tung et al, 2005; Mochida and Hunt, 2007). Hence, addition of calcium into CSF extracts degrades APC/C substrates such as cyclin B and securin and causes exit to interphase, whereas in the absence of calcium the activity of CDK in CSF extracts is high and progress to interphase prevented. "
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    ABSTRACT: The anaphase-promoting complex/cyclosome (APC/C) ubiquitin ligase is tightly regulated to ensure programmed proteolysis in cells. The activity of the APC/C is positively controlled by cyclin-dependent kinase (CDK), but a second level of control must also exist because phosphorylation inactivates Cdc20, a mitotic APC/C co-activator. How Cdc20 is dephosphorylated specifically, when CDK is high, has remained unexplained. Here, we show that phosphatases are crucial to activate the APC/C. Cdc20 is phosphorylated at six conserved residues (S50/T64/T68/T79/S114/S165) by CDK in Xenopus egg extracts. When all the threonine residues are phosphorylated, Cdc20 binding to and activation of the APC/C are inhibited. Their dephosphorylation is regulated depending on the sites and protein phosphatase 2A, active in mitosis, is essential to dephosphorylate the threonine residues and activate the APC/C. Consistently, most of the Cdc20 bound to the APC/C in anaphase evades phosphorylation at T79. Furthermore, we show that the 'activation domain' of Cdc20 associates with the Apc6 and Apc8 core subunits. Our data suggest that dephosphorylation of Cdc20 is required for its loading and activation of the APC/C ubiquitin ligase.
    Full-text · Article · Jun 2012 · The EMBO Journal
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    • "Finally, both UVI4 and Emi1 seem to be required for the repression of APC/C activity during the S-to-G2 transition, coupling DNA replication with mitosis, whereby the E2F transcription factors seemingly regulate the timing of activity of both genes (Hsu et al., 2002). UVI4 and Emi1 are not only structurally and functionally related, but also both possess a homolog that operates specifically in the meiotic cell cycle, namely OSD1 and Erp1/Emi2, respectively (Tung et al., 2005; Ohe et al., 2007; d'Erfurth et al., 2009), that are both essential for the transition from meiosis I to meiosis II (Ohe et al., 2007; d'Erfurth et al., 2010). In Xenopus, this transition occurs through a stabilization of CYCB by Erp1/Emi2 (Ohe et al., 2007), whereas the targets of OSD1 remain to be identified. "
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    ABSTRACT: The anaphase-promoting complex/cyclosome (APC/C) is a multisubunit ubiquitin ligase that regulates progression through the cell cycle by marking key cell division proteins for destruction. To ensure correct cell cycle progression, accurate timing of APC/C activity is important, which is obtained through its association with both activating and inhibitory subunits. However, although the APC/C is highly conserved among eukaryotes, no APC/C inhibitors are known in plants. Recently, we have identified ULTRAVIOLET-B-INSENSITIVE4 (UVI4) as a plant-specific component of the APC/C. Here, we demonstrate that UVI4 uses conserved APC/C interaction motifs to counteract the activity of the CELL CYCLE SWITCH52 A1 (CCS52A1) activator subunit, inhibiting the turnover of the A-type cyclin CYCA2;3. UVI4 is expressed in an S phase-dependent fashion, likely through the action of E2F transcription factors. Correspondingly, uvi4 mutant plants failed to accumulate CYCA2;3 during the S phase and prematurely exited the cell cycle, triggering the onset of the endocycle. We conclude that UVI4 regulates the temporal inactivation of APC/C during DNA replication, allowing CYCA2;3 to accumulate above the level required for entering mitosis, and thereby regulates the meristem size and plant growth rate.
    Full-text · Article · Dec 2011 · The Plant Cell
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