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First published online 23 November 2004
doi: 10.1242/jcs.01567

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Degradation of APC^cdc20 and APC^cdh1 substrates during the second meiotic division in mouse eggs

Cell and Developmental Physiology Research Group, Institute for Cell and Molecular Biosciences, The Medical School, Framlington Place, University of Newcastle, Newcastle, NE2 4HH, UK

^* Author for correspondence (e-mail: k.t.jones{at}newcastle.ac.uk)

Accepted 29 September 2004

Metaphase II-arrested mouse eggs are stimulated to complete meiosis by sperm-induced Ca²⁺ spiking. The Ca²⁺ signal causes activation of the E3 ligase anaphase-promoting complex/cyclosome (APC), leading to the destruction of key proteins necessary for meiotic exit. We show, using western blots of mouse eggs, the presence of both APC activators cdc20 and cdh1, which target D-box and D-box/KEN-box substrates, respectively, for proteolysis. We decided to examine the temporal activation of APC^cdc20 and APC^cdh1 by coupling APC substrates to GFP and examining their destruction in real-time following release from second meiotic division arrest. D-box substrates were degraded quickly after the initiation of sperm-induced Ca²⁺ spiking, such that their degradation was complete by the time of second polar body extrusion. By contrast, KEN-box-containing substrates were degraded when CDK1 activity was low, during the period between polar body extrusion and pronucleus formation. This observation of apparent APC^cdh1 activity in meiosis II based on destruction of exogenous GFP-coupled substrates was then confirmed by observing destruction of endogenous APC^cdh1 substrates. These data are consistent with a model of initial APC^cdc20 activation on sperm-induced activation, followed by APC^cdh1 activation after second polar body extrusion. Interestingly, therefore, we propose that mammalian eggs undergo meiosis II with both APC^cdc20 and APC^cdh1, whereas eggs of other species so far described have APC^cdc20 activity only.

Key words: Calcium, Calcium signalling, Cell cycle proteins, Fertilization, Mammals, Meiosis, Metaphase, Mice, Oocytes, Signal transduction, Ubiquitin-protein ligases

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