Spatial regulation of APCCdh1-induced cyclin B1 degradation maintains G2 arrest in mouse oocytes

Abstract

SUMMARYWithin the mammalian ovary, oocytes remain arrested at G2 for several years. Then a peri-ovulatory hormonal cue triggers meiotic resumption by releasing an inhibitory phosphorylation on the kinase Cdk1. G2 arrest, however, also requires control in the concentrations of the Cdk1-binding partner cyclin B1, a process achieved by anaphase-promoting complex (APC Cdh1 ) activity, which ubiquitylates and so targets cyclin B1 for degradation. Thus, APC Cdh1 activity prevents precocious meiotic entry by promoting cyclin B1 degradation. However, it remains unresolved how cyclin B1 levels are suppressed sufficiently to maintain arrest but not so low that they make oocytes hormonally insensitive. Here, we examined spatial control of this process by determining the intracellular location of the proteins involved and using nuclear-targeted cyclin B1. We found that raising nuclear cyclin B1 concentrations, an event normally observed in the minutes before nuclear envelope breakdown, was a very effective method of inducing the G2/M transition. Oocytes expressed only the a-isoform of Cdh1, which was predominantly nuclear, as were Cdc27 and Psmd11, core components of the APC and the 26S proteasome, respectively. Furthermore, APC Cdh1 activity appeared higher in the nucleus, as nuclear-targeted cyclin B1 was degraded at twice the rate of wild-type cyclin B1. We propose a simple spatial model of G2 arrest in which nuclear APC Cdh1 -proteasomal activity guards against any cyclin B1 accumulation mediated by nuclear import.

Changing expression and subcellular distribution of karyopherins during murine oogenesis

“…To maintain meiotic arrest until the appropriate hormonal cues at ovulation, nuclear maturation promoting factor activity (a dimer of CDK1-cyclin B1) is suppressed by the nuclear presence of CDK1-inactivating kinase WEE1B, nuclear exclusion of CDK1-activating phosphatase CDC25 and nuclear activity of the E3 ubiquitin ligase complex anaphase promoting complex/Fizzy related 1, which promotes degradation of cyclin B1 (Holt et al 2010, Oh et al 2010. The spatial rearrangement of these cell cycle regulators at the onset of meiotic resumption could be regulated by KPNA-mediated nucleo-cytoplasmic shuttling.…”

The APC/C in female mammalian meiosis I

“…Conversely, Emi1 and securin serve to limit such proteolysis, thereby allowing cyclin B1 accumulation and Cdk1 activation required for GVBD (Marangos et al 2007, Marangos & Carroll 2008. Influx of cyclin B1 into the nuclear interior around the time of GVBD is thought to overwhelm nuclearconcentrated APC/C Cdh1 , thereby promoting further cyclin B1 accumulation (Holt et al 2010). Recent data also indicate that cyclin B2 is also important for Cdk1 activation at GVBD and that Hec1 sustains cyclin B2 levels by sequestering it away from APC/C Cdh1 (Gui & Homer 2013).…”

“…Recent evidence indicates that unlike mitosis, G2 arrest in oocytes is very sensitive to intra-nuclear cyclin B1 levels. Thus, a nuclear-targeting cyclin B1 construct markedly increases GVBD in oocytes (Holt et al 2010) whereas a similar construct has little effect on the timing of entry APC/C in female mammalian meiosis I R65 into mitosis (Hagting et al 1998). Consequently, it is especially critical to maintain low cyclin B1 levels within the interior of the GV.…”

“…It turns out that in order to accomplish this, one strategy is to spatially enrich components of the APC/C proteolytic pathway within the GV including the exclusive expression of the a splice variant of Cdh1, which is targeted to the nuclear interior, and not the cytoplasmic b variant (widely expressed in other cell types) (Holt et al 2010). It has been proposed that the primary function of spatially enriched proteolysis within the GV is to maintain the G2/prophase arrest state by maintaining nuclear cyclin at low levels (Holt et al 2010). It is also possible that compartmentalised proteolysis, by helping to preserve a large cytoplasmic reservoir of cyclin B, might assist in averting refractoriness.…”

The APC/C activator FZR1 coordinates the timing of meiotic resumption during prophase I arrest in mammalian oocytes

“…Cyclin B1 overexpression in fully grown oocytes cultured in media designed to maintain high cAMP levels has the ability to induce GVB (Ledan et al, 2001;Marangos and Carroll, 2004;Holt et al, 2010). Oocytes are therefore likely to have a mechanism to prevent cyclin B1 accumulation, and a number of recent studies in mouse have suggested that this is dependent on FZR1, an activator of the anaphase-promoting complex/cyclosome (APC/C) (Reis et al, 2006a;Marangos et al, 2007;Marangos and Carroll, 2008;Homer et al, 2009;Schindler and Schultz, 2009;Holt et al, 2010).…”