https://nova.newcastle.edu.au/vital/access/ /manager/Index en-au 5 https://nova.newcastle.edu.au/vital/access/ /manager/Repository/uon:19735 Wed 11 Apr 2018 17:04:44 AEST ]]> https://nova.newcastle.edu.au/vital/access/ /manager/Repository/uon:22109 Wed 11 Apr 2018 16:41:47 AEST ]]> https://nova.newcastle.edu.au/vital/access/ /manager/Repository/uon:22052 Tue 13 Sep 2022 14:25:52 AEST ]]> https://nova.newcastle.edu.au/vital/access/ /manager/Repository/uon:23055 Thu 28 Oct 2021 13:04:24 AEDT ]]> https://nova.newcastle.edu.au/vital/access/ /manager/Repository/uon:35760 Thu 21 Nov 2019 13:03:58 AEDT ]]> https://nova.newcastle.edu.au/vital/access/ /manager/Repository/uon:36904 Thu 16 Jul 2020 12:54:43 AEST ]]> https://nova.newcastle.edu.au/vital/access/ /manager/Repository/uon:13994 Sat 24 Mar 2018 08:20:35 AEDT ]]> https://nova.newcastle.edu.au/vital/access/ /manager/Repository/uon:10706 Sat 24 Mar 2018 08:12:14 AEDT ]]> https://nova.newcastle.edu.au/vital/access/ /manager/Repository/uon:11246 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 α-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.]]> Sat 24 Mar 2018 08:10:55 AEDT ]]> https://nova.newcastle.edu.au/vital/access/ /manager/Repository/uon:18116 2+-mediated activation of the anaphase-promoting complex/cyclosome (APC/C). Although the loss in activity of the M-phase kinase CDK1 is known to be an essential downstream event of this process, the contribution of phosphatases to arrest and meiotic resumption is less apparent, especially in mammals. Therefore, we explored the role of protein phosphatase 2A (PP2A) in mouse eggs using pharmacological inhibition and activation as well as a functionally dominant-negative catalytic PP2A subunit (dn-PP2Ac-L199P) coupled with live cell imaging. We observed that PP2A inhibition using okadaic acid induced events normally observed at fertilization: degradation of the APC/C substrates cyclin B1 and securin resulting from loss of the APC/C inhibitor Emi2. Although sister chromatids separated, chromatin remained condensed, and polar body extrusion was blocked as a result of a rapid spindle disruption, which could be ameliorated by non-degradable cyclin B1, suggesting that spindle integrity was affected by CDK1 loss. Similar cell cycle effects to okadaic acid were also observed using dominant-negative PP2Ac. Preincubation of eggs with the PP2A activator FTY720 could block many of the actions of okadaic acid, including Emi2, cyclin B1, and securin degradation and sister chromatid separation. Therefore, in conclusion, we used okadaic acid, dn-PP2Ac-L199P, and FTY720 on mouse eggs to demonstrate that PP2A is needed to for both continued metaphase arrest and successful exit from meiosis.]]> Sat 24 Mar 2018 08:04:35 AEDT ]]> https://nova.newcastle.edu.au/vital/access/ /manager/Repository/uon:20418 Sat 24 Mar 2018 08:00:53 AEDT ]]>