Cyclin B Dissociation from CDK1 Precedes its Degradation Upon MPF Inactivation in Mitotic Extracts of Xenopus laevis Embryos

 Abstract

Cyclin B is a regulatory subunit of CDK1 within MPF complex. Degradation of cyclin B via ubiquitin-proteasome pathway seemed to be absolutely required for the M-phase exit. However, inhibition of the proteasome proteolytic activity upon the exit from the meiotic metaphase II-arrest in Xenopus cell-free extract revealed that the proteasome-dependent dissociation of cyclin B from CDK1 is sufficient to inactivate MPF without cyclin B degradation. In this study we analyse whether the same mechanism operates during the exit from mitotic M-phase. We show in Xenopus cell-free extract undergoing the first or the second embryonic mitosis that CDK1 oscillations are not affected by proteasome inhibition with MG132 or ALLN despite effective inhibition of cyclins B degradation. The majority of cyclins B1 and B2 surviving CDK1 inactivation is CDK-free and cyclin B2 becomes resistant to phosphatase ? dephosphorylation. The pool of cyclins B remaining after CDK1 inactivation in the presence of MG132 is mitotically inert, while exogenous or newly synthesised cyclin B activates CDK1. This suggests that cyclins B remain sequestered within the proteasome upon MPF inactivation in the presence of MG132. Comparison of the dynamics of the decline of total and CDK-bound pools of cyclins B1, B2 and B4 upon mitotic exit in absence of protein synthesis reveals that CDK-bound cyclins B diminish clearly faster. Our results thus show that cyclin B dissociation from CDK1 precedes cyclins B degradation upon CDK1 inactivation in mitotic embryo extracts and that proteasome proteolytic activity is dispensable for both activation and inactivation of CDK1 in such extracts.

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1687 - 1698
doi
10.4161/cc.5.15.3123
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Cyclin B Dissociation from CDK1 Precedes its Degradation Upon MPF Inactivation in Mitotic Extracts of Xenopus laevis Embryos