GSK3β mediates acentromeric spindle stabilization by activated PKCζ

Upon fertilization, the mammalian egg undergoes a precise series of signaling events that orchestrate its conversion into a zygote. Mouse eggs contain acentrosomal spindle poles when arrested at meiotic metaphase II. The meiotic spindle is thought to provide a scaffold that mediates spatial and temporal regulation of the signaling pathways orchestrating post-fertilization events. Many kinases have been found to be enriched at the MII meiotic spindle, such as Protein Kinase C (PKC), and are thought to have an important role in regulating signaling events initiated through fertilization. In this study phosphorylated PKC zeta (p-PKC zeta) and Glycogen Synthase Kinase 3 beta (GSK3 beta) were found to be enriched at both acentrosomal spindle poles and the kinetochore region. Phosphorylated PKC zeta (p-PKC zeta) was immunopurified from MII eggs and was found to co-localize with known microtubule stabilizing components found in somatic cells, including GSK3 beta and Partition deficit protein 6 (Par6). Both fluorescence resonance energy transfer (FRET) and immunofluorescence confirmed the existence and close association of these proteins with p-PKC zeta at the meiotic spindle. When GSK3 zeta is phosphorylated on ser9 its activity is inhibited and the spindle is stabilized. However, when GSK3 beta is dephosphorylated (on ser9) it becomes active and the spindle is destabilized. The mechanism by which p-PKC zeta maintains spindle organization appears to be through GSK3 beta and suggests that p-PKC zeta phosphorylates GSK3 beta on the ser9 position inactivating GSK3 beta and consequently maintaining spindle stability during meiotic metaphase arrest. (C) 2008 Elsevier Inc. All rights reserved.