Greatwall is essential to prevent mitotic collapse after nuclear envelope breakdown in mammals

Abstract

Significance Nuclear envelope breakdown (NEB) leads to the exposure of nuclear structures to cytoplasmic activities. Greatwall is a kinase able to inhibit PP2A phosphatases that counteract Cdk-dependent phosphorylation required for mitosis. Here we show that Greatwall, an essential protein in mammals, is exported to the cytoplasm in a Cdk-dependent manner before NEB, thus protecting mitotic phosphates from phosphatase activity.

Regulation of Greatwall kinase by protein stabilization and nuclear localization

“…This finding is in agreement with previous studies in human and Drosophila that reported nuclear export of Gwl before nuclear membrane breakdown. 15,20 However, distinct from the previous study that suggested a requirement of Gwl kinase activity for its nuclear export, 20 we found a similar pattern of nucleocytoplasmic relocalization for G41S Gwl prior to mitotic entry (Fig. 4B).…”

“…15,20 We also observed in Xenopus cells that nuclear Gwl started to diffuse into cytoplasm before centrosomes reached opposite positions. While it remained unclear how this nucleocytoplasmic translocation of Gwl may contribute to the regulation of mitotic entry, previous studies indicated that the translocation was dependent on Gwl phosphorylation by CDK1, and possibly also Plk1.…”

“…15 However, neither of these NLS motifs in Drosophila is conserved in vertebrates. The NLS of human Gwl was found around KR444/445, 20 the corresponding residues of KR456/457 in Xenopus Gwl. Thus, the NLS of Xenopus Gwl, as identified in this study, represents a well-conserved mechanism in vertebrate animals.…”

“…15,20 The Drosophila homolog of Gwl contains 2 essential NLS motifs within the central region, mutation of which disrupted its nuclear location. 15 However, neither of these NLS motifs in Drosophila is conserved in vertebrates.…”

“…While it remained unclear how this nucleocytoplasmic translocation of Gwl may contribute to the regulation of mitotic entry, previous studies indicated that the translocation was dependent on Gwl phosphorylation by CDK1, and possibly also Plk1. 15,20 It was also suggested that the kinase activity of Gwl was required for its nuclear export as a kinasedead form of Gwl harboring a mutation at the ATP-binding site was deficient in nuclear export. 20 However, a different form of kinase-dead Gwl used in our study exhibited a similar pattern of nucleocytoplasmic translocation as the WT.…”