Nucleo-cytoplasmic interactions that control nuclear envelope breakdown and entry into mitosis in the sea urchin zygote

In sea urchin zygotes and mammalian cells nuclear envelope breakdown (NEB) is not driven simply by a rise in cytoplasmic cyclin dependent kinase l-cyc lin B (Cdk1-B) activity; the checkpoint monitoring DNA synthesis can preven t NEB in the face of mitotic levels of Cdk1-B, Using sea urchin zygotes we investigated whether this checkpoint prevents NEB by restricting import of regulatory proteins into the nucleus, We find that cyclin B1-GFP accumulate s in nuclei that cannot complete DNA synthesis and do not break down, Thus, this checkpoint limits NEB downstream of both the cytoplasmic activation a nd nuclear accumulation of Cdk1-B1. In separate experiments we fertilize se a urchin eggs with sperm whose DNA has been covalently cross-linked to inhi bit replication, When the pronuclei fuse, the resulting zygote nucleus does not break down for >180 minutes (equivalent to three cell cycles), even th ough Cdk1-B activity rises to greater than mitotic levels, If pronuclear fu sion is prevented, then the female pronucleus breaks down at the normal tim e (average 68 minutes) and the male pronucleus with cross-linked DNA breaks down 16 minutes later, This male pronucleus has a functional checkpoint be cause it does not break down for >120 minutes if the female pronucleus is r emoved just prior to NEB, These results reveal the existence of an activity released by the female pronucleus upon its breakdown, that overrides the c heckpoint in the male pronucleus and induces NEB, Microinjecting wheat germ agglutinin into binucleate zygotes reveals that this activity involves mol ecules that must be actively translocated into the male pronucleus.