Regulation of the embryonic onset of late DNA replication in Drosophila melanogaster

In Drosophila embryos, the mid-blastula transition (MBT) dramatically remodels the cell cycle during interphase 14. Before the MBT, each cycle is composed of only a short S-phase and mitosis. At the MBT, S-phase is dramatically lengthened by the onset of late replication, and a G2 phase is introduced. These changes set the stage for morphogenesis, including the cellularization of the syncytial embryo and the onset of gastrulation. The introduction of the G2 phase was known to result from the downregulation of Cdc25 phosphatase and the inactivation of Cdk1 kinase; however, the signals modifying S-phase were unknown. We injected cdc25 mRNA to bypass the developmentally programmed downregulation of Cdc25 at the MBT. Introduction of either Cdc25 isoform (String or Twine) or enhanced Cdk1 activity triggered premature replication of late-replicating sequences, even after their specification, and thereby shortened S-phase. Reciprocally, reduction of Cdk1 activity by knockdown of mitotic cyclins extended pre-MBT S-phase. These findings suggest that high Cdc25 and Cdk1 contribute to the speed of the rapid, pre-MBT S-phases.Previous reports suggested that downregulation of Cdc25 activity at the MBT resulted from elimination of cdc25 transcripts. However, premature removal of cdc25 transcripts by RNAi did not affect progression to the MBT. Instead, a new antibody against the Cdc25 isoform Twine showed that Twine protein was abundant and stable until the MBT, when it was destabilized and rapidly eliminated. Twine protein destruction was timed by the nucleo-cytoplasmic ratio and depended on the activation of zygotic transcription at the MBT, including expression of the gene tribbles, whose activity was sufficient to trigger Twine destruction and was required for prompt Twine disappearance.Thus, we propose that the nucleo-cytoplasmic ratio triggers the remodeling of the cell cycle at the MBT by triggering the onset of transcription of a group of genes that inactivate Cdk1 (including through the destruction of Twine), which results in the lengthening of S-phase and addition of the G2 phase