Activating the DNA damage checkpoint in a developmental context.

Abstract

Studies in unicellular systems have established that DNA damage by irradiation invokes a checkpoint that acts to stall cell division. During metazoan development, the modulation of cell division by checkpoints must occur in the context of gastrulation, differential gene expression and changes in cell cycle regulation. To understand the effects of checkpoint activation in a developmental context, we examined the effect of X-rays on post-blastoderm embryos of Drosophila melanogaster. In Drosophila, DNA damage was previously found to delay anaphase chromosome separation during cleavage cycles that lack a G2 phase. In post-blastoderm cycles that included a G2 phase, we found that irradiation delayed the entry into mitosis. Gastrulation and the developmental program of string (Cdc25) gene expression, which normally regulates the timing of mitosis, occurred normally after irradiation. The radiation-induced delay of mitosis accompanied the exclusion of mitotic cyclins from the nucleus. Furthermore, a mutant form of the mitotic kinase Cdk1 that cannot be inhibited by phosphorylation drove a mitotic cyclin into the nucleus and overcame the delay of mitosis induced by irradiation. Developmental changes in the cell cycle, for example, the introduction of a G2 phase, dictate the response to checkpoint activation, for example, delaying mitosis instead of or in addition to delaying anaphase. This unprecedented finding suggests that different mechanisms are used at different points during metazoan development to stall cell division in response to checkpoint activation. The delay of mitosis in post-blastoderm embryos is due primarily to inhibitory phosphorylation of Cdk1, whereas nuclear exclusion of a cyclin-Cdk1 complex might play a secondary role. Delaying cell division has little effect on gastrulation and developmentally regulated string gene expression, supporting the view that development generally dictates cell proliferation and not vice versa.