| After fertilization, Drosophila embryos undergo thirteen rounds of synchronous mitosis without cytokinesis, generating about six thousand nuclei in a common cytoplasm. At this point, embryos pause in the interphase of cycle 14 for about 1 hour and cellularize. The transition from the early synchronous division to a cell cycle pause accompanied by morphogenetic change is known as mid-blastula transition (MBT). The onset of the MBT is controlled monitoring the nucleocytoplasmic (N/C) ratio. Using compound chromosomes to manipulate DNA content of the embryos, I have determined the threshold for the N/C ratio is about 70% of the DNA content normally present at cycle 14. Embryos with DNA contents around this value show intermediate cell cycle behaviors. Some pause at cycle 14, some at cycle 15 and some form patches arrested in different mitotic cycles. Live imaging analysis suggested that the decision of cell cycle pause is made collectively by neighboring nuclei through communication at the MBT.;Another prominent feature of MBT is a massive increase in zygotic transcription and a parallel degradation of maternally supplied RNAs. To determine whether these changes in gene expression are governed by the same N/C ratio that controls cell cycle pause, I compared gene expression in haploid and diploid Drosophila embryos. I found that most maternal RNA degradation and most new transcription correlate with absolute time or developmental stage, and are timed independently of N/C ratio. I did however identify a class of zygotically active genes whose expression depends on the N/C ratio and which is only expressed at cycle 15 in haploids. In embryos with patchy cell cycle behavior due to their threshold DNA content, expression of these genes correlates tightly with the boundaries of mitotic patches, suggesting that the mechanism that measures the N/C ratio and pauses the mitotic cycle is the same, or tightly coupled to the mechanism controlling zygotic transcription of N/C ratio genes at the MBT.;During MBT, embryos monitor cell cycle through regulation of levels of Cdc25, a phosphatase that activates Cdc2. Using the UAS/Gal4 system to drive exogenous expression of fly homologs of cdc25, string and twine, we have shown that the two genes are not only regulated at RNA level, but also at protein level during the MBT. Coincidently, cellularization, the morphogenetic event at the MBT, occurs at the same time as the degradation of cdc25 and the cell cycle pause. The coincidence suggests a coupling mechanism to ensure proper timing for different events at the MBT. However, results from cellularization mutants showed no direct checkpoint on cellularization in the regulation of cdc25 levels, suggesting an indirect regulatory mechanism. Since both cellularization and degradation of string require zygotic transcriptions, we are in search of the candidate zygotic genes involved in regulating the two processes by deficiency screen. |
