Cells of a given type maintain a characteristic cell size to function efficiently in their ecological or organismal context. They achieve this through the regulation of growth rates or by actively sensing size and coupling this signal to cell division. We focus this review on potential size-sensing mechanisms, including geometric, external cue, and titration mechanisms. Mechanisms that titrate proteins against DNA are of particular interest because they are consistent with the robust correlation of DNA content and cell size. We review the literature, which suggests that titration mechanisms may underlie cell-size sensing in Xenopus embryos, budding yeast, and Escherichia coli, whereas alternative mechanisms may function in fission yeast.
During early development, animal embryos depend on maternally deposited RNA until zygotic genes become transcriptionally active. Before this maternal-to-zygotic transition, many species execute rapid and synchronous cell divisions without growth phases or cell cycle checkpoints. The coordinated onset of transcription, cell cycle lengthening, and cell cycle checkpoints comprise the midblastula transition (MBT). A long-standing model in the frog, Xenopus laevis, posits that MBT timing is controlled by a maternally loaded inhibitory factor that is titrated against the exponentially increasing amount of DNA. To identify MBT regulators, we developed an assay using Xenopus egg extract that recapitulates the activation of transcription only above the DNA-to-cytoplasm ratio found in embryos at the MBT. We used this system to biochemically purify factors responsible for inhibiting transcription below the threshold DNA-to-cytoplasm ratio. This unbiased approach identified histones H3 and H4 as concentration-dependent inhibitory factors. Addition or depletion of H3/H4 from the extract quantitatively shifted the amount of DNA required for transcriptional activation in vitro. Moreover, reduction of H3 protein in embryos induced premature transcriptional activation and cell cycle lengthening, and the addition of H3/H4 shortened post-MBT cell cycles. Our observations support a model for MBT regulation by DNA-based titration and suggest that depletion of free histones regulates the MBT. More broadly, our work shows how a constant concentration DNA binding molecule can effectively measure the amount of cytoplasm per genome to coordinate division, growth, and development.early vertebrate development | systems biology | cell size control | maternal zygotic transition | transcription activation T he midblastula transition (MBT) is the first major developmental transition in many fast-developing organisms. Before the MBT, proteins encoded by maternally supplied RNAs drive the early, synchronous cell divisions of the embryo. At the MBT, the cell cycles lengthen to incorporate growth phases, and the zygotic genome becomes transcriptionally active (1, 2). In addition, the MBT is often associated with the emergence of cell cycle checkpoints, cell motility, and in Drosophila, cellularization of the syncytium.The onset of the MBT during early development has been attributed to two general classes of mechanisms. The first class posits that embryos monitor the amount of time since fertilization or count the number of divisions. The second class posits that embryos monitor changes in the DNA-to-cytoplasm ratio in individual cells. Before the MBT, synchronously dividing cells of the embryo do not grow, and thus, the DNA-to-cytoplasm ratio doubles as the cell volume is halved with each division. In D. melanogaster, some genes respond to the timer mechanism, and other genes, including those that control cell cycle lengthening and syncytium cellularization, respond to the DNA-to-cytoplasm ratio (3-7). In Xenopus, the degradation of the cell cycl...
The early embryos of many animals, including flies, fish and frogs, have unusually rapid cell cycles and delayed onset of transcription. These divisions are dependent on maternally supplied RNAs and proteins including histones. Previous work suggests that the pool size of maternally provided histones can alter the timing of zygotic genome activation (ZGA) in frogs and fish. Here, we examine the effects of under-and overexpression of maternal histones in Drosophila embryogenesis. Decreasing histone concentration advances zygotic transcription, cell cycle elongation, Chk1 activation and gastrulation. Conversely, increasing histone concentration delays transcription and results in an additional nuclear cycle before gastrulation. Numerous zygotic transcripts are sensitive to histone concentration, and the promoters of histone-sensitive genes are associated with specific chromatin features linked to increased histone turnover. These include enrichment of the pioneer transcription factor Zelda, and lack of SIN3A and associated histone deacetylases. Our findings uncover a crucial regulatory role for histone concentrations in ZGA of Drosophila.
Highlights d Most nuclear histone H3 is not DNA-bound in the early cleavage cycles d Later cycles lose free nuclear H3 due to titration of maternal stores d H3 on chromatin decreases with each early cycle d H3.3 replaces H3 on chromatin during ZGA
Forward genetic screens remain at the forefront of biology as an unbiased approach for discovering and elucidating gene function at the organismal and molecular level. Past mutagenesis screens targeting maternal-effect genes identified a broad spectrum of phenotypes ranging from defects in oocyte development to embryonic patterning. However, earlier vertebrate screens did not reach saturation, anticipated classes of phenotypes were not uncovered, and technological limitations made it difficult to pinpoint the causal gene. In this study, we performed a chemically-induced maternal-effect mutagenesis screen in zebrafish and identified eight distinct mutants specifically affecting the cleavage stage of development and one cleavage stage mutant that is also male sterile. The cleavage-stage phenotypes fell into three separate classes: developmental arrest proximal to the mid blastula transition (MBT), irregular cleavage, and cytokinesis mutants. We mapped each mutation to narrow genetic intervals and determined the molecular basis for two of the developmental arrest mutants, and a mutation causing male sterility and a maternal-effect mutant phenotype. One developmental arrest mutant gene encodes a maternal specific Stem Loop Binding Protein, which is required to maintain maternal histone levels. The other developmental arrest mutant encodes a maternal-specific subunit of the Minichromosome Maintenance Protein Complex, which is essential for maintaining normal chromosome integrity in the early blastomeres. Finally, we identify a hypomorphic allele of Polo-like kinase-1 (Plk-1), which results in a male sterile and maternal-effect phenotype. Collectively, these mutants expand our moleculargenetic understanding of the maternal regulation of early embryonic development in vertebrates.
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