Embryonic development depends upon the utilization of messenger RNA stored in the egg as well as synthesized by the embryo. Defective development, which results in spontaneous abortion or birth defects in humans, can thus be the result of genetic or environmental perturbations of gene expression during oogenesis or embryogenesis. These investigations will assess the representation, properties, and roles of messenger RNA transcribed from the maternal and zygotic genomes in sea urchin embryos. Newly synthesized RNA of eggs and embryos labelled with thiophosphate and selected by affinity chromatography on an organomercury column, will be compared to maternal RNA or RNA synthesized at other stages by nucleic acid hybridization and electrophoretic analyses of the products of cell-free translation of the RNA. The possible levels of gene regulation responsible for the restricted expression of many paternal genes in interspecies hybrid embryos will be examined using cloned recombinant DNA probes. This, in turn, should provide information about the regulation of expression of these genes in embryos of the paternal species. DNA clones corresponding to maternal messenger RNAs which persist throughout embryonic development, if any, will be identified. The structural and translation properties of these transcripts will be examined in eggs and embryos. DNA clones will be identified corresponding to maternal messenger RNA sequences which disappear during development; the kinetics of decay of these transcripts will be determined using the cloned DNA probes. Genes undergoing major changes in their relative rates of transcription during development will be identified by hybridizing RNA synthesized in isolated nuclei to collections of DNA clones. The state of methylation and sensitivity to nuclease digestion in chromatin of the DNA of these genes will be examined during development. Finally, we will relate these various cloned genes to the products of their expression by electrophoretic separations of the products of translation of their hybrid selected messenger RNAs. These products, in turn, will be localized in eggs and embryos by immunocytochemical staining techniques. Ultimately these investigations should help lead to an understanding of how gene expression is regulated and integrated during embryonic development and how this leads to the formation of increasingly complex structural and functional components of the embryo.
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