Embryonic development depends upon the utilization of messenger RNA and proteins stored in the egg as well as synthesized by the embryo. Developmental defects, which result in spontaneous abortion or birth defects in humans, can thus be the result of genetic or environmental perturbations of gene expression occurring during oogenesis or embryogenesis. This project is designed to assess the interaction between stored maternal components and those encoded by the embryonic genome in a convenient model system, the sea urchin embryo. Echinoid interspecies hybrid embryos fail to accumlate a variety of mRNAs of one species. The expression of one such gene encoding metallothionein (MT) is restricted posttranscriptionally in hybrids. The hypothesis that this is the result of a competition betwen MT mRNA for factors resulting in mRNA stabilization will be tested by measuring MT mRNA stabilities and by microinjection into eggs of MT mRNAs. The control of stability of other mRNAs will be analyzed by the microinjection of radioactive synthetic mRNAs produced from chimeric DNA molecules. The autoregulation of tubulin gene expression during embryogenesis will be analyzed using similar approaches, with the expectation that coding sequence elements of the mRNA are responsible for the control of mRNA stability by the level of unpolymerized tubulin dimers. The sequence elements responsible for the temporal and spatial specificity, as well as the induction by heavy metals and repression by chelators, of MT gene expression will be assessed in transient assays of expression of microinjected DNA mutated in vitro. This should result in the development of a very active inducible promoter system potentially useful for the conditional production of heterologous mRNAs and antisense RNA applicable to the analysis of gene function. Ultimately, these investigations should help establish an understanding of how gene expression is regulated and integrated during embryomic development and how this leads to the formation of increasingly complex structural and functional components of the embryo.
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