The objective of this work is to understand early vertebrate development at the molecular level. Approaches include the isolation of genes that play a critical role, analysis of their expression and function, and examination of the mechanisms whereby patterns of gene expression are altered in early development. The amphibian Xenopus has been chosen as the experimental animal because the embryos are large and easy to manipulate surgically, and the function of macromolecules, such as mRNA or protein, can be assayed by injection into living embryos. Furthermore, the early stages of development when the dorso-ventral and antero-posterior axes form are readily accessible to biochemical and embryological analysis. The molecular mechanisms which determine cell fate are conserved among vertebrates, so the findings from this work will further the understanding of development of all vertebrate species, including man. The objectives of this proposal are: 1) to determine the molecular basis for instability of a class of maternal RNAs that include homeobox-containing transcripts; the endonuclease that initiates degradation of the mRNAs will be purified, and its specificity and control elucidated. 2) to determine the mechanisms whereby specific genes are regulated to produce the detailed pattern of the embryo. Molecular markers will be used to determine the molecular basis for induction of different regions of the neural plate by underlying mesoderm and neighboring ectoderm. The role of peptide growth factors in regional induction will be scrutinized. Regional induction of the neural plate depends on prior regionalization of the dorsal mesoderm. To further understand the progressive restriction of mesodermal cell fate, the formation of muscle tissue will be used as a model. Progressive localization of MyoD transcripts and the role of a newly identified gene expressed in myotomes will be explored. A new structural motif in this gene was identified using antibodies to a heterologous protein. The same approach has been used to identify other motifs including a protein expressed on the surface of dorsal gastrula cells. The function of these newly identified proteins will be assayed in the embryo and in cell culture.
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