We propose to identify and characterize genes required for vertebrate somitogenesis, the process by which a 'homogeneous' field of pre- somatic cells is divided into reiterated segments called somites. Zebrafish are well suited for this task, as mutational screens can be performed with relative ease. Additionally, wild-type and mutant cells can be studied at single cell resolution, a critical feature if we are to understand how cells on one side of a somite boundary become distinct from their neighbors on the other side, just one cell diameter away. Recently, vertebrate homologs of Drosophila hairy/Enhancer of split [E(sp)] genes (her or hes genes) have been described that are expressed in wave-like fashion in the pre-somatic mesoderm, cycling with a periodicity equal to the time it takes to generate a somite. Two of the mutations we have isolated, one deleting her gene function and one causing ectopic her gene expression, will e used to understand the specific role of her genes and to investigate her genes are regulated during somitogenesis. To isolate additional genes, we will continue our sensitive genetic screen to isolate mutations that disrupt the segmental gene expression of her1, a zebrafish hairy/E(sp1)-related gene. Regulatory mechanisms governing segmentation in flies and dish appear distinct, likely reflecting the basic mechanistic differences between segmentation in these diverse organisms. For example, Notch/Delta signaling has been implicated in vertebrate somitogenesis, but is not required for fly segmentation. The experiments proposed will identify molecules involved in cell specification and cell-cell communication and begin to establish a regulatory hierarchy for their action. These studies will highlight molecules of potential importance in human development, as well as generate potential models for human genetic disease. Notch signaling has been implicated in somitogenesis, cell proliferation, cell fate decisions, organ and stem cell differentiation programs, and cancer, although the mechanisms by which notch signalling controls these diverse developmental decisions is largely unknown. The proposed screen will likely isolate mutations in genes already implicated in somite formation, but importantly, its unbiased approach will likely lead to the identification of previously undescribed molecules involved in Notch/Delta signaling, cell specification, and cell-cell communication.
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