Research in my laboratory is focused on the fundamental issue of pattern formation during development using the zebrafish as a model. We are particularly interested in the molecular mechanisms governing patterning of the skeleton. Towards this end we have been studying the roles of inductive interactions mediated by the secreted signals, Sonic hedgehog (Shh), Nodal, and Endothelin-1 (ET-1), in craniofacial development. Shh and Nodal are signals expressed at the embryonic midline that could provide positional cues to skeletal precursors of the developing palate. ET-1 is expressed in the developing pharyngeal arches and is thought to pattern the lower jaw. Defects in these signals underlie several common human skeletal malformations such as cleft palate, holoprosencephaly and velocardiofacial syndrome. We are using a collection of mutations in these genes to study: 1) their roles in controlling the movements and division patterns of skeletal precursors, 2) interactions between the different signaling pathways, and 3) the sources of the signals and distances over which they act. We trace cells in living zebrafish, either microinjected with lineage tracers or in which green fluorescent protein expression is driven by cell type-specific promoters. We are finding that the clonal progeny of both neural crest and mesoderm that generate the craniofacial skeleton are disrupted both in their migration and proliferation patterns near the midline in Shh and Nodal mutants. We have created embryos carrying mutations in both Shh and Nodal signaling components and find that these pathways appear to synergize in patterning the midfacial skeleton. We are also using cell transplantation to create genetic mosaic embryos to determine which cells require each of these signals and the nature of the cellular responses.