The initial specification of cell types along dorsal-ventral axis of the vertebrate spinal cord requires the activities of the Sonic hedgehog (Shh), Nodal and BMP signaling pathways. Mutations in many of the components in these signaling pathways are known to cause human birth defects and tumors. Thus understanding how these signaling pathways function in vivo is of clear importance for human health. The lab has identified seven ENTJ-induced mouse mutations that change cell fate along the dorsalventral axis of the spinal cord. The experiments proposed will define how these mutations affect neural patterning. One mutation is allelic to smoothened, an essential component of the Shh pathway. One mutation is an allele of the open brain (opb) gene, which the lab showed encodes Rab23 and acts as a negative regulator of the Shh pathway. The other five genes appear to be novel, because they map to regions that do not include genes previously implicated in neural patterning. Genetic analysis will be used to determine which step in the Shh pathway is controlled Opb/Rab23. The subcellular localization of Rab23 will be characterized and its role in controlling the localization of proteins known to act in the Shh signaling pathway will be determined. The Drosophila homologue of Rab23 will be studied. If the Drosophila gene acts in the Hedgehog pathway, reagents and techniques available in Drosophila will be used to define the mechanism of action of this gene. A novel mouse gene, wimple (wim), will be characterized. wim mutants lack ventral cell types in the spinal cord lack normal left-right asymmetry, and preliminary data suggest that wim may act downstream of Patched in the Shh pathway. Map-based strategies will be used to identify the wim gene, and the function of the Wim protein will be characterized. Four additional novel mutations identified in the screen that affect early dorsal-ventral patterning the spinal cord will be characterized genetically. These experiments will identify new components of the signaling pathways that control neural patterning and will define the processes that are essential for early dorsal-ventral neural cell type specification.
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