The broad objective of this ongoing research project is to use C. elegans dauer formation as a genetically tractable model for chemosensation. The dauer larva is a dispensable alternative third larval stage whose formation is regulated by a combination of chemosensory signals and nutritional status. Previous work defined a large number of Daf-C (dauer formation constitutive and Daf-d (defective) genes that regulate dauer formation, and defined their participation in a complex set of genetic pathways. These pathways are molecularly and cellularly diverse. Chemosensory neurons use a cGMP mediated primary sensory transduction process to control dauer formation, partly through release of a TGF-related ligand. Downstream steps include the TGF-response pathway and an insulin- signaling pathway that integrates with the sensory pathways. It is likely that the main dauer-regulating output of these pathways is the ligand for the orphan daf-12 nuclear hormone receptor, which initiates dauer- regulating output of these pathways is the ligand for the orphan daf-12 nuclear hormone receptor, which initiates dauer differentiation. Our genetic approach to dauer formation has lead us to study diverse elements of this pathway. Here, we propose to genetically analyze a set of new daf genes that we have identified, most of which appear to function in the insulin-signaling pathway, and to molecularly study a select few of these genes. We will similarly analyze two genes that we have newly identified that may act at a downstream step in the TGF-beta pathway. The last known step in the insulin-signaling branch of the dauer pathway is the winged-helix transcription factor daf-16. In two aims, we will study genes that act close to or downstream of daf-16. In a genetic approach, we will isolate suppressors of a Daf-c allele of the protein kinase pdk-1, thought to act just upstream of daf-16. In a biochemical approach, we will use a combination of binding site selection, genome searches, and DNA-array based transcript analysis to search directly for transcriptional targets for daf-16. Finally, we have recently found that daf-19, a gene long thought to regulate sensory neuron development, encodes an RFX-type transcription factor that controls expression of proteins that comprise the core sensory cilium. We will use a combination of genome searches, transgenic expression tests, and DNA-array analysis to further investigate the role of daf-19. All of the pathways under study here correspond to important regulatory pathways in humans, and many of the genes are affected in various inherited disorders.
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