DESCRIPTION (from applicant's abstract): This application examines how a neuron is specified to express a defined set of differentiated characteristics. Dr. Emmons has been examining the morphogenesis of the male tail in the nematode C. elegans. The male tail is composed of nine morphologically distinct pairs of rays in an acellular fan. The 3 cells of each ray, 2 neurons and a structural cell are derived from a ray precursor cell, Rn. By characterizing mutants with abnormal tail morphologies, several genes responsible for tail morphogenesis have been identified. Repeated expression of the ray sublineage is dependent on expression of lin-32, a bHLH transcription factor of the achaete/scute family. Differences among the rays are dependent on several transcription factors of the C. elegans Hox cluster, mab-5, egl-5, and Pax-6. The expression of these transcription factors may be regulated by extracellular factors. One extracellular factor that is involved in ray 5, 7, and 9 formation is dbl-1, which encodes a BMP2 homolog. Dbl-1 may act through SMA-6 and DAF-4, type I and type II BMP receptors, respectively, and the products of the sma-2, sma-3, and sma-4, which are SMAD proteins. Among the differentiated characteristics of neurons is the expression of their complement of transmitters. Dr. Emmons proposes to focus on how neurotransmitter phenotype is specified, and will examine the specification of dopamine (DA) in three bilateral pairs of sensory neurons in rays 5, 7, and 9 of the male tail. Transgenic animals carrying a TH::GFP reporter construct were examined in dbl-1, sma-3, egl-5, and mab-21 loss of function backgrounds. These animals expressed GFP ectopically in other rays, suggesting that the expression of DA is independent of BMP but DA patterning is dependent on BMP. Overexpression of the DBL-1 protein by a hs::dbl-1 transgene resulted in GFP expression in rays 3-9, but never in rays 1 or 2. These data suggest that DBL-1 may provide instructive cues. Since no expression is seen in rays 1 or 2, these rays may be incompetent to respond to the DBL-1 signal. The investigator proposes the model that DA specification is based on three patterning processes: patterned expression of the ligand (DBL-1), pre-patterning of cell competence by egl-5 and some other unidentified gene, and lateral inhibition. The aims of this application are to: (1) examine the role of DBL-1, Hox genes, and other genes in inducing DA phenotype. TH::GFP expression will be examined in animals with mutations in BMP and Hox genes. This will determine whether all cells have equal competence to take on a DA phenotype, to distinguish between whether ray cells are prepatterned to respond to BMP or whether BMP is spatially patterned, and to determine whether Hox genes play a role in establishing competence to cells to respond to the BMP signal. It will also be determined whether there is an inhibitory pathway restricting DA expression. Animals with mutations in mab-21, which encodes a novel protein, will be examined for TH::GFP expression; wild-type animals will also be ablated for different ray precursor cells. (2) determine whether common cis-regulatory elements are present in genes involved in DA synthesis. Promoter regions necessary for DA expression in rays will be delimited and examined for conserved motifs. (3) screen for mutants (including ts alleles) in which TH::GFP expression is lost and characterize the corresponding genes molecularly. (4) identify genes involved in the axon connectivity of DA neurons.
|Effective start/end date||7/16/99 → 6/30/03|
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