9507409 Bucher The long term goal of this pilot project is to understand the genetic mechanisms of muscle cell specification in a developing organism. In vitro studies of vertebrate muscle gene regulation have revealed a complexity of transcription factors involved in muscle cell differentiation and determination. The analysis of muscle gene regulation in vivo should extend these studies to understand genetic pathways and to define regulatory factors that can only be discovered in vivo. This pilot project undertakes one aim, comprised of transgenic studies, to investigate the in vivo sequence requirements for temporal and oviduct lineage specific expression of the C. elegans troponin T (CeTnT) muscle gene. C. elegans is an organism having well established genetic and molecular transgenic approaches and the C. elegans oviduct is an ideal tissue to undertake these studies: the oviduct is comprised of only 10 muscle cells that arise through a simple stereotyped series of cell divisions. Furthermore, similar to some vertebrate muscle types, the oviduct dose not express the CeMyoD transcription factor and thus other factors may be revealed. The CeTnT gene structure is simple and provides an ideal molecular target to study gene expression. Different CeTnT genomic sequences will be cloned upstream of reporter genes. Reporter gene expression will then be assayed in individual cells of transgenic worms to characterize the in vivo requirements for oviduct specific expression. Dr. Bucher will test the hypothesis that MEF2 and E-box consensus sequences are required for oviduct expression. She will also test the hypothesis that novel consensus sequences are required for oviduct expression. Identification of new cis sequences elements that direct oviduct expression will provide the basis for future experiments involving the cloning of new transcription factors using molecular/biochemical approaches and by genetic suppression strategies. The expectation is that these appro aches will ultimately elucidate in vivo genetic pathways and define new genes involved in extracellular and intracellular signalling, as well as transcriptional regulators of the myogenic regulatory pathway. ***
