The long term goal of this project is to understand how skeletal muscle growth and differentiation are regulated during embryonic development. The broad aims able are to determine how growth factor-mediated signals function to stimulate myoblast proliferation and to repress differentiation; and how deprivation of specific mitogens during the G1-cell cycle phase activates muscle specific gene expression and leads to the acquisition of a post-mitotic phenotype.
Specific aims are to purify and characterize the fibroblast growth factor receptor (FGFR), to isolate the FGFR gene, to identify components in the FGFR intracellular mitogenic signaling pathway, and to determine how FGFR gene expression and signal pathway components may change as proliferating myoblasts acquire the post-mitotic phenotype of terminally differentiated muscle. With respect to muscle-specific gene regulation, specific goals are to complete a delineation of the cis-acting regulatory elements of the mouse M- creatine kinase (MCK) gene, and to identify the factors with which these interact. Particular attention will be focused on isolating the muscle-specific factors that interact with an enhancer element of the MCK gene--the eventual goal being to clone and analyze the developmental control of the MCK regulatory gene that is responsible for muscle-specific factor production. Major methodologies consist of cell culture; growth factor receptor analysis; enzyme assays; protein purification and partial sequencing; polyclonal and monoclonal antisera production and use in affinity purification and screening protocols; cDNA and gene cloning and sequencing; construction of fusion genes; cell transfection; analysis of transient and stably integrated fusion gene expression; and the production and analysis of transgenic mice carrying various fusion gene constructs. Health related aspects of the project are its contribution toward the understanding of growth control and gene expression during embryonic development. Possible medical applications are to neuromuscular disease (particularly muscular dystrophy), to muscle tissue repair, and to normal and neoplastic growth regulation.
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