This proposal represents an application for renewal of a FIRCA grant to continue collaborative studies on the characterization of skeletal muscle membrane associated proteoglycans. The long range goal of this project is to understand the role of extracellular matrix (ECM) and ECM receptors in skeletal muscle cell differentiation. The terminal differentiation of skeletal muscle cells is dependent on signals arising from soluble growth factors and from ECM. The molecular basis of the effect of ECM contact on myogenesis is not understood. During the previous funding period we determined that the synthesis of sulfated proteoglycans was essential for the differentiation of myoblasts into myotubes. We also identified and characterized 2 principal membrane associated heparan sulfate proteoglycans synthesized by differentiating muscle cells, glypican and syndecan- 1. The hypothesis underlying the proposed studies is that these proteoglycans function as cell surface receptors for adhesive proteins of the ECM, and play essential roles in muscle cell-- ECM interactions during differentiation. We will utilize a well characterized mouse myoblast cell line, C2C12, that can be induced to undergo normal terminal differentiation in culture. One of the specific aims of the proposed work is to identify specific extracellular ligands that bind to glypican and/or syndecan- 1 isolated from skeletal muscle cells. We will test a panel of well characterized potential ligands for this activity, as well as attempt to identify potential novel muscle cell-secreted ligands for these proteins. The second aim is to determine the functional consequences of inhibition of glypican on syndecan-1 synthesis on skeletal muscle cell terminal differentiation. Inhibition of their expression will be accomplished by transfection of myoblasts with antisense expression vectors. These experiments should provide important new information on the function of these proteoglycans during skeletal muscle cell differentiation.