EXPRESSION OF PERLECAN, A PROTEOGLYCAN THAT BINDS MYOGENIC INHIBITORYBASIC FIBROBLAST GROWTH-FACTOR, IS DOWN-REGULATED DURING SKELETAL-MUSCLE DIFFERENTIATION

Heparan sulfate proteoglycans (HSPG) have been shown to be involved in the activation of tyrosine kinase receptors by basic fibroblasts grow th factor (bFGF), a strong inhibitor of skeletal muscle differentiatio n, Skeletal muscle fibers contact extracellular matrix (ECM) that surr ounds individual fibers (endomysium) and bundles of several fibers (pe rimysium). Perlecan is a HSPG; present in the majority of basement mem branes. In this study we evaluated the expression and localization of perlecan during differentiation of C2C12 skeletal muscle cells. C2C12 myoblasts incubated with [S-35]Na2SO4 synthesize a HSPG that can be sp ecifically immunoprecipitated with antibodies against murine perlecan. The immunoprecipitated HSPG eluted from a Sepharose CL-4B with a Kav of 0.44, Analysis of the core protein of the HSPG immunoprecipitated f rom [S-35]methionine-labeled C2C12 after treatment with heparitinase r evealed two polypeptides of 170 and over 300 kDa. The amount of polype ptides immunoprecipitated decreased with muscle differentiation, Immun ocytolocalization studies indicate that perlecan is localized on the m yoblast surface and by immunogold staining we have demonstrated that i t is associated with patches of incipient extracellular matrix. The ex pression of perlecan mRNA decreased substantially during skeletal musc le differentiation, in contrast to the increase in transcripts for spe cific skeletal muscle proteins such as myogenin and creatine kinase. B y immunofluorescence microscopy almost no perlecan staining associated with the surface of myotubes was observed. All these results suggests that perlecan, a HSPG that binds myogenic inhibitory bFGF, normally a ssociated with basement membranes in adult tissues is present on the s urface of myoblasts and its expression is down regulated during skelet al muscle differentiation. (C) 1997 Academic Press.