EFFECTS OF THIOL PROTEASE INHIBITORS ON MYOBLAST FUSION AND MYOFIBRILASSEMBLY IN-VITRO

To investigate the roles of thiol proteases such as cathepsins and cal pains in muscle differentiation, we have treated primary cultures of p ectoralis muscle with a variety of protease inhibitors and examined th e effects these agents have on myoblast fusion and myofibrillogenesis. We have found that a membrane-permeable inhibitor, E64D, has dramatic effects on both events of muscle differentiation. Cells treated with this inhibitor display gross morphological changes, severe delays in m yofibril assembly, and reduced ability to fuse in culture. These morph ological changes are correlated with a build up of beta 1-integrin thr oughout the cytoplasm. These effects could also be produced using NH4C l, a lysosomotrophic agent. In addition, we show that two nonpermeable inhibitors (leupeptin and E64) slightly decrease myoblast fusion, but have no effects on the ability of the cells to form mature myofibrils . These results are discussed in terms of their relevance to the inher itable disease of muscular dystrophy and I-cell disease (mucolipodosis II). Cell Motil. Cytoskeleton 40:354-367, 1998. (C) 1998 Wiley-Liss, Inc.