The dystrophin-glycoprotein complex plays an important role in muscle function. One of the components of the complex, a 156-kDa cell surface glycoprotein (a-dystroglycan) binds to laminin, thereby connecting the basal lamina and muscle cells. We have examined the progressive appearance of a-dystroglycan and laminin in muscle cells that differentiate in culture. We find that nondifferentiated cultures of C2C12 myoblasts express low amounts of dystroglycan mRNA and, in contrast, this gene is prominently expressed in differentiated myotubes. Immunofluorescence analysis with a monoclonal antibody against a-dystroglycan shows its progressive appearance during myoblast differentiation into myotubes. lrnmunostaining with a monoclonal antibody against laminin shows that it is not present on the surface of undifferentiated myoblasts. Subsequently, laminin becomes apparent on the surface of differentiated myotubes where it codistributes with immunostained a-dystroglycan. lmmunoblotting of isolated surface membrane proteins of differentiated myotubes with antibodies against a-dystroglycan identifies a broad band of about 140-1 60 kDa, resembling a-dystroglycan from rabbit muscle. The composite results indicate that a-dystroglycan and laminin appear and become co-distributed on the surface of cultured C2C12 during the progression of differentiation.