Interaction of Integrin α7β1 in C2C12 Myotubes and in Solution with Laminin

Abstract The dimer of integrin α7 and β1 is a major laminin-binding receptor in skeletal muscle. We studied interactions of integrin α7β1 with the extracellular matrix protein laminin in solution and in intact cells. Integrin α7β1 bound to EHS laminin (laminin-1, composed of α1, β1, and γ1 chains), but not to endogenous laminin expressed in C2C12 myotubes. Northern blot analysis demonstrated that C2C12 myotubes synthesized laminin-1 α, β, and γ subunits mRNAs. C2C12 laminin was, however, immunologically distinct from EHS laminin; it was not recognized by 5D3 anti-laminin-1 monoclonal antibody, whereas 5A2 and LT3 antibodies reacted equally well with C2C12 and EHS laminins. Following deglycosylation of EHS laminin, separation of the subunits by SDS–PAGE, Western blotting, and partial amino acid sequencing of the protein bands, the epitope recognized by 5D3 antibody was localized to the γ1 laminin chain. Following bindingin vitro,the complex of EHS laminin and integrin α7β1 was subject to chemical cross-linking. The two proteins did not undergo cross-linking at the cell surface, consistent with the fact that in intact, resting myotubes integrin α7β1 interacted poorly with EHS laminin, which may reflect a limited accessibility of integrin α7β1 in the membrane to laminin or an inactive state of the integrin.