We cloned and characterized a novel member of the tenascin family of extracellular matrix proteins - the murine orthologue of zebrafish tenascin-W. Full-length recombinant tenascin-W was expressed and purified from mammalian cell cultures. Rotary shadowing followed by electron microscopy showed that tenascin-W forms hexabrachions. We studied its expression during development and in the adult by immunohistochemistry, in situ hybridization and immunoblotting. Tenascin-W is expressed during palate formation, osteogenesis and smooth muscle development. In the adult, tenascin-W is found in the kidney, cardiac semilunar valves, corneal limbus and periosteum. Tenascin-W and tenascin-C expression overlap in many of these areas. Bone-morphogenic-protein-2 treated C2C12 cells secrete tenascin-W and are able to adhere to and to extend actin-rich processes on a tenascin-W substratum. In vitro, cells bind to tenascin-W in an RGD-dependent manner. This adhesion is increased by transfection of α8 integrin, which localizes with tenascin-W in the periosteum and kidney.