Plectin is a versatile cytoskeletal linker protein and preferentially localized at interfaces between intermediate filaments and the plasma membrane in muscle, epithelial cells and other tissues. Its deficiency leads to disorganization of myofibrils and disruption of sarcolemmal membrane integrity in skeletal muscle. To get a better understanding of plectin's functional roles at the sarcolemma of skeletal muscles and to gain some insights into the underlying mechanism of plectin-deficient muscular dystrophy, we investigated both the ultrastructural localization and the molecular relationships of plectin to subsarcolemmal cytoskeletal components, such as desmin, dystrophin and vinculin, in rat skeletal muscles. Immunoelectron microscopy revealed that plectin fine threads tethered desmin intermediate filaments onto subsarcolemmal dense plaques overlying Z-lines. These subsarcolemmal dense plaques were found to contain dystrophin and vinculin, and thus may be the structural basis of costameres. Furthermore, the costameric dense plaques were apparently in register with -dystroglycan-specific gold labels on the outer surface of sarcolemma. Immunoprecipitation experiments showed in vivo association of plectin with desmin, vinculin, meta-vinculin, dystrophin and actin. Treatment of plectin immunoprecipitates with gelsolin reduced actin, dystrophin, vinculin, and meta-vinculin but not desmin, implicating that subsarcolemmal actin could at least partly mediate the interaction between plectin and dystrophin or (meta-) vinculin. Altogether, our data suggest that plectin, along with desmin intermediate filaments, might serve a vital structural role in the stabilization of the subsarcolemmal cytoskeleton, contributing to the maintenance of sarcolemmal integrity against shear stresses imposed during muscle contraction. © 2002, The Kitakanto Medical Society. All rights reserved.