Raver1, a ubiquitously expressed protein was originally identified as a ligand for metavinculin, the muscle specific isoform of the microfilament-associated protein vinculin.The protein resides primarily in the nucleus, where it colocalizes and may interact with the polypyrimidine-tract binding protein PTB, which is involved in alternative splicing processes.During skeletal muscle differentiation, raver1 translocates to the cytoplasm and eventually targets to the Z-line of sarcomeres. Here, it colocalizes with metavinculin, vinculin and alphaactinin, all of which have biochemically been identified as raver1 ligands. To obtain more information on the potential role of raver1 in muscle structure and function, we investigated its distribution and fine localization in striated and smooth muscle of mouse. Three monoclonal antibodies that recognize epitopes in different regions of the raver1 protein were employed in immunofluorescence and immuno electron microscopy. Our results show that cytoplasmic accumulation of raver1 is not confined to skeletal muscle, but occurs also in heart and smooth muscle. Unlike vinculin and metavinculin, cytoplasmic raver1 is not restricted to costameres, but also represents an integral part of the sarcomere. In isolated myofibrils and in ultrathin sections of skeletal muscle, raver1 was found concentrated at the I-Z-I band. In addition, a minor fraction of raver1 was present in the nuclei of all three types of muscle.These data indicate that during muscle differentiation raver1 might link gene expression with structural functions of the contractile machinery of muscle.2