The long term goals of this proposal are to understand the structures, functions and protein interactions of muscle-specific alpha-actinins and to apply this information to the study of human neuromuscular disorders. Alpha-actinins are a family of closely related actin-binding proteins that serve to cross link and anchor actin filaments. In non-muscle cells, calcium-sensitive cytoskeletal isoforms are found in association with f- actin-containing stress fibers and at adhesion plaques where they participate in linking the internal cytoskeleton to the extracellular matrix. In muscle, calcium-insensitive alpha-actinins are a major component of Z lines and discs where they constitutively anchor the actin/nebulin-containing thin filaments. A number of human neuromuscular diseases have been shown to result from mutations of muscle-specific cytoskeletal elements. Similarly, several inherited cardiomyopathies are caused by mutations in genes for cardiac-specific isoforms of sarcomeric proteins. We hypothesize that some human neuromuscular diseases may be caused by mutations in muscle-specific alpha-actinin genes, and/or in genes for proteins that interact with alpha-actinin. We have cloned and characterized genes for human alpha-actinins and have recently identified several patients with congenital muscular dystrophy who lack expression of one of the muscle-specific isoforms. This proposal entails: 1) extending this observation in additional patients with primary disorders of muscle and identifying mutations in the alpha-actinin genes of deficient patients: 2) further characterizing actin-actinins in normal tissues: 3) developing cell culture and transgenic mouse models of alpha- actinin dysfunction: 4) identifying genes for novel proteins that interact with alpha-actinins: 5) characterizing these new genes and proteins: and 6) assessing these as candidate genes for other human neuromuscular diseases. Direct clinical benefits will include accurate pre- and postnatal diagnoses for these disorders better understanding of their underlying etiology, and insights into potential therapies. These experiments will also increase our understanding of alpha-actinin function and identify new interactions with existing and novel muscle proteins at the Z-line and elsewhere.