The α-actinin-3 gene (ACTN3) R577X polymorphism is one of the most highly studied genetic variations associated with human skeletal muscle function and performance. This chapter summarizes the current research in ACTN3 R577X and discusses the molecular mechanisms that underlie the consequences of a-actinin-3 deficiency, with new insights into implications for the general population beyond athletic performance — in healthy ageing, muscle disuse, and disease. The α-actinins are a family of four actin-binding proteins that have evolved from repeated gene duplication events to perform similar roles in different cell types. The role of sarcomeric α-actinins in the maintenance of ordered myofibrillar arrays in skeletal muscle make them prime candidate genes for human muscle disease. The varied allele frequencies observed between the different ethnic groups and specialized protein expression in fast glycolytic muscle fibers suggest that selective forces may be acting on the ACTN3 577X allele during modern human evolution.