Enhanced muscle contractile activity increases antioxidant enzyme in oxidative skeletal muscle, in association with elevated Nrf2 expression, nuclear translocation, and DNA-binding activity. A critical molecular mechanisms for Nrf2 nuclear translocation involves enhanced p62 phosphorylation, which increases its binding affinity to Keap1 leading to dissociation of Nrf2. Importantly, loss of p62 in muscle significantly reduced regular exercise-mediated increase of antioxidant enzyme expression mimicking observations in muscle-specific Nrf2 knockout mice. Altogether these findings reveal an essential cooperation between p62 and Nrf2 in the regulation of antioxidant enzyme expression in oxidative skeletal muscle. These may have implications for the development of new therapies to preserve skeletal muscle mass and function in conditions that benefit from regular exercise such as in aging and chronic metabolic and/or cardiac diseases.