The concern for the quality of food, its composition and contribution towards nutrition and health is widespread among public and private food system actors. The increasing interest in locally integrated supply chains leads to reconsider the configuration of the food system in relation to sustainability and health outcomes. This article focuses on the relationship between processing practices and nutritional value in the wheat-tobread sector, illustrated by a case study on the Tuscan Bread Protected Designation of Origin (PDO). By adopting a food system perspective, the case study shows how the different actors have mobilized to respond to multiple drivers of change. A mixed research method approach is adopted to illustrate the relationship between processing practices and nutritional value outcomes: practice-based indicators for each step of the chain are complemented with performance-based indicators of the chemical, physical and sensorial profile of Tuscan Bread PDO. Furthermore, the implications on food system governance of a differentiation strategy based on territorial origin and enhanced nutrition are discussed.