The simplicity and effectiveness of calorie restriction (CR) in lifespan and healthspan extension have fascinated generations searching for the Fountain of Youth. CR reduces levels of oxidative stress and damage, which have been postulated in the free radical theory of aging as a major cause of aging and diseases of aging. This reduction has long been viewed as a result of passive slowing of metabolism. Recent advances in nutrient sensing have provided molecular insights into the oxidative stress response and suggest that CR triggers an active defense program involving a cascade of molecular regulators to reduce oxidative stress. Physiological studies have provided strong support for oxidative stress in the development of aging-associated conditions and diseases but have also revealed the surprising requirement for oxidative stress to support normal physiological functions and, in some contexts, even slow aging and prevent the progression of cancer. Deciphering the molecular mechanisms and physiological implications of the oxidative stress response during CR will increase our understanding of the basic biology of aging and pave the way for the design of CR mimetics to improve healthspan.