The thymus has been viewed as the main site of tolerance induction to self-antigens that are specifically expressed by thymic cells and abundant blood-borne self-antigens, whereas tolerance to tissue-restricted self-antigens has been ascribed to extrathymic (peripheral) tolerance mechanisms. However, the phenomenon of promiscuous expression of tissue-restricted self-antigens by medullary thymic epithelial cells has led to a reassessment of the role of central T-cell tolerance in preventing organ-specific autoimmunity. Recent evidence indicates that both genetic and epigenetic mechanisms account for this unorthodox mode of gene expression. As we discuss here, these new insights have implications for our understanding of self-tolerance in humans, its breakdown in autoimmune diseases and the significance of this tolerance mode in vertebrate evolution.