The negative selection of T cells in the thymus is necessary for the maintenance of self tolerance. Medullary thymic epithelial cells have a key function in this process as they express a large number of tissue-specific self antigens that are presented to developing T cells. Mutations in the autoimmune regulator (AIRE) protein cause a breakdown of central tolerance that is associated with decreased expression of self antigens in the thymus. In this Review, we discuss the role of AIRE in the thymus and recent advances in our understanding of how AIRE might function at the molecular level to regulate gene expression.