Naturally occurring CD4⁺ regulatory T (Treg) cells, which specifically express the transcription factor FoxP3 in the nucleus and CD25 and CTLA-4 on the cell surface, are a functionally distinct T cell subpopulation actively engaged in the maintenance of immunological self-tolerance and homeostasis. Recent studies have facilitated our understanding of the cellular and molecular basis of their generation, function, phenotypic and functional stability, and adaptability. It is under investigation in humans how functional or numerical Treg anomalies, whether genetically determined or environmentally induced, contribute to immunological diseases such as autoimmune diseases. Also being addressed is how Tregs can be targeted to control physiological and pathological immune responses, for example, by depleting them to enhance tumor immunity or by expanding them to treat immunological diseases. This review discusses our current understanding of Treg immunobiology in normal and disease states, with the perspective on the realization of Treg-targeting therapies in the clinic. Expected final online publication date for the Annual Review of Immunology, Volume 38 is April 26, 2020. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.