Regulatory T (Treg) cells enforce T cell homeostasis and maintain peripheral T cell tolerance. Here we report a previously unappreciated phenomenon of acute T cell lymphopenia in secondary lymphoid organs and non-lymphoid tissues triggered by Treg cell depletion that precedes the expansion of self-reactive T cells. Lymphopenia affects both neonates and adults indicating a dominant role of Treg cells in maintaining peripheral T cell numbers regardless of the developmental stage. The lymphopenia was neither triggered by caspase-dependent apoptosis nor macrophage-mediated clearance of T cells, nor diminished survival of naïve or recently activated T cells due to paucity of IL-7. It is possible that transient lymphopenia associated with congenital or acute Treg cell deficiency may contribute to the development of T cell mediated autoimmune disorders.