A number of mechanisms control T-cell reactivity to prevent autoimmune disease, including clonal deletion, anergy and regulatory T (Treg) cell activity. The challenge is to understand the molecular mechanisms of how processes cooperate to preserve immunological tolerance. Here, we have explored this issue by testing the synergy between genes that drive thymic deletion (via apoptosis) or anergy (inactivation). The combined loss of two key mediators of deletion, Bim and Aire, did not cause severe autoimmunity. By contrast, the combined loss of Aire with the anergy genes, Cblb or Pdcd1, caused a severe, fatal autoimmune disease that targets many different organs. Surprisingly, Cblb-/-Bim-/- and Pdcd1-/-Bim-/- mice did not succumb to fatal autoimmunity, indicating that Bim does not cooperate with these anergy genes. Together, these data suggest that: 1) the most potent synergy exists between the molecular mediators of different tolerance mechanisms (e.g. deletion and anergy), and; 2) Cbl-B- and PD-1-mediated anergic tolerance is dispensible for tolerance in Bim-, but not Aire-deficient mice. This differential reliance upon peripheral tolerance mechanisms may be explained by an expansion of Treg cells observed in Bim-/- mice, but not Aire-/-; these might act to maintain immunological tolerance in Bim-deficiency when anergy is impaired. These studies reveal differential requirements for cooperation among deletional and non-deletional tolerance mechanisms to prevent autoimmunity.