Summary: CD4+CD25+ regulatory T cells play a crucial role in preventing autoimmune disease and can also modulate immune responses in settings such as transplantation and infection. We have developed a transgenic mouse system in which the role that T-cell receptor (TCR) specificity for self-peptides plays in the formation of CD4+CD25+ regulatory T cells can be examined. We have shown that interactions with a single self-peptide can induce thymocytes bearing an autoreactive TCR to undergo selection to become CD4+CD25+ regulatory T cells and that thymocytes bearing TCRs with low affinity for the selecting peptide do not appear to undergo selection into this pathway. In addition, thymocytes with identical specificity for the selecting self-peptide can undergo overt deletion versus abundant selection to become CD4+CD25+ regulatory T cells in response to variations in expression of the selecting peptide in different lineages of transgenic mice. Finally, we have shown that CD4+CD25+ T cells proliferate in response to their selecting self-peptide in the periphery, but these cells do not proliferate in response to lymphopenia in the absence of the selecting self-peptide. These studies are determining how the specificity of the TCR for self-peptides directs the thymic selection and peripheral expansion of CD4+CD25+ regulatory T cells.