CTLA-4-deficient mice develop a fatal lymphoproliferative disorder, characterized by polyclonal expansion of peripheral lymphocytes. To examine the effect of restricting the CD4⁺ TCR repertoire on the phenotype of CTLA-4-deficient mice and to assess the influence of CTLA-4 on peptide-specific CD4⁺ T cell responses in vitro, an MHC class II-restricted T cell receptor (AND TCR) transgene was introduced into the CTLA-4^−/− animals. The expression of the AND TCR transgene by CD4⁺ T cells delays but does not prevent the lymphoproliferation in the CTLA-4^−/− mice. The CD4⁺ T cells become preferentially activated and expand. Interestingly, young AND TCR⁺ CTLA-4^−/− mice carrying a null mutation in the rag-1 gene remain healthy and the T cells maintain a naive phenotype until later in life. We demonstrate that CTLA-4 regulates the peptide-specific proliferative response generated by naive and previously activated AND TCR⁺ RAG^−/− T cells in vitro. The absence of CTLA-4 also augments the responder frequency of cytokine-secreting AND TCR⁺ RAG^−/− T cells. These results demonstrate that CTLA-4 is a key regulator of peptide-specific CD4⁺ T cell responses and support the model that CTLA-4 plays a differential role in maintaining T cell homeostasis of CD4⁺ vs. CD8⁺ T cells.