Autoreactive CD4+ T cells play a major role in the pathogenesis of autoimmune diabetes in nonobese diabetic (NOD) mice. We recently showed that the non-MHC genetic background controlled enhanced entry into the IFN-γ pathway by NOD vs B6.G7 T cells. In this study, we demonstrate that increased IFN-γ, decreased IL-4, and decreased IL-10 production in NOD T cells is CD4 T cell intrinsic. NOD CD4+ T cells purified and stimulated with anti-CD3/anti-CD28 Abs generated greater IFN-γ, less IL-4, and less IL-10 than B6.G7 CD4+ T cells. The same results were obtained in purified NOD.H2b vs B6 CD4+ T cells, demonstrating that the non-MHC NOD genetic background controlled the cytokine phenotype. Moreover, the increased IFN-γ:IL-4 cytokine ratio was independent of the genetic background of APCs, since NOD CD4+ T cells generated increased IFN-γ and decreased IL-4 compared with B6.G7 CD4+ T cells, regardless of whether they were stimulated with NOD or B6.G7 APCs. Cell cycle analysis showed that the cytokine differences were not due to cycle/proliferative differences between NOD and B6.G7, since stimulated CD4+ T cells from both strains showed quantitatively identical entry into subsequent cell divisions (shown by CFSE staining), although NOD cells showed greater numbers of IFN-γ-positive cells with each subsequent cell division. Moreover, 7-aminoactinomycin D and 5-bromo-2′-deoxyuridine analysis showed indistinguishable entry into G0/G1, S, and G2/M phases of the cell cycle for both NOD and B6.G7 CD4+ cells, with both strains generating IFN-γ predominantly in the S phase. Therefore, the NOD cytokine effector phenotype is CD4+ T cell intrinsic, genetically controlled, and independent of cell cycle machinery.