CD8+ T cells are crucial for host defense against invading pathogens and malignancies. However, relatively little is known about intracellular signaling events that control the genetic program of their activation and differentiation. Using CD8+ T cells from TCR-transgenic mice crossed to protein kinase C-θ (PKCθ)-deficient mice, we report that PKCθ is not required for Ag-induced CD8+ T cell proliferation, but is important for T cell survival and differentiation into functional, cytokine-producing CTLs. Ag-stimulated PKCθ−/− T cells underwent accelerated apoptosis associated with deregulated expression of Bcl-2 family proteins and displayed reduced activation of ERKs and JNKs. Some defects in the function of PKCθ−/− T cells (poor survival and reduced Bcl-2 and Bcl-x L expression, CTL activity, and IFN-γ expression) were partially or fully restored by coculture with wild-type T cells or by addition of exogenous IL-2, whereas others (increased Bim EL expression and TNF-α production) were not. These findings indicate that PKCθ, although not essential for initial Ag-induced proliferation, nevertheless plays an important role in promoting and extending T cell survival, thereby enabling the complete genetic program of effector CD8+ differentiation. The requirement for PKCθ in different types of T cell-dependent responses may, therefore, depend on the overall strength of signaling by the TCR and costimulatory receptors and may reflect, in addition to its previously established role in activation, an important, hitherto unappreciated, role in T cell survival.