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The biologic role of each CD3 chain and their relative contribution to the signals transduced through the TCR/CD3 complex and to downstream activation events are still controversial: they may be specialized or redundant. We have immortalized peripheral blood CD4+ and CD8+ T lymphocytes from a human selective CD3 gamma deficiency using Herpesvirus saimiri. The accessibility of the mutant TCR/CD3 complex to different Abs was consistently lower in immortalized CD8+ cells when compared with CD4+ cells, relative to their corresponding CD3 gamma-sufficient controls. Several TCR/CD3-induced downstream activation events, immediate (calcium flux), early (cytotoxicity and induction of surface CD69 or CD40L activation markers or intracellular TNF-alpha) and late (proliferation and secretion of TNF-alpha), were normal in gamma-deficient cells, despite the fact that their TCR/CD3 complexes were significantly less accessible than those of controls. In contrast, the accumulation of intracellular IL-2 or its secretion after CD3 triggering was severely impaired in gamma-deficient cells. The defect was upstream of protein kinase C activation because addition of transmembrane stimuli (PMA plus calcium ionophore) completely restored IL-2 secretion in gamma-deficient cells. These results suggest that the propagation of signals initiated at the TCR itself can result in a modified downstream signaling cascade with distinct functional consequences when gamma is absent. They also provide evidence for the specific participation of the CD3 gamma chain in the induction of certain cytokine genes in both CD4+ and CD8+ human mature T cells. These immortalized mutant cells may prove to be useful in isolating cytosolic signaling pathways emanating from the TCR/CD3 complex.