IL-12 and IFN-gamma-inducing factor (IGIF) have the capacity to stimulate IFN-gamma production by T cells. Using an IL-12-responsive T cell clone, 2D6, we investigated how these two cytokines collaborate for IFN-gamma production. 2D6 obtained from cultures containing rIL-12 produced IFN-gamma in response to rIGIF. 2D6 from cultures deprived of IL-12 for 24 h produced only marginal levels of IFN-gamma production following stimulation with either rIL-12 or rIGIF alone. However, simultaneous stimulation of these 2D6 cells with both cytokines resulted in strikingly enhanced levels of IFN-gamma production. 2D6 could also be maintained in the presence of rIL-2 instead of rIL-12. 2D6 lines maintained with rIL-12 (2D6(IL-12)) or rIL-2 (2D6(IL-2)) exhibited differential IGIF responsiveness: both lines responded similarly to rIL-2 or rIL-12, whereas the 2D6(IL-12) or 2D6(IL-2) exhibited high or marginal IGIF responsiveness, respectively. The 2D6(IL-12) line expressed IGIF receptor (IGIFR), whereas the 2D6(IL-2) did not. Overnight exposure of the 2D6(IL-12) to rIL-2 reduced IGIFR expression and conversely, exposure of the 2D6(IL-2) to rIL-12 restored IGIFR expression. IGIFR expression by these 2D6 lines correlated with the capacity to produce IFN-gamma in response to rIGIF. Purified naive T cells stimulated with anti-CD3 plus anti-CD28 mAb and subsequently cultured with rIL-12 were also found to express IGIFR and induce enhanced IFN-gamma production following IGIF stimulation. These results indicate that the induction of IGIFR by IL-12 represents one of the mechanisms underlying the synergy between IL-12 and IGIF in IFN-gamma production.