We previously identified a Fas ligand regulatory element (FLRE) in the Fas ligand (fasL) promoter that binds Egr family proteins and demonstrated that Egr-3 (PILOT) but not Egr-1 (NGFI-A, Krox-24, Tis-8, and Zif-268) induces transcription of fasL. The aberrant CD4(-)CD8(-) T cells from lpr/lpr and gld/gld mice, which have mutations in the genes encoding Fas and FasL, respectively, have an activated phenotype and constitutively express high levels of fasL mRNA, prompting us to ask what role if any the FLRE and Egr family proteins have in this aberrant expression of fasL. Unstimulated MRL-lpr/lpr and C3H-gld/gld CD4(-)CD8(-) T cells constitutively contained high levels of two proteins that bound to the FLRE. Supershift analysis revealed these proteins to be Egr-1 and Egr-2 (Krox-20); Egr-3 was not detected. Activation of normal lymph node cells resulted in increased expression of Egr-1, -2, and -3. As with egr-3, expression of egr-2 was blocked by cyclosporin A. Although overexpressed Egr-1 was ineffective, overexpressed Egr-2 was as potent as Egr-3 in inducing fasL promoter-dependent reporter constructs in T cell hybridomas and HeLa cells, and both up-regulated endogenous fasL mRNA in HeLa cells. FasL-dependent reporter constructs in MRL-lpr/lpr and C3H-gld/gld CD4(-)CD8(-) T cells were constitutively active, and this activity was largely prevented by mutation of the critical Egr family binding element. Thus, Egr-2, in addition to Egr-3, regulates FasL expression in activated normal T cells, and Egr-2 is likely to play a direct role in aberrant fasL up-regulation in lpr/lpr and gld/gld CD4(-)CD8(-) T cells.