FIGURE 4.

Egr-1 enhances the transcriptional activity mediated by the P1 enhancer element. A, Egr-1 enhances the transcriptional activity of the IL-4 P1 but not the IL-2 NFAT-binding element. Luciferase (Luc) reporter constructs containing four copies of either the human IL-4 P1 or the human IL-2 NFAT-binding element were co-transfected with the Egr-1 (4 μg) or the empty (4 μg) expression vector into Jurkat T cells. After overnight recovery, cells were split and either left unstimulated or stimulated with PMA (10 ng/ml)/ionomycin (1 μm) (PMA/iono.) for 8 h, followed by determining luciferase activity. B, Egr-1 cooperates with NFAT to enhance the transcriptional activity of the P1 element synergistically. The P1 luciferase reporter construct was co-transfected with Egr-1 (1 μg), NFATp (1 μg), NFATc (1 μg), or empty expression vector (1 μg) alone or in combinations into Jurkat T cells. After overnight recovery, cells were split and stimulated as in A. C, Egr-1 cooperates with NF-κB to enhance the transcriptional activity of the P1 element synergistically. The P1 luciferase reporter construct was co-transfected with Egr-1 (1 μg), p65 (1 μg), or empty expression vector alone or in combination into Jurkat T cells. After overnight recovery, cells were split and stimulated as in B. Results are representative of two (A) or three (B and C) independent experiments, each with triplicate transfections (error bars indicate S.D.).