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. 1998 May;18(5):2815–2824. doi: 10.1128/mcb.18.5.2815

FIG. 2.

FIG. 2

FIG. 2

FIG. 2

Both c-Jun and TAM-67 synergize with C/EBPβ to activate the TNFα promoter. (A) Effects of C/EBPβ, c-Jun, and c-Jun mutants on PMA-induced TNFα promoter activation. Jurkat T lymphocytes were transfected by the DEAE-dextran method with plasmid vectors expressing C/EBPβ, c-Jun, the c-Jun mutants c-Jun-LZ, c-Jun-DBD, and TAM-67, and the TNFα promoter reporter construct (3 μg/transfection) containing 120 bp 5′ of the transcription start site. The C/EBPβ plasmid was transfected at a suboptimal concentration (2 μg), while the plasmids expressing wild-type and mutant c-Jun and the CMV vector control were added at increasing concentrations (0.25, 0.5, 1, 2, and 3 μg, represented by the columns from left to right), with the total DNA added kept constant (8 μg). Luciferase activity was reported as RLU, corrected for the total protein in each lysate. Fold activation is expressed as a mean ± 1 SE for three or more experiments. (B) Effects of cotransfection of C/EBPβ with c-Jun or its mutants on PMA-induced activation of the TNFα promoter. Jurkat T lymphocytes were transfected with plasmid vectors expressing C/EBPβ plus c-Jun or its mutants and the −120 TNFα promoter reporter construct (3 μg). Various concentrations of control, c-Jun, or mutant c-Jun (0.25 to 3 μg) were added to C/EBPβ (2 μg). In addition, to more readily compare the results of different experiments, prior to analyses, the results of each experiment were normalized to the values obtained for C/EBPβ, which was defined as 100%. Data for wild-type c-Jun are the means ± 1 SE of six experiments, while those involving the mutants are the means of four experiments. Differences between C/EBPβ alone and with mutant or wild-type Jun were determined by t test for matched pairs. (C) Effects of TAM-67 and CMV vector control on c-Jun-induced activation of TRE(IL-2)-Luc promoter-reporter in PMA-stimulated Jurkat T cells. The TRE(IL-2)-Luc promoter reporter contains two copies of the c-Jun binding site of the IL-2 promoter. TRE(IL-2)-Luc (3 μg) was transfected with an optimal concentration of c-Jun (0.25 μg) and increasing concentrations of TAM-67 (0.25, 2, and 5 μg) or control CMV vector (0.25, 2, and 5 μg). The results presented are the means ± 1 SE of a single experiment that was representative of four experiments.