Figure 3

A: Gadd34 co-activation with EBNA3C requires aa483-610, and is independent of PP1 recruitment. A) BJAB cells (1 × 10⁷) were transfected with 5 μg of (-512/+72) LMP1p-Luc reporter construct, and 2 μg of psg5-EBNA2 (all lanes). psg5-EBNA3C (5 μg) was transfected(lanes 2-7). Gadd34 expression constructs were transfected as indicated (lanes 3-7) Luciferase values were normalized over beta-galactosidase levels obtained with co-transfection of 5 μg pgk-β-Gal plasmid. Reporter activation observed with EBNA2 alone is normalized to 1 (lane 1). Values are averages of duplicate observations in each experiment (repeated 3 times) plus standard error. A representative experiment is shown. B: Gadd34 180-483 is dominant negative in a low-dose EBNA3C co-activation assay. Reporter assay in BJAB cells with the indicated quantities of expression plasmids, as in figure 3a except that 100 ng (versus 5 μg) of psg5-EBNA3C expression plasmid was used. Each Gadd34 plasmids was titrated to determine stochiometry effects on EBNA3C, transfected at 0.1, 1.0 or 10.0 μg of DNA. C: Gadd34 does not co-activate transcription with EBNA2 in the absence of EBNA3C Reporter assay in BJAB cells with the indicated quantities of expression plasmids as in figure 3b. A representative experiment is shown. Protein expression levels of flag-tagged Gadd34, EBNA2 and EBNA3C are shown below.