Effect of MDM2 on p65-dependent transcription.

EU-4 cells were cotransfected with 2 μg E-selectin promoter-CAT reporter construct or 5 μg p65 expression plasmid alone (lane 2), either with increasing amounts (1, 5, and 10 μg) of wt p53 (lanes 3-5) or with increasing amounts (1, 5, and 10 μg) of MDM2-sense in the presence (lanes 6-8) or absence (lanes 9-11) of a fixed amount of wt p53 (10 μg). MDM2 sense and antisense alone (10 μg), respectively, were cotransfected with E-selectin promoter-CAT as controls (lanes 12, 13). Controls also included cotransfection with E-selectin, p65, and MDM2 antisense (lane 14). Total concentration of DNA was adjusted to 27 μg per transfection with empty pCMV-neo-Bam vector. Transfection and CAT ELISA assay were as described in Figure 4.