Figure 3.

(A) HEK-293 cells were transfected with expression constructs expressing activated H-Ras paired with and without an expression construct for NORE1A. 24 h post-transfection the cells were lysed and levels of endogenous MDM2 were analyzed. When activated Ras was over-expressed, levels of MDM2 were found increased. However, when NORE1A was co-expressed with the activated Ras, this effect was reversed and the MDM2 levels were suppressed compared to the empty vector control; (B) HepG2 cells were transfected with a scramble siRNA control or a siRNA designed against NORE1A. 48 h post-transfection, the cells were lysed and the levels of endogenous MDM2 were analyzed. In the cells where NORE1A expression was suppressed, levels of MDM2 were observed to increase, again suggesting that indeed NORE1A is negatively regulating MDM2 protein levels. Relative intensities are represented in fold change to the control sample and representative blots of 2 independent experiments are shown.