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. 2007 Feb;18(2):536–546. doi: 10.1091/mbc.E06-05-0447

Figure 3.

Figure 3.

Mel-18 regulates Bmi-1 in human cells. (A) Stable expression of Mel-18 in MCF10A and MCF7 cells leads to down-regulation of Bmi-1 and c-Myc oncoprotein. Total cell lysate from indicated cells was analyzed by Western blot analysis using antibody against Bmi-1, Mel-18, c-Myc, and α-tubulin (loading control). (B) Transient overexpression of Mel-18 in 293T cells leads to the down-regulation of c-Myc and Bmi-1 in a dose-dependent manner. 293T cells were transiently transfected with increasing amounts pLPC-Mel-18, and 48 h after transfection total cell lysate was analyzed by Western blot analysis using antibody against Mel-18, Bmi-1, GFP (transfection control), and α-tubulin (loading control). (C) Stable knockdown of Mel-18 expression using the RNAi approach in MCF10A, WI-38, and BJ cells leads to up-regulation of c-Myc and Bmi-1 expression. MCF10A (left panel), WI-38 (middle panel), and BJ (right panel) cells were infected with pRS vector expressing either Mel-18 shRNA no. 1 (#1i), Mel-18 shRNA no. 2 (#2i), or an irrelevant control shRNA (Ctrl i), selected in puromycin and analyzed for the expression of Mel-18, c-Myc, and Bmi-1 by Western blot analysis.