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Figure 3 | BMC Cancer

Figure 3

From: MDM2 negatively regulates the human telomerase RNA gene promoter

Figure 3

Mdm2 represses pRb and Sp1 mediated hTR promoter transactivation a) mdm2 suppresses Sp1 activation of hTR promoter activity. 5637 cells were co-transfected with 1.5 μg of the pLh2023(-796/+69) construct, 2.0 μg of the expression vector for either Sp1 and a titration of mdm2. Total input DNA amount for transfection was adjusted with Salmon Sperm DNA to ensure a constant amount in all transfections. After 48 hour of culture, cells were harvested, and the cell lysates assayed for firefly and Renilla luciferase activity. The data is expressed as fold induction of luciferase activity relative to the promoter alone and data are the mean of three independent experiments performed in duplicate. b) mdm2 suppresses pRB activation of hTR promoter activity 5637 cells were co-transfected with 1.5 μg of the pLh2023(-796/+69) construct, 2.0 μg of the expression vector for either pRb and a titration of mdm2. Total input DNA amount for transfection was adjusted with Salmon Sperm DNA to ensure a constant amount in all transfections. After 48 hour of culture, cells were harvested, and the cell lysates assayed for firefly and Renilla luciferase activity. The data is expressed as fold induction of luciferase activity relative to the promoter alone and data are the mean of three independent experiments performed in duplicate. c) Sp1 sites in the hTR promoter are not essential for repression by mdm2 5637 cells were co-transfected with 3.0 μg of the 2923 wild type or 2923 mSp1(4) mutant constructs [13], together with mdm2. After 48 hour of culture, cells were harvested, and the cell lysate were assayed for firefly and Renilla luciferase activity. The data is expressed as fold induction of luciferase activity relative to the promoter alone and the means of the three independent experiments performed in duplicate.

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