Figure 6. Inhibition of Sall2 expression by p53.

Early passages MEFs p53^ER/ER were treated with 4 hydroxytamoxifen (4-OH Tamoxifen) or vehicle for 4 hours before doxorubicin treatment. A. Western blot analysis of SALL2 and p21, using whole-cell lysates from early-passage MEFs p53^ER/ER that were cultured in either the presence (4-OH Tamoxifen) or the absence (vehicle) of 4-hydroxytamoxifen. β-actin and GAPDH show equal loading. Shown below densitometric analysis of the data using ImageJ. Sall2 band intensities were normalized by GAPDH. Results are expressed as fold changes relative to control vehicle treated cells, and representative of three independent experiments with similar results. B. RT-PCR analysis of Sall2 on total RNA isolated from same experiment as in A. Cyclofilin is used as normalizing gene. Shown below densitometric analysis of Sall2 mRNA levels normalized to cyclofilin. Results are expressed as fold changes relative to control vehicle treated cells. C. Western blot analysis of SALL2 and p53 using total protein lysates obtained from Rat PC12 pheocromocitoma cells transfected with various concentrations of p53. Lysates were subjected to SDS-PAGE and levels of endogenous SALL2 and exogenous p53 were evaluated by western blotting. Actin shows equal loading. D. Rat PC12 pheocromocitoma cells were transfected with 4 µg of wild type p53 and lysates were collected after 7, 14, and 24h post transfection. Endogenous SALL2 and exogenous p53 levels were evaluated by western blotting. GAPDH shows equal loading. E. Western blot analysis of endogenous SALL2 and p53 in rat PC12 cells treated with 10µM Etoposide for 12h. Actin shows equal loading. F. Western blot analysis of endogenous SALL2 and p53 in human HCT116 (p53+/+) colon cancer cells treated with 10µM etoposide for 24h. Actin shows equal loading. For all western blot molecular weight markers are indicated on the left side.