Supplemental Figure 3.

WT TP53 and p53 depletion with siRNA alters endogenous WT and mutant p53 protein levels and p21 protein levels in human and mouse ovarian cancer cell lines. (A) Human cancer cell lines were transfected with WT p53 and treated without or with nutlin-3a for 24 hours. Cell lysates were prepared, and Western blots were done to analyze the levels of p21 protein. Note that P21 is increased by WT p53 in all cell lines and is reduced by nutlin-3a by mechanisms not yet understood. (B) Human cells were treated with 20 nM siRNA for 24 hours. Lysates were prepared, and Western blots were done to determine the relative reduction of cellular levels of p53 and p21 in each sample. Note that SKOV3 cells lacking p53 do not express p21 and that OVCA420 and OVCAR3 cells expressing homozygous mutant alleles express low levels of p21 protein. (C–D) Mouse ovarian cancer cells expressing WT p53, heterozygous alleles (WT/R172H), or homozygous alleles (R172H) were treated with p53 siRNA at 0, 20, and 50 nM or 50 nM, respectively, for 72 hours. Note that the mouse cell lines are less sensitive to p53 siRNA knockdown of p53 and p21. Representative of 3 separate experiments.