Fig. 3.
The effect of DNMT1 siRNA knockdown and treatment with other DNMT inhibitors on p53R2 expression in RKO cells. A, RKO cells were treated with the indicated concentrations of wildtype or mutant DNMT1 siRNA for five days. LF2000 indicates cells treated with only the transfection reagent. Alternatively, RKO cells treated with the indicated concentrations of decitabine (DAC) and harvested five days post treatment. DNA extracts were obtained, and genomic 5-methyldeoxycytidine (5mdC) levels were determined as described in Materials and Methods. B, p53R2 expression was measured by qRT-PCR under the same set of conditions described in A. C, The activity of the p53R2 intron 1 construct was measured under the same set of conditions described in A. D, p53R2 expression in RKO cells was measured by qRT-PCR five days post-treatment with the indicated concentrations of decitabine (DAC), 5-azacytidine (5-aza), zebularine, or RG108. RKO cells were treated with the indicated concentrations of drugs at day 0 and day 3 and cells were harvested at day 5 post treatment. PBS serves as the vehicle control for the 1st three drugs, while DMSO serves as the vehicle control for RG108.