FIGURE 2.

Knocking down L29 or L30 stabilizes p53 and induces p53 activation. A, p53 induction and activation by knockdown of L29 or L30. U2OS cells were transfected with scrambled, L29, or L30 siRNA. Cell lysates were assayed for expression of p53, p21, MDM2, and L30 by IB using antibodies as indicated. The knockdown of endogenous L29 was determined by RT-PCR detection of the L29 mRNA (bottom panels). B, knockdown of endogenous L29 or L30 increases the mRNA expression of p53 targets mdm2 and p21. Total RNAs were extracted from U2OS cells transfected with siRNAs as in A and subjected to RT-qPCR assays. Relative expression of p21 and mdm2 genes was normalized against the expression of GAPDH. C, knockdown of endogenous L29 or L30 stabilizes p53. U2OS cells transfected with siRNAs as in A were treated with 50 μg/ml CHX and harvested at different time points as indicated. The cell lysates were assayed for levels of p53 and tubulin by IB. The bands were quantified and normalized with loading controls determined by tubulin expression and plotted in the bottom panel. D, knockdown of L29 or L30 induces p53 in WI38 cells. WI38 cells were transfected with scrambled, L29, or L30 siRNA. Cell lysates were assayed for expression of p53, p21, MDM2, and L30 by IB using antibodies as indicated.