Figure 4.

miR-16-5p suppresses WIP1 expression, affects p53 stabilization and its activity, and enhances sensitivity to RG7388. (A) miR-16-5p sequences and its putative binding sites in the 3′-UTR of PPM1D; (B) Nalm6 cells were transfected with negative control miRNA and miR-16-5p mimic (200nM). Total RNA was extracted from DMSO (0.5%) treated Nalm6, mock control, and those transfected with negative control miRNA or miR-16-5p mimic at 48 h after transfection. miR-16-5p levels were measured by quantitative RT-PCR shown significant ectopic overexpression of miR-16-5p in transfected cells with miR-16-5p compared to DMSO control, mock control and negative control miRNA; (C) ectopic overexpression of miR-16-5p suppresses WIP1 expression, enhances p53 stabilization and upregulates expression of p53 target genes, p21^WAF1 and MDM2 in wild-type TP53 Nalm6 treated with RG7388. Nalm6 cells were transfected with miR-16-5p (200nM) or scrambled miRNA. Cells were treated with RG7388 (0.5 μM) at 48 h after transfection and their protein levels analyzed at 6 h after RG7388 treatment. The intensity of p53 blots (P < 0.05 or P < 0.1) (D) and p21^WAF1 blots (P < 0.05) (E) was quantified by Image J software and normalized with the DMSO control. Data shown are the average of three independent experiments and error bars represent SEM. Ctrl, Control. *P < 0.05, **P < 0.01