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. 2015 Jul 17;43(15):7577–7589. doi: 10.1093/nar/gkv728

Figure 3.

Figure 3.

Enhancement of miRNA function by Y27632 is mediated by increased PAIP2 expression. (A) β-globin reporter mRNA levels were determined using a β-globin construct carrying let-7 wt binding sites. Let-7-mediated mRNA decay was enhanced by Y27632 when transcription from this construct was stopped by doxycycline, but not when ActD was applied. β-globin reporter mRNA levels in 293T cells were determined by qRT-PCR 8 h after adding Y27632 with Dox or ActD treatment. Data represent the means ± S.D. of three experiments. *P< 0.05. (B) PAIP2 mRNA levels in 293T cells were determined by qRT-PCR at 6 h after Y27632 treatment. Data represent the means ± S.D. of three experiments. *P< 0.05. Similar results were obtained using Caco2 cells. (C) PAIP2 protein levels in Caco2 cells treated with Y27632 were determined at the indicated time points by western blotting. (D) Confirmation of reduced PAIP2 protein levels in PAIP2-knockdown Caco2 cells. (E, F) Y27632 did not enhance miRNA function in PAIP2-knockdown Caco2 cells. Cells were transfected with a reporter construct and the corresponding miRNA precursor-expressing plasmid [let-7 in (E) and miR-122 in (F)] and the effect of Y27632 was determined. Data represent the means ± S.D. of three experiments. *P< 0.05.