Figure 4

PRDX1 is a direct target of microRNA 510. (A) Schematic representation of the predicted target site of miR-510 in the 3'UTR of peroxiredoxin 1 (PRDX1) mRNA. The numbers 1 to 294 represent base pairs in the 3'UTR of PRDX1. Complementary base sequences are highlighted in upper case letters. Western blot (B) and quantitative PCR (C and D) analysis of endogenous PRDX1 (B and C) and miR-510 (D) levels in the cell lines used in the study. GAPDH is shown as a loading control for western blot in B and is used for normalization in C and D. (E) Luciferase activity of HEK293 cells transfected with pGL3 promoter luciferase reporter vector alone (EV), the PRDX1 3'UTR reporter construct (PRDX 3'UTR) or PRDX1 3'UTR reporter construct mutated in the miR510 seed sequence binding site (PRDX mut). (F) Luciferase activity of MDA MB 231 cells stably transfected with scrambled control (dark gray bars) or miR-510 (light gray bars) transiently transfected with reporter constructs described in (E). All luciferase assays were normalized to Renilla luciferase activity. The data are expressed as the mean ± SD for three experiments conducted in triplicate.