Figure 5

miR-26 directly targets CHD1, GREB1 and KPNA2 genes. (a) Putative miR-26-binding sequences in the 3′ untranslated region (UTR) of CHD1, GREB1 and KPNA2 mRNA. Mutations were generated on the CHD1, GREB1 and KPNA2 3′ UTR sequence in the complementary site for the seed region of miR-26. (b) Luciferase reporter assays in T47D cells, with co-transfection of wild-type (wt) or mutant (mut) 3′ UTR and microRNA (miRNA) as indicated. (c) Luciferase reporter assays in T47D cells, with co-transfection of wt or mut 3′ UTR and miRNA sponge as indicated. (d) Forced expression of miR-26a or miR-26b in MCF-7 and T47D cells decreased protein levels of CHD1, GREB1 and KPNA2 by western blot analysis. (d, e) After forced expression or depletion of miR-26, the expression of CHD1, GREB1 and KPNA2 protein were analyzed by western blot assay. (f, g) Western blotting analysis of the expression of CHD1, GREB1 and KPNA2 protein in 17β-estradiol (E2)-treated MCF-7 and T47D cells transfected with the indicated miRNA molecules. *P < 0.01.