Fig. 1

miR-101 targets Rhes mRNA 3′UTR. a Scheme of Rhes mRNA structure and miRNAs that bind to Rhes mRNA 3′UTR as predicted by DIANA-microT. b Luciferase assay using a part of the Rhes mRNA 3′UTR and miRNAs. (Upper) Luciferase activity was inhibited by miR-101. HEK293 cells were co-transfected with a reporter vector without the insert (Control) or with Rhes 3′UTR (Rhes) and 50 nM miRNA mimics (miR-sc, miR-101, or miR-132). At 24 h after transfection, the cells were lysed, and the luciferase activity of the cell lysates was measured. The activity was normalized to that of a control transfected with miR-sc. Data are presented as the mean ± SEM, n = 5. (Lower) miR-101 inhibited luciferase activity in a dose-dependent manner. HEK293 cells were co-transfected with a reporter vector containing Rhes mRNA 3′UTR and the miR-101 mimic at specified concentrations. Data are presented as the mean ± SEM, n = 5. c Sequence of the part of the Rhes mRNA 3′UTR (1731–1770) containing the predicted binding site for miR-101. Mut, substitution mutant of Rhes mRNA 3′UTR; Del, deletion mutant of Rhes mRNA 3′UTR. d The mutant form of Rhes mRNA 3′UTR eliminated the luciferase activity inhibition by miR-101. HEK293 cells were co-transfected with a reporter vector and 50 nM miRNA mimics. The activity was normalized to that of a control transfected with miR-sc. Data are presented as the mean ± SEM, n = 5. *p < 0.05, **p < 0.01