Role of MicroRNAs in Islet Beta-Cell Compensation and Failure during Diabetes
Figure 1
Role of miR-9 in insulin mRNA and promoter activity. (a) Effect of miR-9 on insulin mRNA. The RNA duplex containing the mature form of miR-9 [26] and a siRNA directed against miR-9 (si-miR-9) or a control oligonucleotide was transfected in MIN6 cells for 48 hrs. The expression of the preproinsulin mRNA was measured by quantitative PCR. The mRNA level was normalized against the housekeeping acidic ribosomal phosphoprotein P0 gene (Rplp0) and the expression level in cells transfected with the control siRNA was set to 100%. Data are the mean of ± SEM of 3 independent experiments. (b) Effects of miR-9 and dominant negative Oc2 mutant on the activity of an insulin reporter construct in MIN6 cells. MIN6 cells were transiently transfected with miR-9 RNA duplexes containing the mature form of miR-9 [26] or the dominant negative Oc2 mutant [26]. The cells were cotransfected with a luciferase reporter construct driven by a 600 bp fragment of the rat insulin promoter (Ripluc) and with pRLSV40, a construct producing a renilla luciferase activity under the control of the constitutive SV40 promoter. The firefly luciferase activity produced by Ripluc was normalized to the renilla luciferase activity to rule out differences in the transfection efficiency. The empty pGL3 basic (luc) was used as control. Each experiment was performed at least three times in triplicate.
