miR-375 targets PDK1 and regulates glucose-induced biological responses in pancreatic β-cells

Abstract

Objective: MicroRNAs are short non-coding RNAs that regulate gene expression. We hypothesized that the PI3kinase cascade known to be important in β-cell physiology could be regulated by microRNAs. Here, we focused on the pancreas-specific miR-375 as a potential regulator of its predicted target PDK1 and we analyzed its implication in the response of insulin-producing cells to elevation of glucose levels.

Research Design and Methods: We used INS-1E cells to analyze the effects of miR-375 on : PDK1 protein level and downstream signaling using western blotting, glucose-induced insulin gene expression using qRT-PCR and DNA synthesis by measuring thymidine incorporation. Moreover, we analyzed the effect of glucose on miR-375 expression in both INS-1E cells and primary rat islets. Finally, miR-375 expression in isolated islets was analyzed in diabetic Goto-Kakizaki rats.

Results: We found that miR-375 targets directly PDK1 and reduces its protein level resulting in decreased glucose-stimulatory action on insulin gene expression and DNA synthesis. Further, glucose leads to a decrease in miR-375 precursor level and a concomitant increase in PDK1 protein. Importantly, regulation of miR-375 expression by glucose occurs also in primary rat islets. Finally, miR-375 expression was found to be decreased in fed diabetic Goto-Kakizaki rat islets.

Conclusions: Our findings provide evidence for a role of a pancreatic specific microRNA, miR-375, in the regulation of PDK1, a key molecule in PI3kinase signaling in pancreatic β-cells. The effects of glucose on miR-375 are compatible with the idea that miR-375 is involved in glucose regulation of insulin gene expression and β-cell growth.