Thesis

REGULATION OF MICRORNA-375

MicroRNAs are important regulators of eukaryotic cellular processes and their misregulation is associated with many human diseases. MicroRNA-375, a regulator of insulin secretion in pancreatic beta cells, is misregulated in human type 2 diabetes. The aim of this study is to elucidate the regulation of microRNA-375. My hypothesis is that microRNA-375 is regulated in pancreatic beta cells through protein kinase A. With this hypothesis, I aimed to uncover fundamental mechanisms which lead to type 2 diabetes. Using bioinformatics, I identified putative binding sites in the microRNA-375 promotor for transcription factors in the protein kinase A pathway. With microRNA target prediction algorithms, I found some of these transcription factors may in turn be targeted by microRNA-375. Luciferase reporter assays indicated that cyclic-AMP increases the transcriptional activity of discrete regions of the microRNA-375 promoter. Chromatin immunoprecipitation assays revealed RNA polymerase II associates with the microRNA-375 promoter in response to cyclic-AMP. Cell treatments and real time PCR indicate that microRNA-375 is regulated by glucose and cyclic-AMP in a protein kinase A dependent manner.

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