Epigenetic regulation of the MEG3-DLK1 microRNA cluster in human Type 2 diabetic islets

Type 2 diabetes mellitus (T2DM) is a multi-factorial disease characterized by the inability of beta-cells in the endocrine pancreas to produce sufficient amounts of insulin to overcome insulin resistance in peripheral tissue. To investigate the function of miRNAs in T2DM, we sequenced the small RNAs of human islets cells from diabetic and non-diabetic organ donors and identified a cluster of miRNAs in an imprinted locus on human chromosome 14 to be dramatically down-regulated in T2DM islets. These miRNAs are highly and specifically expressed in human beta-cells. The down-regulation of this imprinted locus strongly correlates with increased methylation of its promoter in T2DM islets, providing evidence for an epigenetic modification that contributes to the pathogenesis of T2DM. Targets of the Chr 14q32 cluster of miRNAs were identified by high-throughput sequencing of cross-linked and immunoprecipitated RNA (HITS-CLIP) of Argonaute. We have also identified a unique class of sequences, termed chimeric reads, that represent an in vivo ligation of miRNAs and their targets while in complex with Argonaute, and which allow for the direct identification of miRNA:target relationships in vivo. Overall design: There are three experiments in this submission. All are in human islets or islet cell types. The first is a comparison of miRNA levels in sorted alpha versus beta cells. There is one replicate for this experiment. The second experiment is to measure the expression of miRNAs in whole islets as a function of glucose levels. There are three levels and one replicate for each condition. The third exeriment is a comparison of whole islets taken from human donors that were suspected/confirmed Type 2 diabetic or considered controls. There are 3 controls and 4 T2D samples.

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Kameswaran V, Bramswig NC, McKenna LB, Penn M, Schug J, Hand NJ, Chen Y, Choi I, Vourekas A, Won KJ, Liu C, Vivek K, Naji A, Friedman JR, Kaestner KH