Fig. 6

Characterization of miRNAs bound to the Pax6 3’UTR in pancreatic α cells. a Identification of miRNAs associated with the Pax6 3’UTR in αTC1–6 cells using TaqMan multiplex qPCR arrays. Normalized relative quantity (NRQ) greater than 1 indicates more target miRNA was purified with the Pax6 3’UTR than the control lacking a Pax6 3’UTR. Some miRNAs expressed in α cells that also have predicted MREs in the Pax6 3’UTR did not associate with the Pax6 3’UTR-containing transcript: miR-101, 1187, 144, 15a, 196b, 335-3p, 362-3p, 365, 410, 466f, 466 k, 495, 501–3p, 96. Results represent four independent experiments. Geometric mean +/− 95% confidence intervals are shown. b Landscape of Pax6 3’UTR miRNA interaction in αTC1–6 cells. Average NRQ value for each interacting miRNA is indicated as a peak at the 3’UTR position(s) of the predicted MRE(s). Predicted MREs for interacting miRNAs are labeled in black, and non-interacting miRNAs in grey. Each peak has a 25-nucleotide width on either side of the MRE position. Overlap between interaction peaks spaced 8–50 nucleotides apart is indicated in red, and these interacting miRNAs may be capable of mediating cooperative regulation of Pax6. MREs for Pax6 3’UTR-interacting miRNAs were found to cluster into three regions, i-iii (orange boxes), located approximately at nucleotide positions 250–350, 420–550 and 600–810. Conservation of the Pax6 3’UTR sequence between orthologous placental mammal sequences is shown for the 876-nucleotide length. Secondary, poorly conserved MREs for interacting miRNAs having well conserved predicted MREs are not shown. c miRNA interaction with the Pax6 3’UTR is not directly related to miRNA abundance in αTC1–6 cells. miTRAP ratio and relative miRNA level for αTC1–6-interacting miRNAs is shown. miTRAP ratio was calculated by dividing the relative abundance of each miRNA with the Pax6 3’UTR by the relative abundance in αTC1–6 cells. Larger miTRAP values indicated greater enrichment of the miRNA with the Pax6 3’UTR relative to cellular abundance