A porcine microRNA and its targets

Only a few porcine microRNAs (miRNAs) are known. Because of their close phylogenetic distance to humans, pigs serve as a suitable model for studying intestinal development or disease. Recent studies indicate that miRNAs are key regulators of intestinal development and their aberrant expression leads to intestinal malignancy. The aim of the study was to identify and validate the regulative interactions between porcine miRNAs and their targets. These miRNAs were identified from porcine intestinal tissue after deep sequencing and microarray validation. We focused on a cluster including eight miRNAs, which showed increased expression in distal jejunum and ileum compared with other intestinal loci. One of the identified miRNAs was predicted to target all members of the Trefoil factor family (TFF1-3). TFFs are known as protective mucosal peptides mediating mucosal healing as well as restitution and are of particular interest in the developing intestine. RNAhybrid[1] analysis revealed significant binding probabilities between the identified miRNA and all TFFs (PTFF1<0.02, PTFF2<0.02 and PTFF3<0.005). The 3´UTR of each TFF member was cloned into reporter plasmids to verify the biological function of predicted interactions. The miRNA precursor was cloned into a second expression plasmid. The porcine intestinal epithelial cell line IPEC-J2 was cotransfected using the reporter-, expression- as well as normalisation plasmids. Our initial results point to a down-regulation of TFF3 after cotransfection. Further miRNA mimic experiments as well as the investigation of the two other members will evaluate the role of the identified miRNA in the regulation of this peptide family.