Analysis of conserved microRNA targets in the nematode Caenorhabditis elegans and the fruit fly Drosophila melanogaster.

MicroRNA (miRNA) is small regulatory non-coding single stranded RNA molecule that can repress protein expression either at transcriptional or translational level. Since their discoveries in nematodes in the early 1990´s extensive research have shown that this mechanism is conserved across species. Because the miRNA is so small, about 22 nucleotides (nt) long and only requires a minimum of 6nt to interact imperfect with its intended target 3´UTR, therefore a single miRNA could potentially have hundreds of potential targets, which have been suggested by computational prediction. The goal of the project is to experimentally verify three predicted Caenorhabditis elegans mir-2 miRNA­ targets in cell culture, with as candidate targets fos-1, mek-1 and sel-5. In addition C. elegans mir-2 and its mechanism is conserved in Drosophila Melanogaster, miR-2. We want to elucidate if not only mir-2 miRNA is cross species conserved but also it targets. To test this hypothesis we selected the following predicted mir-2 target candidate genes: C. Elegans iff-1 and Drosophila Melanogaster protein ortholog eIF-5A. Validation of miRNA and its functionality was done by transfecting cells with a luciferase-3´UTR reporter only or luciferase-3´UTR and a miRNA-expression plasmid. After the reporter gene was induced, cells were harvested and the luciferase activity measured and the results normalized and compared. Unfortunately our data were inconclusive and future experiments are needed to give a clear picture