BMC Genomics 2005, 6:88 http://www.biomedcentral.com/1471-2164/6/88 DFirgousroeph1ila let-7 miRNA targets the abrupt 3' UTR Drosophila let-7 miRNA targets the abrupt 3' UTR. A. Predicted sites of let-7 interaction with the ab 3' UTR. The schematic at top shows the relative positions of the sites as vertical bars in the ab 3' UTR. The predicted pairings are shown below, with the free energies (in kcal/mol) and exact positions in the ab 3' UTR indicated. B. Luciferase expression in transfected S2 cells. Expression from the luciferase/ab mRNA with no added miRNA is shown at left, set to a relative value of 1. The other bars indi- cate the results of coexpression with let-7 or miR-92b. In this figure and in Figure 2, all values represent the average luciferase expression from at least 5 experiments, and error bars represent standard deviation. can repress expression in the assay system of a reporter mRNA bearing the 3' UTR of a predicted miR-92b target (below), but has only a small effect on luc/ab expression, much less than the effect of let-7 (Fig. 1B). We conclude that let-7 is specifically targeting the ab 3' UTR. Two additional predicted targets were tested for miRNA- dependent regulation. The CrebA mRNA is predicted to have three target sites for miR-92b and mir-312 (Table 1); these miRNAs are closely related, sharing the same nts in positions 2–9 and 15–21. Both miRNAs repress expres- sion of the luc/CrebA reporter (Figure 2B). The Eip74EF mRNA is predicted to have three target sites for miR-34 (Table 1; Figure 2A), and miR-34 represses expression of the luc/Eip74EF reporter (Figure 2B). Flexible MovingTargets search to identify potential miRNAs targeting specific genes In addition to validating a subset of our predicted targets, we wondered if we could use the flexibility of MovingTar- gets to identify a miRNA that regulates a gene known to be under another form of translational control. One such gene is tramtrack (ttk), which encodes a transcription fac- tor that determines non-neuronal identity in developing Page 7 of 11 (page number not for citation purposes)