ID	Duplex structure	Position
1	miRNA 3' cguguCGGGGGCAGGGAGGGa 5' ||||| | ||||||| Target 5' ctgtcGCCCCAGGCCCTCCCt 3'	3 - 23

- Mol Carcinog, 2010

microRNAs (miRNAs) play vital roles in several biological processes, including apoptosis, by negatively regulating the expression of target genes. The molecular mechanisms of the key survival signal, Akt family, have been widely explored. However, it remains to be ascertained whether Akt1, the predominant isoform in most tissue, is a direct target of miRNA. In this study, we identified Akt1 and E2F1 to be two direct targets of miR-149* and b-Myb to be an indirect target by reporter assays and Western blot analyses. Ectopic expression of miR-149*-induced apoptosis in Be2C, a neuroblastoma cell line, and in HeLa cells. Silencing of Akt1 or E2F1 expression also led to similar apoptotic changes in these two cell lines, suggesting that the pro-apoptotic effects of miR-149* were exerted by repressing Akt1 and E2F1 expressions. Importantly, analysis of primary neuroblastoma samples revealed a significant inverse correlation of miR-149* with E2F1 expressions (P = 0.026). Interestingly, using the reporter assays, excess miR-149 introduced by transfection to simulated its preponderance in the in vivo condition, could not overcome the repressive function of miR-149* on the target genes. This implies that the pro-apoptotic function of miR-149* may not be dampened by its predominant cognate, miR-149, in vivo. Our findings not only provided the first evidence that Akt1 is a direct target of miRNA but also demonstrated that miR-149* is a pro-apoptotic miRNA by repressing the expression of Akt1 and E2F1. (c) 2010 Wiley-Liss, Inc.