Suppression of miR-92a–induced erythroleukemogenesis by miR-17 is associated with target gene down-regulation. (A) Survival analysis of mice injected with miR-92a (n = 20), miR-17 (n = 18), miR-17+miR-92a (n = 26) and empty vector control (n = 17) retroviruses. (B) Quantitative RT-PCR analysis of precursor miR-17 expression in pooled-populations of SB1 cells infected with miR-17 or empty vector retroviruses. Results were normalized relative to β-actin expression in all experimental samples. Each experiment was performed in quadruplicate, where error bars represent SD; **P < .005. (C) Proliferation rates of miR-17 and control vector infected SB1 cells, as determined on the indicated days using the Trypan blue exclusion assay. (D) p53, Gata-1, (E) STAT-5, Bcl-2, and Jak2 protein expression in SB1 cells transduced with miR-17 or control vector retroviruses. TK6 cells were used as a positive control for p53 expression. Schematic representation (top panel) of the bcl-2 3′UTR (E), STAT-5 3′UTR (F), and jak2 3′UTR (G) containing potential miR-17 binding sites. Positions of mutations generated within these binding sites (bcl-2-mu, STAT-5-mu, and jak2-mu) are indicated in gray. Luciferase assays (bottom panel) of the reporter vector containing the bcl-2 3′UTR (E), STAT-5 3′UTR (F), jak2 3′UTR (G), and their corresponding mutations after transient transfection in NIH-3T3 cells stably expressing miR-17. Data are representative of 5 separate experiments, where error bars represent SD; **P < .005. The expression is presented relative to that obtained with transfection of control vector alone.