Figure 3. miR-204 binds to both UCA1 and CXCR4 in vitro and in vivo.

(A) Schematic representation of luciferase reporter constructs. The PGK promoter drives constitutive transcription of a chimeric mRNA containing the firefly luciferase coding sequence fused to the wild-type or mutated UCA1 and CXCR4 3′-UTRs. (B) and (C) Relative activity of the luciferase gene fused with the wild-type or mutant UCA1 3′-UTR in PC-3 and DU-145 cells. (D) Relative activity of the luciferase gene fused with wild-type of CXCR4 in PC-3 and DU-145 cells. The data was normalized to renilla luciferase activity. The data are represented as means ± S.D. from separate transfections (n=3). Statistical analysis was performed using Mann–Whitney U test to compare fold changes between transfection and control groups. All statistical tests were two-sided. **P<0.01. (E) RNA binding protein immunoprecipitation (RIP) assay was performed to investigate the interaction between UCA1, miR-204, and Ago2 protein. The pull-downed RNA was used to analyzed relative expression of miR-204 and UCA1 in PC-3 and DU-145 cells (n=3). **P<0.01. Mann–Whitney U test was used to compare fold changes between control and Ago2 groups.

Luc%, percentage of relative luminescence; Mut, mutant type; WT, wild type.