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. Author manuscript; available in PMC: 2019 Apr 1.
Published in final edited form as: Mol Cancer Res. 2018 Jan 12;16(4):669–681. doi: 10.1158/1541-7786.MCR-17-0321

Figure 6. RBL1, KLF5, TIMP2, and SMAD4 are targeted by miR-888 in prostate cells.

Figure 6

A, Luciferase reporter assays measured renilla/firefly levels of prostate cells transfected with the dual luciferase psiCHECK2 vector that contained cloned 3′UTR elements (harboring predicted miR-888 binding sites) for RBL1, SMAD4 (MiRNA Complementary Site 1 or 2), TIMP2, or KLF5 downstream of the renilla cassette. PC3-N cells stably overexpressing miR-888 (lenti-888) significantly repressed renilla expression compared to cells stably overexpressing SCR control mimics (lenti-SCR). Reduced mRNA (B) and protein (C) levels of TIMP2, SMAD4, RBL1 and KLF5 were observed in vitro for PC3-N lenti-888 cells relative to lenti-SCR control cells. D, Western blot analysis of paired PC3-N prostate tumors overexpressing miR-888 or SCR control mimics grown subcutaneously for 5 weeks in the flanks of ten NOD/SCID mice. TIMP2 and SMAD4 protein was reduced in miR-888 treated tumors compared to SCR tumors. Expression was normalized to human 18S rRNA (qRT-PCR) or GAPDH protein (Western blot).