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Fig. 5 | BMC Cancer

Fig. 5

From: Micro-RNA-186-5p inhibition attenuates proliferation, anchorage independent growth and invasion in metastatic prostate cancer cells

Fig. 5

Tumor suppressor AKAP12 is a target of miR-186-5p in PCa cells. AKAP12 transcript and protein expression were measured using qRT-PCR and western blot analysis, respectively. a Three potential miR-186-5p binding sites in AKAP12 were identified; site 1) mirSVR score: − 0.5641 PhastCons score: 0.6188; site 2) mirSVR score: − 0.7981 PhastCons score: 0.5553; and site 3) mirSVR score: − 0.2640 PhastCons score: 0.6218 (values from www.microrna.org). b AKAP12 transcript expression was significantly lower in metastatic LNCaP (p = 0.0011) and MDA-PCa-2b (p < 0.0001) cells and higher in PC-3 cells relative to normal prostate RWPE1 cells (p = 0.0011). c Stable anti-miR-186-5p expression in PC-3 cells increased AKAP12 transcript expression (p = 0.0013). AKAP12 expression was normalized to GAPDH. d HEK 293 T cells were transfected with scrambled miR control or miR-186-5p mimic. A representative western blot shows ectopic expression of miR-186-5p in HEK 293 T cells decreased AKAP12 protein by 30% 72 h post-transfection (p = 0.0063). All analyses involved at least three independent experiments. e and f Representative western blots of AKAP12, β-catenin and pAKT expression in HEK 293 T (72 h), PC-3 (48 h), and MDA-PCa-2b (24 h) cells post-transfection. Data analyses included three independent experiments for e and two experiments for f. g and h Quantitation of western blots presented as black bar (scramble control) and grey bar (miR-186-5p inhibitor). AKAP12 protein was increased in PC-3 (p = 0.0049) and MDA-PCa-2b (p = 0.0318) cells transiently transfected with miR-186-5p inhibitor. In contrast, β-catenin/ β-actin protein was decreased in PC-3 (p = 0.0434) and MDA-PCa-2b (p = 0.0048) cells transfected with miR-186-5p inhibitor. Data analysis was based on mean ± S.D. of target protein relative to β-actin in the same blot. (*p-value < 0.05, **p-value < 0.007, **** p-value < 0.002)

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