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Figure 4 | Molecular Cancer

Figure 4

From: miR-300 inhibits epithelial to mesenchymal transition and metastasis by targeting Twist in human epithelial cancer

Figure 4

Twist is a direct target of miR-300. A. The predicted binding of miR-300 with Twist 3′UTR was illustrated. B. A dual-luciferase reporter system analysis was performed to validate miR-300 target genes. A 3′UTR fragment containing the predicted miR-300 targeting sites of Twist was fused downstream of the firefly luciferase gene. A miR-300 mutated binding site (mut) was also constructed. Cotransfection of Twist 3′UTR with miR-300 mimic led to a strong reduction of the luciferase signal. Mutagenesis of the seed region resulted in a reversal of this inhibition. C. Endogenous study showed that cotransfection of Twist 3′UTR with miR-300 inhibitor caused an elevated luciferase signal in both HN-4 and MCF-7 cells. D, E. Western blot analysis of Twist protein expression in HN-12 and MDA-MB-231 cells treated with miR-153 mimic (D) or in HN-4 and MCF-7 cells treated with miR-153 inhibitor F, G, H, I. Overexpression of Twist rescued the repressive effects of miR-300 on EMT, leading to morphological changes (F), molecular changes consistent with EMT (G, H), and elevated invasive abilities in transwell assays (I). * P < 0.05.

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