Figure 6

Requirement of Snail for the miR-130b antagonism effect on downstream gene expressions in vitro.

(a) MiR-130bi restored SNAIL expression but further downregulated E-CADHERIN induced by Snail siRNA as shown by immunofluorescence and (b) quantification of the staining intensity. Arrowheads indicated co-localized SNAIL and E-CADHERIN. (c) MiR-130b inhibitor attenuated the silencing effect of Snail siRNA on the expression of Snail, Vimentin and Collagen IV and downregulated E-cadherin by qRT-PCR and (d) Western blot analyses. (e) MiR-130b and its putative binding sequence in the 3′-UTR of Snail. The mutant Snail binding site was generated in the complementary site for the seed region of miR-130b. (f) MiR-130b inhibitor led to a noticeable increase in the luciferase activity of wt 3′-UTR of Snail. Results are presented as mean ± SD of three independent experiments. *P < 0.05; ^#P < 0.0001. NT: non-targeting; siRNA: small interfering RNA; miR-iNC: miRNA inhibitor negative control; miR-130bi: miR-130b inhibitor; wt: wild type; mt: mutant type.