Figure 5

ZEB1 is another miR-223 target gene, functioning as an inhibitor of myoblast differentiation. (a) The potential binding site of miR-223 in the ZEB1 mRNA 3′ UTR. (b) The potential binding site (red) of miR-223 in the ZEB1 mRNA 3′ UTR is highly conserved among vertebrates. (c) Dual-luciferase reporter assay indicated that miR-223 can bind to the predicted binding site of the ZEB1 mRNA 3′ UTR. (d) miR-223 overexpression inhibited ZEB1 mRNA and protein expression in CPM. (e) miR-223 overexpression inhibited ZEB1 mRNA expression in qm-7. (f) miR-223 inhibition promoted ZEB1 mRNA and protein expression in CPM. (g) miR-223 inhibition promoted ZEB1 mRNA expression in qm-7. (h) ZEB1 protein expression during CPM differentiation. (i) ZEB1 inhibition reduced ZEB1 mRNA and protein expression in CPM. (j) ZEB1 inhibition reduced ZEB1 mRNA expression in qm-7. (k) Expression level of muscle differentiation marker genes after CPM transfected with ZEB1 siRNA. (l) Expression level of muscle differentiation marker genes after qm-7 transfected with ZEB1 siRNA. (m) CPM was co-transfected with miR-223 and pSDS-IGF2, miRNA mimic NC and control vector, miR-223 and control vector, respectively, and expression level of muscle differentiation marker genes was then analyzed. Results are shown as the mean±S.E.M. of three independent experiments. Independent sample t-test was used to analysis the statistical differences between groups. *P<0.05; **P<0.01