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. 2014 Oct 24;5(21):10870–10885. doi: 10.18632/oncotarget.2539

Figure 6. Msi1 Binds to the 3′UTR of p21, p27 and p53 and represses their translation.

Figure 6

(A) Secondary RNA structure surrounding the Msi1 binding sites in p21, p27 and p53 as predicted by the RNAstructure 4.6 software. p21 has only one proven Msi1 binding site, p27 has two predicted Msi1 binding sites (p27–1 and p27–2), and p53 has two predicted Msi1 binding sites (p53–1 and p53–2). (B) Schematic representation of the reporter construct containing the firefly luciferase coding sequence fused to the p21 3′UTR/p27 3′UTR/p53 3′UTR. The predicted binding site regions within the wild-type p21 3′UTR/p27 3′UTR/p53 3′UTR are noted in colored ovals, and the residues altered in the p21 3′UTR/p27 3′UTR/p53 3′UTR mutants construct are marked on the ovals. Luciferase reporter assays are shown for p21, p27 and p27 targeted by Msi1 in HeLa-Msi1 (C) and SiHa-Msi1 cells (D). The histogram shows the ratio of firefly to Renilla luciferase activity normalized to empty vector-transfected cells (baseline). The data are shown as the mean± S.E.M. *P < 0.05. (E) The RNA-Msi1 binding assay shows associations of binding sites of p21, p27–1, p27–2, p53–1 and p53–2 with Msi1 protein in HeLa-Msi1 cells(upper) and SiHa-Msi1 cells(lower). (F) The expression of Msi1 and p21, p27 and p53 in 13 cervical cancers specimens. The expression levels of two representative samples are shown. The Msi1, p21, p27 and p53 staining were scored from 1 to 12. The correlation was determined by Pearson's correlation test. (G) The correlation between Msi1 and p21 was significant (r=-0.5907, P=0.0335). (H) The correlation between Msi1 and p27 was significant (r=-0.5586, P=0.0472). (I) The correlation between Msi1 and p53 was significant (r=-0.5590, P=0.0470).