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. 2018 Dec 17;17(3):1509–1516. doi: 10.3892/etm.2018.7106

Figure 3.

Figure 3.

SP1 is a novel target gene of miR-375. (A) Bioinformatics software was used to identify a conserved binding site for miR-375 in the 3′UTR of SP1. The sequence highlighted in red was the mutant sequence used to demonstrated binding between miR-375 and 3′UTR of SP1. (B) KYSE450 and KYSE150 cells were co-transfected with miR-375 mimic or negative control miRNA and luciferase reporter constructs SP1 3′UTR or SP1 3′UTR (mut). The relative luciferase activity was measured using the Dual-Luciferase Reporter Assay System. (C) The protein expression level of SP1 was determined by western blot analysis in tumor and matched adjacent healthy tissue samples from patients with ESCC. The expression level of SP1 was normalized to GAPDH. (D) The protein expression level of SP1 was determined by western blot analysis in ESCC cell lines KYSE450 and KYSE150 and the normal esophageal endothelial cell line Het-1A. (E) The protein expression level of SP1 was determined by western blot analysis in ESCC cell lines following transient transfection with miR-375 mimic or NC miRNA. *P<0.05, **P<0.01 ***P<0.001 and as indicated. (F) An inverse correlation between miR-375 and SP1 protein expression in ESCC cells was identified (Pearson's correlation r=−2.106; P<0.05). NS, not significant miR, microRNA; SP1, specificity protein 1; ESCC, esophageal squamous cell carcinoma; OD, optical density; 3′UTR, 3′untranslated region; mut, mutant; NC, negative control; KYSE450 and KYSE150, ESCC cell lines; Het-1A, normal esophageal endothelial cell line; miR-375 mimic, ESCC cell (KYSE450 or KYSE150) transfected with miR-375 mimic; NC miRNA, ESCC cell (KYSE450 or KYSE150) transfected with NC miRNA.