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. 2018 Aug 7;8(16):4409–4428. doi: 10.7150/thno.26467

Figure 5.

Figure 5

Figure 5

Suppression of Chi3L1 expression by miR-125a-3p induction is associated with USF1. (A) ChIP analysis for USF1 and Chi3L1 promoter binding activity. Relative binding activity of USF1 and the Chi3L1 promoter was analyzed by qPCR. *, p<0.05; **, p<0.01. (B) Chi3L1 transcriptional activity was measured by luciferase expression using transfection of luciferase-expressing plasmid DNA vector. Luciferase activity was analyzed 12 h after siUSF1 treatment. *, p<0.05. (C) miR-24, -125a-3p, and -342-3p levels were measured in lung tumor tissues of Chi3L1 KD mice compared to normal mice. *, p<0.05. (D) Relative expression of Chi3L1 mRNA in miR-24- and miR-125a-3p mimic-treated A549 and H460 cell lines. *, p<0.01. (E) Relative expression of Chi3L1 mRNA in siUSF1-treated A549 and H460 cell lines with miR-24 and miR-125a-3p mimic. *, p<0.05; #, p<0.05. (F) Relative Chi3L1 mRNA levels by 3'UTR-contained luciferase vector in A549 and H460 cell lines. WT vector contains original Chi3L1 mRNA 3'UTR. MT vector contains miR-125a-3p binding site deleted Chi3L1 mRNA 3'UTR. * p<0.05. (G) Expression of miR-125a-3p in Chi3L1 KO mice. *, p<0.05. (H) Expression of miR-125a-3p by treatment with rhChi3L1 in A549 and H460 cells. *, p<0.05. (I) Expression of miR-125a-3p in siChi3L1 and siUSF1 co-treated A549 and H460 cell lines. *, p<0.05; #, p<0.05. (J) miR-125a-3p transcriptional activity in siUSF1 or 6×Myc-tagged USF1-transfected A549 and H460 cell lines. *, p<0.05; #, p<0.05; ##, p<0.01. (K) Expression of miR-125a-3p in USF1 KD mice. **, p<0.01. All experiments were repeated three times, and samples in each experiment were prepared in duplicate.