Fig. 7

FOXM1 transcriptionally activates ASPM in HCC cells. A The HCC cell line HepG2 was used to establish ASPM-overexpressing cells by lentivirus infection. Total and endogenous ASPM mRNA levels were detected by qRT-PCR, via the primers at CDS or 3’UTR region in ASPM mRNA (shown in Additional file 1: Fig. S5A). ASPM OE, overexpression of ASPM. EV, empty vector. B HepG2 cells were transfected with the indicated plasmids for 48 h. CHX (50 μg/mL) was then added at 0, 6, 12, 18, and 24 h. Lysates were collected at the indicated time points and immunoblotted with the indicated antibodies. FOXM1 OE, overexpression of FOXM1. EV, empty vector. C The SNU-739 and HepG2 cells were used to establish FOXM1-knockdown cells by lentivirus mediated shRNA delivery system. The protein levels of FOXM1 and ASPM were detected using Western blot. D The qRT-PCR assay was performed to detect the mRNA expression of ASPM and FOXM1 in FOXM1-knockdown cells. The data were presented as the mean ± SD of three independent experiments. The significance was analyzed by one-way ANOVA. **p < 0.01, ***p < 0.001. E The binding regions of FOXM1 on ASPM promoter were predicted by bioinformatics. TSS, Transcriptional Start Site. F Dual luciferase reporter assay of FOXM1 and ASPM promoter. The data were presented as the mean ± SD of three independent experiments. The significance was analyzed by Student’s t test. **p < 0.01. FOXM1 OE, overexpression of FOXM1. EV, empty vector. G The ChIP-qPCR was used to determine the direct binding of FOXM1 on the promoter region of ASPM in SNU-739 cell lines. Design primers according to Fig. 7E. The data were presented as the mean ± SD of three independent experiments. The significance was analyzed by Student’s t test. ns, no significance; **p < 0.01; ***p < 0.001