Fig 2: KIAA0101 is a critical target of FoxM1 in promoting migration and invasion of liver cancer cells. After transfection with si-FoxM1 and cotransfection with si-FoxM1 and KIAA0101 as well as NC, the protein and mRNA expression levels of FoxM1 and KIAA0101 in HepG2 and Huh7 cells were measured by western blot (A) and qRT-PCR (B). A transwell assay was applied to assess the invasion and migration ability of the transfected cells. Cell counts from five random microscopic fields were analyzed (C and D. Magnification: ×200). Migration ability was also assessed by a wound-healing assay (E and F. Magnification: ×40). All data are presented as the means±SDs of at least three independent experiments. *P<0.05, **P<0.01, compared with the NC group; #P<0.05, ##P<0.01, compared with the si-FoxM1 group.

Fig 3: FoxM1 and KIAA0101 were both upregulated in HCC samples. Western blot (A), qRT-PCR (B), and immunohistochemical staining (C) showed the expression of FoxM1 and KIAA0101 in HCC tissues and adjacent normal tissues. N: normal tissue; T: HCC tissue. Magnification: ×50 and ×400. All data are presented as the means±SDs of at least three independent experiments. *P<0.05, **P<0.01.

Fig 4: KIAA0101 promotes the migration and invasion of liver cancer cells. After transfection with si-KIAA0101 as well as NC, the protein and mRNA expression of KIAA0101 in HepG2 and Huh7 cells was measured by western blot (A) and qRT-PCR (B). A transwell assay was applied to assess the invasion and migration ability of the transfected cells. Cell counts from five random microscopic fields were analyzed (C and D. Magnification: ×200). Migration ability was also assessed by a wound-healing assay (E and F. Magnification: ×40). All data are presented as the means±SDs of at least three independent experiments. *P<0.05, **P<0.01, compared with the NC group; #P<0.05, ##P<0.01, compared with the si-FoxM1 group.

Fig 5: KIAA0101 is a direct target of the transcriptional factor FoxM1. After transfection with si-FoxM1 or negative control (NC), the protein and mRNA expression of FoxM1 and KIAA0101 in HepG2 and Huh7 cells were measured by western blot (A) and qRT-PCR (B and C). Schematic illustration of the KIAA0101 promoter region and the binding site of FoxM1. The KIAA0101 promoter was determined by using https://genome.ucsc.edu. The sequence is numbered from the transcription start site (exon 1). The primer set was designed to amplify the region of the promoter to amplify the potential binding site (D). ChIP assays with anti-FoxM1 antibody or IgG were performed to verify the binding between FoxM1 and the KIAA0101 promoter in HepG2 and Huh7 cells. After the reversal of cross-linking, the coimmunoprecipitated DNA was amplified by PCR using primers amplifying the FoxM1 binding-site-containing region (E). In addition, a representative gel image of the ChIP results is presented (F). The WT and MUT promoter sequences of the putative promoter regions (G). A luciferase assay revealed that FoxM1 regulated KIAA0101 mRNA expression by binding to the promoter of KIAA0101 (H). All data are presented as the means±SDs of at least three independent experiments. *P<0.05, **P<0.01.