Fig. 5

KIFC1 interacts with HMGA1 and regulates its transcriptional activity in HCC cells. a A silver staining assay was used to distinguish differentiated proteins between control and KIFC1-overexpressing HepG2 cells. HMGA1 (17 kD) is indicated by a black bar. b HEK-293 T cells were transfected with Flag-KIFC1 or Myc-HMGA1 or both vectors. A coIP assay was performed to test the interaction (upper part). HepG2-vector and HepG2-KIFC1-Flag cells were also tested by the coIP assay (lower part). Flag-tag antibody (14793 s, Cell signaling technology, 1:1000) and Myc-tag antibody (M192-3B, MBL, 1:1000) were used in the upper part. Flag tag antibody and HMGA1 (ab129153, Abcam, 1:1000) were used in the lower part. c The fold change in HMGA1 mRNA expression levels in 40 paired HCC and adjacent nontumor tissues. The correlation between KIFC1 and HMGA1 was tested in 40 paired specimens and data from TCGA cohort. Data are presented as the mean ± SD, * P < 0.01, ** P < 0.001. d IF assay demonstrated the colocalization of KIFC1 and HMGA1 in HepG2 and 7701 cells. The bar indicated 40 μm. e ChIP assay was performed using HMGA1 antibody (ab129153, Abcam). IgG was used as the negative control. Quantitative PCR was conducted at the promoter regions of Twist1, vimentin, Stat3, MMP2, and E-cadherin in 7701-shnc and HepG2 vector cells. f Binding levels of HMGA1 and E-cadherin, vimentin, Twist1, MMP2 and Stat3 promoters after KIFC1 knockdown or overexpression