Fig. 6

The D1 domain is the key region for VCP exhibiting biological function in HCC. A The scheme of VCP mutants. The WT VCP including 806 amino acids was divided into four domains (△N, △D1, △D2, and △C tail). The gray area indicated the deleted region. B 293T cells were transfected with His-tagged full-length VCP and its mutants with deletion of various regions. Cell lysates were immunoprecipitated with anti-His antibody. And western blot was used to detect the expression of His probe and HMGB1. C The scheme of HMGB1. The full length of HMGB1 included 215 amino acids and was divided into three regions including A box, B box, and C-tail. The box with gray color indicates the deleted domain of HMGB1. D 293 T cells were transfected with WT HMGB1 and its various mutants with the expression of the Flag probe. Cell lysates were collected for immunoprecipitation with anti-Flag antibody. Then the expression of Flag probe and VCP were determined by western blot. E The protein expression of WT VCP and D1 mutant in MHCC-LM3 cells. F The ability of cell proliferation was measured by CCK8 assay in MHCC-LM3 cells with transfection of WT VCP and D1 mutant, separately. G The movement capacity of MHCC-LM3 cells with transfection of WT VCP or D1 mutant was evaluated by wound-healing assay. H Transwell system indicates the cell's ability of invasion and migration in various groups. The representative images and the corresponding statistical results were shown. WT: wild type. All ***P < 0.001, ****P < 0.0001, and ns no significance