2.1. Cell Lines, Constructs and Primers

Human embryonic kidney 293 cells, and human HCC cell lines that included Huh-7, SMMC-7721, MHCC-97L and EHBC-512, were maintained as previously described [24]. The ASPH coding sequence was amplified from a cDNA library of EHBC-512. The enzymatic loss ASPH mutant was prepared through site mutagenesis of histidine-679, a reported essential residue for ASPH catalytic activities [19,25], to alanine using an in vitro mutagenesis system (Promega, Madison, WI). The coding sequence of vimentin was synthesized by Shanghai Genepharma (Shanghai, China). The shRNA sequences for silencing ASPH and vimentin were 5′-GCGCAGTGTGAGGATGAT-3′ and 5′-GCTAACTACCAAGACACTATT-3′, respectively. The lentivirus of vimentin, wild-type (WT) or mutant ASPH and the shRNA against human ASPH or vimentin were constructed, packaged and harvested by Shanghai Genepharma. The FLAG-ASPH and HA-vimentin plasmids were constructed through the introduction of ASPH and vimentin coding sequence into pFLAG-CMV (Sigma, St. Louis, MO) and pCMV-HA vectors (Clontech, Kusatsu, Shiga Japan). The primers are listed in Table S1.