FIGURE 7.

GnT-III overexpression in HepG2 cells significantly increased cell migration, but knock-out of ST6GAL1 in this cell line conferred GnT-III with the opposite effect on cell motility. Overexpression of ST6GAL1 was able to neutralize the anti-migratory role of GnT-III in HeLa cells. A, cell lysates from the control and GnT-III-overexpressing HepG2 cells were immunoblotted with E4-PHA and DSA lectins to check the overexpression efficiency of GnT-III. B, their cell migration toward FN was determined by Transwell assay. Cells that migrated through the Transwell membrane were stained with 0.5% crystal violet. Shown is a representative example recorded by phase contrast microscopy. Scale bar, 100 μm. C, the migrated cells were counted under a microscope. The quantitative data were obtained from three independent experiments. D, ST6GAL1 knock-out HepG2 cells were incubated with (bold line) or without (gray shading) biotin-conjugated SNA, followed by incubation with streptavidin Alexa Fluor 647 conjugate and subjected to FACS analysis. E, the migratory ability of cells indicated toward FN was determined by Transwell assay. Cells that migrated through the Transwell membrane were stained with 0.5% crystal violet. Shown is a representative example recorded by phase contrast microscopy. Scale bar, 100 μm. F, the migrated cells were counted under a microscope. The quantitative data were obtained from three independent experiments. The p values were calculated using one-tail unpaired t test. Error bars indicate standard derivation. *, p < 0.01. Con, control; OE, GnT-III-overexpressing cells.