Figure 4.

TM4SF5-mediated lipogenic activity upon excessive fructose intake involves GLUT8. (A) HepG2 cells (endogenously expressing TM4SF5) were transiently transduced with lentivirus for either a non-specific sequence (NS) or TM4SF5 target sequences (#2 or #4) together with transfections of either empty vector (EV-FLAG), or GLUT2-HA and GLUT8-FLAG plasmids for 24 h, glucose-starved for 16 h, and then treated with fructose (450 mg/dL) for 30 min prior to whole cell lysate preparation for standard western blot analysis. (B) Tissue extracts from livers of WT or KO mice fed HFD or HFFD for 10 weeks were prepared for immunoblotting. (C) WT or KO mice were fed with HFD for 1 week and siRNA against control or GLUT8 sequence (siGLUT8) was intravenously injected twice per week (2.5 mg/kg/animal). The animals were starved for 5 h and orally injected with excessive fructose (4 g/kg in H₂O), 90 min before preparation of liver tissue extracts and immunoblotting. (D) Huh7 cells (endogenously expressing TM4SF5) were transiently transfected with either TM4SF5-Strep, GLUT2-HA, or GLUT8-FLAG constructs. One day later, the cells were glucose-starved for 16 h before treatment with fructose for 15 min. The cells were then harvested both for co-pulldown experiments using streptavidin-conjugated agarose beads (B) and for standard western blot analysis of the indicated molecules (C). The data shown are representative of three independent experiments.