Figure 2
FBP1 is a degradation substrate of E3 ubiquitin ligase TRIM28. (a) HepG2 and SK-Hep-1 cells were infected with control or two independent TRIM28-specific shRNAs. Cells were harvested for western blot analysis after 48 h transfection. (b) Western blot analysis of whole-cell lysate of HepG2 cells transfected with the indicated constructs. Cells were treated with or without 20 μM of MG132 for 8 h before harvest. (c) HepG2 cells were infected with control and TRIM28-specific shRNAs. After 48 h, cells were treated with 50 μg/μl cycloheximide (CHX). At different time points, cells were harvested for western blot analysis. At each time point, the intensity of FBP1 was normalized to the intensity of ERK2 (loading control) first and then to the value at the 0-h time point. (d) HepG2 cells were transfected with indicated plasmids for 24 h followed by western blot analysis. (e) Western blot analysis of whole-cell lysate of HepG2 cells transfected with the indicated constructs. (f) Western blot analysis of whole-cell lysate of HepG2 cells transfected with the indicated constructs. (g) Western blot analysis of whole-cell lysate of HepG2 cells transfected with the indicated constructs. Cells were treated with or without 20 μM of MG132 for 8 h before harvest. (h) Measurement of glucose consumption and l-lactate production in spent medium of HepG2 cells 48 h after transfected with indicated constructs and western blot analysis of whole-cell lysate of HepG2 cells. *P< 0.01.
