FIG. 3.

PPP1R3G stimulates glycogen synthesis in hepatocytes. A: Primary hepatocytes isolated from overnight-fasted mice were cultured overnight and then treated with 25 multiplicity of infection (MOI) of Ad-GFP or Ad-PPP1R3G with a Flag tag at the N-terminus. Twenty-four hours later, the cells were harvested for immunoblotting with antibodies as indicated. B: Primary hepatocytes isolated from overnight fasted mice were infected with Ad-GFP or Ad-PPP1R3G at different MOI, followed by glycogen measurement. ***P < 0.001 in comparison with the Ad-GFP group. C: The cells as in A were treated for 24 h with various glucose concentrations, followed by measurement of glycogen content. Data are shown as mean ± SD. **P < 0.01 as comparison between the two groups. D and E: The cells as in A were treated with 100 nmol/L insulin (for D) or 25 μmol/L forskolin (for E) for 24 h, followed by measurement of glycogen content. Data are shown as mean ± SD. **P < 0.01 and ***P < 0.001 as comparison between the treated and untreated groups. F: Hepatocytes isolated from overnight-fasted mice were cultured overnight and then treated with 25 MOI of Ad-shRNA as indicated. Cells were harvested 72 h later and analyzed by RT-qPCR. The fold change of PPP1R3G mRNA in Ad-Scrambled shRNA group is set to 1. **P < 0.01 in comparison with the Ad-Scrambled shRNA group. G: Cell lysate from hepatocytes of G was used in immunoblotting with the antibodies as indicated. H: Cells as in G were used to measure glycogen content. ***P < 0.001 as compared with the scrambled shRNA group. I: Primary hepatocytes isolated from overnight-fasted mice were cultured overnight and then treated with 25 MOI of adenovirus as indicated. Forty-eight hours later, the cells were treated at various glucose concentrations for 24 h, followed by measurement of glycogen content. Data are shown as mean ± SD. *P < 0.05 and **P < 0.01 compared between the two groups.