Metabolic impact of adenovirus-mediated overexpression of the glucose-6-phosphatase catalytic subunit in hepatocytes.
Glucose-6-phosphatase (G6Pase) catalyzes the hydrolysis of glucose 6-phosphate (Glu-6-P) to free glucose and, as the last step in gluconeogenesis and glycogenolysis in liver, is thought to play an important role in glucose homeostasis. G6Pase activity appears to be conferred by a set of proteins localized to the endoplasmic reticulum, including a glucose-6-phosphate translocase, a G6Pase phosphohydrolase or catalytic subunit, and glucose and inorganic phosphate transporters in the endoplasmic reticulum membrane. In the current study, we used a recombinant adenovirus containing the cDNA encoding the G6Pase catalytic subunit (AdCMV-G6Pase) to evaluate the metabolic impact of overexpression of the enzyme in primary hepatocytes. We found that AdCMV-G6Pase-treated liver cells contain significantly less glycogen and Glu-6-P, but unchanged UDP-glucose levels, relative to control cells. Further, the glycogen synthase activity state was closely correlated with Glu-6-P levels over a wide range of glucose concentrations in both G6Pase-overexpressing and control cells. The reduction in glycogen synthesis in AdCMV-G6Pase-treated hepatocytes is therefore not a function of decreased substrate availability but rather occurs because of the regulatory effects of Glu-6-P on glycogen synthase activity. We also found that AdCMV-G6Pase-treated-cells had significantly lower rates of lactate production and [3-3H]glucose usage, coupled with enhanced rates of gluconeogenesis and Glu-6-P hydrolysis. We conclude that overexpression of the G6Pase catalytic subunit alone is sufficient to activate flux through the G6Pase system in liver cells. Further, hepatocytes treated with AdCMV-G6Pase exhibit a metabolic profile resembling that of liver cells from patients or animals with non-insulin-dependent diabetes mellitus, suggesting that dysregulation of the catalytic subunit of G6Pase could contribute to the etiology of the disease.
Seoane
J
J
Department de Bioquímica i Biologia Molecular, Facultat de Química, Universitat de Barcelona, E08028 Barcelona, Spain.
Trinh
K
K
O'Doherty
R M
RM
Gómez-Foix
A M
AM
Lange
A J
AJ
Newgard
C B
CB
Guinovart
J J
JJ
D000256:Adenoviridae; D000818:Animals; D002478:Cells, Cultured; D005952:Glucose-6-Phosphatase / Q000096:biosynthesis / Q000378:metabolism*; D019298:Glucose-6-Phosphate / Q000378:metabolism; D006019:Glycolysis; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D008112:Liver Glycogen / Q000378:metabolism; D046911:Macromolecular Substances; D008297:Male; D051381:Rats; D017208:Rats, Wistar; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D014162:Transfection
D008112:Liver Glycogen; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D019298:Glucose-6-Phosphate; D005952:Glucose-6-Phosphatase