Effect of glutathione deficiency on the adipocyte Mg(2+)-dependent phosphatidate phosphohydrolase.
Previous studies from this laboratory [Jamdar S. C. and Cao W. F. (1994) Biochem. J. 301, 793-799] show that the adipocyte Mg(2+)-dependent phosphatidate phosphohydrolase (MGPPH), a major regulatory enzyme in adipose triacylglycerol metabolism, requires an active thiol group for its activity and perturbation of this group results in the loss of enzyme activity. Since glutathione (GSH) is important in maintaining the intracellular thiol state, we have used GSH-deficient animals and adipocytes to test the possibility that intracellular GSH concentration is critical in controlling the MGPPH activity. The MGPPH was measured in the presence of aqueous dispersed phosphatidate, and the release of P1 was taken as a measure of enzyme activity. The GSH deficiency in animals and isolated adipocytes was produced in the presence of diethylmaleate (DEM) or buthionine sulfoximine (BSO). Intraperitoneal administration of BSO into animals (3 mmoles/kg) showed 10-25% reduction in the blood and adipose GSH and 25% decline in the adipose MGPPH activity. However, DEM (0.3 ml/kg) was more effective and caused over 70% reduction of the blood and adipose tissue GSH content and 75% decline in the adipose MGPPH activity within 4 hr of drug administration. After 24 hr, these values returned to normal. Adipocytes incubated with 2.5 mM DEM for 60 min at 37 degrees C also showed a significant reduction in the GSH content and the MGPPH activity present in the cytosol and membrane fractions. The loss of membrane MGPPH was associated with decreased rates of triacylglycerol formation from [14C]palmitate. Pre-incubation of adipocyte homogenates with 1 mM DEM also resulted in > 90% decline in the MGPPH activity, which was preventable in the presence of GSH and dithiothreitol. Therefore, these studies suggest that the sulfhydryl environment offered by glutathione is critical for the maintenance of adipocyte MGPPH activity.
Jamdar
S C
SC
Department of Anesthesiology, College of Physicians and Surgeons, Columbia University, New York, NY 10032, USA.
Cao
W F
WF
D017667:Adipocytes / Q000378:metabolism*; D000818:Animals; D019328:Buthionine Sulfoximine / Q000494:pharmacology; D005978:Glutathione / Q000172:deficiency*; D008298:Maleates / Q000494:pharmacology; D010711:Phosphatidate Phosphatase / Q000378:metabolism*; D051381:Rats
D008298:Maleates; D019328:Buthionine Sulfoximine; D010711:Phosphatidate Phosphatase; C014476:diethyl maleate; D005978:Glutathione