title: Phosphatidate accumulation in hormone-treated hepatocytes via a phospholipase D mechanism.
abstract: Isolated rat hepatocytes responded to a variety of Ca2+-mobilizing agents (vasopressin, angiotensin II, epinephrine, epidermal growth factor, ATP, and ADP) with a rapid increase in phosphatidate mass, as measured by a sensitive new method. When hepatocytes were incubated with vasopressin (10(-8) M), phosphatidate levels increased 2-3-fold in 2 min, but there was no significant increase in diacylglycerol at this time. Changes in the fatty acid composition of phosphatidate also preceded those in diacylglycerol. De novo synthesis of phosphatidate from [3H]glycerol was unaffected by vasopressin in short-term incubation. Incubation of washed rat liver plasma membranes with GTP gamma S caused a time-dependent increase in phosphatidate. When membranes were incubated with GTP gamma S and [gamma-32P]ATP, no incorporation of 32P into phosphatidate was observed. This excludes the phospholipase C-diacylglycerol kinase pathway and suggests that a phospholipase D activity produced the phosphatidate. At submaximal concentrations of GTP gamma S, ATP and ADP stimulated membrane phosphatidate formation, presumably by acting through P2-purinergic receptors. Only phosphatidylcholine, among the major phospholipids, decreased in the membranes in response to GTP gamma S. The fatty acid composition of the phosphatidate produced in response to vasopressin in hepatocytes also suggests that phosphatidylcholine may be the source of hormonally elicited phosphatidate. We conclude that Ca2+-mobilizing hormones mainly increase phosphatidate levels in hepatocytes by a mechanism that does not involve phosphorylation of diacylglycerol or de novo synthesis but involves a guanine nucleotide-binding protein coupled to phospholipase D.
mesh_terms: D000227:Adenine Nucleotides / Q000494:pharmacology; D000804:Angiotensin II / Q000494:pharmacology; D000818:Animals; D000001:Calcimycin / Q000494:pharmacology; D004815:Epidermal Growth Factor / Q000494:pharmacology; D004837:Epinephrine / Q000494:pharmacology; D005227:Fatty Acids / Q000032:analysis; D006728:Hormones / Q000494:pharmacology*; D066298:In Vitro Techniques; D007700:Kinetics; D008055:Lipids / Q000302:isolation & purification; D008099:Liver / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D008563:Membrane Lipids / Q000032:analysis; D010712:Phosphatidic Acids / Q000378:metabolism*; D010739:Phospholipase D / Q000378:metabolism*; D010740:Phospholipases / Q000378:metabolism*; D010743:Phospholipids / Q000032:analysis; D051381:Rats; D014667:Vasopressins / Q000494:pharmacology
chemical_list: D000227:Adenine Nucleotides; D005227:Fatty Acids; D006728:Hormones; D008055:Lipids; D008563:Membrane Lipids; D010712:Phosphatidic Acids; D010743:Phospholipids; D014667:Vasopressins; D000804:Angiotensin II; D000001:Calcimycin; D004815:Epidermal Growth Factor; D010740:Phospholipases; D010739:Phospholipase D; D004837:Epinephrine