We studied the mechanism whereby insulin activates de novo phosphatidic acid synthesis in BC3H-1 myocytes. Insulin rapidly activated glycerol-3-phosphate acyltransferase (G3PAT) in intact and cell-free preparations of myocytes in a dose-related manner. The apparent Km of the enzyme was decreased by treatment with insulin, whereas the Vmax was unaffected. No activation was found by ACTH, insulin-like growth factor-I, angiotensin II, or phenylephrine, but epidermal growth factor, which, like insulin, is known to activate de novo phosphatidic acid synthesis in intact myocytes, also stimulated G3PAT activity. In homogenates or membrane fractions, the effect of insulin on G3PAT was fully mimicked by nonspecific or phosphatidylinositol (PI)-specific phospholipase C (PLC). An antiserum raised against PI-glycan-PLC completely blocked the effect of insulin on G3PAT. Although the above findings suggested involvement of a PLC in insulin-induced activation of G3PAT, neither diacylglycerol nor protein kinase C activation appeared to be involved. On the other hand, insulin stimulated the release of a cytosolic factor, which activated membrane-associated G3PAT. This cytosolic factor had a molecular weight of less than 5K as determined by Sephadex G-25 chromatography. NaF, a phosphatase inhibitor, blocked the activation of G3PAT by insulin, suggesting involvement of a phosphatase. Insulin-induced activation of G3PAT was also blocked by pretreatment of intact myocytes with pertussis toxin and by prior addition, to homogenates, of an antiserum that recognizes the C-terminal decapeptide of Giα. Our results suggest that insulin activates a pertussis toxin sensitive, Giα-protein-requiring PI-glycan-PLC in BC3H-1 myocytes, resulting in the release of a cytosolic, low molecular weight factor, which decreases the Km of G3PAT, probably by a phosphatasemediated mechanism. This activation of G3PAT may account for insulin-induced increases in de novo synthesis of phosphatidic acid, which, in turn, may amplify diacylglycerol-protein kinase C signaling and provide a mechanism to replenish phospholipids that are hydrolyzed during insulin action. © 1990, American Chemical Society. All rights reserved.
Documento: | Artículo |
Título: | Insulin Activates Glycerol-3-phosphate Acyltransferase (de Novo Phosphatidic Acid Synthesis) through a Phospholipid-Derived Mediator. Apparent Involvement of Giα and Activation of a Phospholipase C |
Autor: | del Vila, M.C.; Milligan, G.; Standaert, M.L.; Farese, R.V. |
Filiación: | Facultad de Ciencias Exactas Naturales, Departamento de Chemica Biologica, University of Buenos Aires, Argentina Department of Biochemistry, University of Glasgow, Glasgow G128QQ, United Kingdom Research Service, James A. Haley Veterans Hospital and Departments of Internal Medicine and Biochemistry and of Molecular Biology, University of South Florida College of Medicine, Tampa, Florida 33612, United States |
Palabras clave: | 1-phosphatidylinositol phosphodiesterase; Bordetella virulence factor; fluoride sodium; glycerol 3 phosphate acyltransferase; Glycerol 3 Phosphate O Acyltransferase; GTP Binding Proteins; guanine nucleotide binding protein; inositol phosphate; inositol phosphate glycan; insulin; phosphatidic acid; phosphatidylinositol phosphodiesterase; phosphodiesterase; polysaccharide; diacylglycerol; glycerol 3 phosphate acyltransferase; insulin; phospholipase c; animal; article; biosynthesis; cell culture; chemistry; cytosol; drug effect; enzyme activation; kinetics; metabolism; mouse; muscle; signal transduction; muscle cell; nonhuman; priority journal; signal transduction; Animal; Cells, Cultured; Cytosol; Enzyme Activation; Glycerol-3-Phosphate O-Acyltransferase; GTP-Binding Proteins; Inositol Phosphates; Insulin; Kinetics; Mice; Muscles; Phosphatidic Acids; Phosphoric Diester Hydrolases; Polysaccharides; Signal Transduction; Sodium Fluoride; Support, Non-U.S. Gov't; Support, U.S. Gov't, P.H.S.; Virulence Factors, Bordetella |
Año: | 1990 |
Volumen: | 29 |
Número: | 37 |
Página de inicio: | 8735 |
Página de fin: | 8740 |
DOI: | http://dx.doi.org/10.1021/bi00489a033 |
Título revista: | Biochemistry |
Título revista abreviado: | Biochemistry |
ISSN: | 00062960 |
CAS: | fluoride sodium, 51668-54-3, 7681-49-4, 79933-27-0; glycerol 3 phosphate acyltransferase, 9029-96-3; inositol phosphate, 15421-51-9; insulin, 9004-10-8; 1-phosphatidylinositol phosphodiesterase, EC 3.1.4.10; Glycerol-3-Phosphate O-Acyltransferase, EC 2.3.1.15; GTP-Binding Proteins, EC 3.6.1.-; inositol phosphate glycan; Inositol Phosphates; Insulin, 11061-68-0; Phosphatidic Acids; Phosphoric Diester Hydrolases, EC 3.1.4; Polysaccharides; Sodium Fluoride, 7681-49-4; Virulence Factors, Bordetella |
Registro: | https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_00062960_v29_n37_p8735_delVila |