Diacylglycerol molecular species in plasma membrane and microsomes change transiently with endothelin-1 treatment of glioma cells.
Agonist-induced intracellular signal transduction often involves activation of protein kinase C by diacylglycerol (DAG) released from membrane phospholipids by phospholipases. Using either DAG kinase or HPLC assays to quantitatively determine DAG mass, we observed a time-dependent increase in DAG accumulation upon incubation of rat C6 glioma cells with 200 nM endothelin-1 (ET-1). Total cell DAG rapidly increased by 25-35% from a basal level of 4.5 +/- 0.3 nmol/mg protein during one min of ET-1 treatment and remained constant or slightly decreased between 1 and 2 min. Thereafter, DAG increased to a maximum (1.6-fold above basal) by 5-10 min. and remained elevated to 30 min. Resolution of DAG molecular species by HPLC after incubation of cells with ET-1 revealed that accumulation of DAG species differed in total cell lysate and subcellular compartments. In plasma membrane, major DAG species increased at 1 min. followed by a decrease at 10 min. whereas in microsomes DAG species did not change at 1 min. and decreased at 10 min. Although phospholipid sources of DAG species were not identified specifically, there was preferential hydrolysis of molecular species of phospholipid for DAG production. We propose that molecular species of DAG produced at the plasma membrane may be transferred to the endoplasmic reticulum so that phospholipid resynthesis can replenish molecular species initially utilized in signal transduction.
Laurin
D E
DE
Atlantic Research Center, Department of Pediatrics, Dalhousie University, Halifax, Nova Scotia, Canada.
Byers
D M
DM
Palmer
F B
FB
Cook
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HW
D000804:Angiotensin II / Q000494:pharmacology; D000818:Animals; D001920:Bradykinin / Q000494:pharmacology; D001932:Brain Neoplasms / Q000378:metabolism / Q000473:pathology*; D002462:Cell Membrane / Q000187:drug effects*; D002851:Chromatography, High Pressure Liquid; D004075:Diglycerides / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism; D019332:Endothelin-1 / Q000494:pharmacology*; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D005910:Glioma / Q000378:metabolism / Q000473:pathology*; D006801:Humans; D008563:Membrane Lipids / Q000145:classification / Q000378:metabolism*; D051379:Mice; D008861:Microsomes / Q000187:drug effects; D009496:Neurotensin / Q000494:pharmacology; D010743:Phospholipids / Q000145:classification / Q000378:metabolism*; D051381:Rats; D012701:Serotonin / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects*; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism
D004075:Diglycerides; D019332:Endothelin-1; D008563:Membrane Lipids; D010743:Phospholipids; D000804:Angiotensin II; D012701:Serotonin; D009496:Neurotensin; D001920:Bradykinin