title: The activation of phospholipase D by endothelin-1, angiotensin II, and platelet-derived growth factor in vascular smooth muscle A10 cells is mediated by small G proteins of the ADP-ribosylation factor family.
abstract: We show here that A10 cells express the phospholipase D (PLD) isoforms PLD1b and PLD2. The activation of PLD in these cells by angiotensin II (AngII), endothelin-1 (ET-1), and platelet-derived growth factor (PDGF) was found to be sensitive to inhibitors of the activation of ADP-ribosylation factor (ARF) but not to blockers of Rho protein function. PDGF, AngII, and ET-1 induced the binding of ARF proteins to cell membranes in a permeabilized cell assay. Cells permeabilized and depleted of ARF were no longer sensitive to stimulation with AngII, ET-1, or PDGF, but the addition of recombinant myristoylated human ARF1 restored agonist-dependent PLD activity. Expression of dominant negative ARF mutants blocked receptor-dependent activation of PLD. PLD activity was also potently stimulated by treatment with phorbol esters, but this activity was only partially inhibited by brefeldin A or by the overexpression of ARF dominant negative mutants. Transient expression of catalytically inactive mutants of PLD2, but not PLD1, inhibited significantly PDGF- and AngII-dependent PLD activity. We conclude: 1) the activation of PLD by cell surface receptors occurs primarily by an ARF-dependent mechanism in A10 cells, whereas the activation of PLD by protein kinase C-dependent pathways is only partially dependent on the regulation of ARF proteins; and 2) cell surface receptors, such as AngII and PDGF, signal primarily via PLD2 in A10 cells.
authors.affiliation: Department of Pharmacology of the University of Pittsburgh School of Medicine, Pennsylvania 15261, USA.
authors.forename: K
authors.initials: K
authors.lastname: Shome
authors.forename: M A
authors.initials: MA
authors.lastname: Rizzo
authors.forename: C
authors.initials: C
authors.lastname: Vasudevan
authors.forename: B
authors.initials: B
authors.lastname: Andresen
authors.forename: G
authors.initials: G
authors.lastname: Romero
mesh_terms: D036002:ADP Ribose Transferases / Q000494:pharmacology*; D020727:ADP-Ribosylation Factors / Q000235:genetics / Q000502:physiology*; D000804:Angiotensin II / Q000494:pharmacology*; D001905:Botulinum Toxins; D020126:Brefeldin A / Q000494:pharmacology; D002460:Cell Line; D003014:Clostridium botulinum / Q000201:enzymology; D019332:Endothelin-1 / Q000494:pharmacology*; D004789:Enzyme Activation / Q000187:drug effects; D015870:Gene Expression; D049452:Green Fluorescent Proteins; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics; D020559:Monomeric GTP-Binding Proteins / Q000502:physiology*; D009131:Muscle, Smooth, Vascular / Q000201:enzymology*; D009154:Mutation; D010739:Phospholipase D / Q000378:metabolism*; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014162:Transfection
chemical_list: D019332:Endothelin-1; D008164:Luminescent Proteins; D010982:Platelet-Derived Growth Factor; D011993:Recombinant Fusion Proteins; D000804:Angiotensin II; D049452:Green Fluorescent Proteins; D020126:Brefeldin A; D036002:ADP Ribose Transferases; C063007:exoenzyme C3, Clostridium botulinum; C105606:phospholipase D2; D010739:Phospholipase D; D001905:Botulinum Toxins; D020727:ADP-Ribosylation Factors; D020559:Monomeric GTP-Binding Proteins