title: ADP-ribosylation factor proteins mediate agonist-induced activation of phospholipase D.
abstract: The role of small G proteins of the ADP-ribosylation factor (ARF) and Rho families on the activation of phospholipase D (PLD) by platelet-derived growth factor (PDGF) and phorbol esters (PMA) has been investigated. The activation of PLD by PDGF and PMA was blocked by brefeldin A (BFA), an inhibitor of ARF activation, but not by Clostridium botulinum C3 exotoxin, an inhibitor of the activity of Rho. PDGF and PMA, in the presence of GTPgammaS, promoted the association of ARF and RhoA with cell membranes. Cells depleted of ARF and Rho by digitonin permeabilization showed a significant reduction of the activity of phospholipase D. Recombinant ARF was sufficient to restore agonist-induced PLD activity to digitonin-permeabilized, cytoplasm-depleted cells. In contrast, isoprenylated recombinant RhoA had no effects in this reconstitution assay. HIRcB cells were transiently transfected with wild-type and dominant-negative mutants of ARF1 and ARF6. Neither wt-ARF1 nor wt-ARF6 had any effects on agonist-dependent PLD activity. However, dominant-negative ARF1 and ARF6 mutants blocked the stimulation of PLD by PDGF but only partially inhibited the effects of PMA. These results demonstrate that ARF rather than Rho proteins mediate the activation of PLD by PDGF and phorbol esters in HIRcB fibroblasts.
authors.affiliation: Department of Pharmacology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15261, USA.
authors.forename: K
authors.initials: K
authors.lastname: Shome
authors.forename: Y
authors.initials: Y
authors.lastname: Nie
authors.forename: G
authors.initials: G
authors.lastname: Romero
mesh_terms: D036002:ADP Ribose Transferases / Q000494:pharmacology; D020823:ADP-Ribosylation Factor 1; D020727:ADP-Ribosylation Factors; D000255:Adenosine Triphosphate / Q000378:metabolism; D001905:Botulinum Toxins; D020126:Brefeldin A / Q000494:pharmacology; D002478:Cells, Cultured; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism; D010739:Phospholipase D / Q000378:metabolism*; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D011493:Protein Kinase C / Q000378:metabolism; D051571:Protein Kinase C-alpha; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D020742:rhoA GTP-Binding Protein
chemical_list: D007527:Isoenzymes; D010982:Platelet-Derived Growth Factor; D011500:Protein Synthesis Inhibitors; D020126:Brefeldin A; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D000255:Adenosine Triphosphate; D036002:ADP Ribose Transferases; C063007:exoenzyme C3, Clostridium botulinum; C496628:PRKCA protein, human; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D010739:Phospholipase D; D001905:Botulinum Toxins; D019204:GTP-Binding Proteins; D020823:ADP-Ribosylation Factor 1; D020727:ADP-Ribosylation Factors; D020742:rhoA GTP-Binding Protein; D013755:Tetradecanoylphorbol Acetate