title: Differential phospholipase D activation by bradykinin and sphingosine 1-phosphate in NIH 3T3 fibroblasts overexpressing gelsolin.
abstract: Gelsolin, an actin-binding protein, shows a strong ability to bind to phosphatidylinositol 4,5-bisphosphate (PIP(2)). Here we showed in in vitro experiments that gelsolin inhibited recombinant phospholipase D1 (PLD1) and PLD2 activities but not the oleate-dependent PLD and that this inhibition was not reversed by increasing PIP(2) concentration. To investigate the role of gelsolin in agonist-mediated PLD activation, we used NIH 3T3 fibroblasts stably transfected with the cDNA for human cytosolic gelsolin. Gelsolin overexpression suppressed bradykinin-induced activation of phospholipase C (PLC) and PLD. On the other hand, sphingosine 1-phosphate (S1P)-induced PLD activation could not be modified by gelsolin overexpression, whereas PLC activation was suppressed. PLD activation by phorbol myristate acetate or Ca(2+) ionophore A23187 was not affected by gelsolin overexpression. Stimulation of control cells with either bradykinin or S1P caused translocation of protein kinase C (PKC) to the membranes. Translocation of PKC-alpha and PKC-beta1 but not PKC-epsilon was reduced in gelsolin-overexpressed cells, whereas phosphorylation of mitogen-activated protein kinase was not changed. S1P-induced PLC activation and mitogen-activated protein kinase phosphorylation were sensitive to pertussis toxin, but PLD response was insensitive to such treatment, suggesting that S1P induced PLD activation via certain G protein distinct from G(i) for PLC and mitogen-activated protein kinase pathway. Our results suggest that gelsolin modulates bradykinin-mediated PLD activation via suppression of PLC and PKC activities but did not affect S1P-mediated PLD activation.
authors.affiliation: Department of Biochemistry, Gifu University School of Medicine, Tsukasamachi-40, Gifu 500-8705, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Banno
authors.forename: H
authors.initials: H
authors.lastname: Fujita
authors.forename: Y
authors.initials: Y
authors.lastname: Ono
authors.forename: S
authors.initials: S
authors.lastname: Nakashima
authors.forename: Y
authors.initials: Y
authors.lastname: Ito
authors.forename: N
authors.initials: N
authors.lastname: Kuzumaki
authors.forename: Y
authors.initials: Y
authors.lastname: Nozawa
mesh_terms: D016475:3T3 Cells; D000818:Animals; D001920:Bradykinin / Q000494:pharmacology*; D000001:Calcimycin / Q000494:pharmacology; D002462:Cell Membrane / Q000201:enzymology; D018076:DNA, Complementary; D004533:Egtazic Acid / Q000494:pharmacology; D004789:Enzyme Activation; D018260:Gelsolin / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism; D007700:Kinetics; D008246:Lysophospholipids; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D037342:Pertussis Toxin; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000494:pharmacology; D010739:Phospholipase D / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism; D064546:Protein Kinase C beta; D051571:Protein Kinase C-alpha; D051744:Protein Kinase C-epsilon; D011994:Recombinant Proteins / Q000378:metabolism; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014162:Transfection; D010738:Type C Phospholipases / Q000378:metabolism*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
chemical_list: D018076:DNA, Complementary; D018260:Gelsolin; D007527:Isoenzymes; D008246:Lysophospholipids; D019269:Phosphatidylinositol 4,5-Diphosphate; D011994:Recombinant Proteins; D010566:Virulence Factors, Bordetella; C060506:sphingosine 1-phosphate; D000001:Calcimycin; D004533:Egtazic Acid; D037342:Pertussis Toxin; C497256:Prkce protein, mouse; C496628:PRKCA protein, human; C497255:PRKCE protein, human; C496630:Prkca protein, mouse; D011493:Protein Kinase C; D064546:Protein Kinase C beta; D051571:Protein Kinase C-alpha; D051744:Protein Kinase C-epsilon; D020928:Mitogen-Activated Protein Kinases; D010738:Type C Phospholipases; D010739:Phospholipase D; D013110:Sphingosine; D013755:Tetradecanoylphorbol Acetate; D001920:Bradykinin