title: Selective activation of nuclear phospholipase D-1 by g protein-coupled receptor agonists in vascular smooth muscle cells.
abstract: Recent studies highlight the existence of an autonomous nuclear lipid metabolism related to cellular proliferation. However, the importance of nuclear phosphatidylcholine (PC) metabolism is poorly understood. Therefore, we were interested in nuclear PCs as a source of second messengers and, particularly, nuclear phospholipase D (PLD) identification in membrane-free nuclei isolated from pig aorta vascular smooth muscle cells (VSMCs). Using immunoblot experiment, in vitro PLD assay with fluorescent substrate and confocal microscopy analysis, we demonstrated that only PLD1 is expressed in VSMC nucleus, whereas PLD1 and PLD2 are present in VSMC. Inhibition of RhoA and protein kinase Czeta (PKCzeta) by C3-exoenzyme and PKCzeta pseudosubstrate inhibitor, respectively, conducted a decrease of phosphatidylethanol production. On the other hand, treatment of intact VSMCs, but not nuclei, with phosphoinositide 3-kinase (PI3K) inhibitors prevented partially nuclear PLD1 activity, indicating for the first time that PI3K may have a role in nuclear PLD regulation. In addition, lysophosphatidic acid (LPA) or angiotensin II treatment of VSMCs resulted in an increase of intranuclear PLD activity, whereas platelet-derived growth factor and epidermal growth factor have no significant effect. Moreover, pertussis toxin induced a decrease of LPA-stimulated nuclear PLD1 activity, suggesting that heterotrimeric G(i)/G(0) protein involvement in intranuclear PLD1 regulation. We also show that LPA-induced nuclear PLD1 activation implied PI3K/PKCzeta pathway activation and PKCzeta nuclear translocation as well as nuclear RhoA activation. Thus, the characterization of an endogenous PLD1 that could regulate PC metabolism inside VSMC nucleus provides a new role for this enzyme in control of vascular fibroproliferative disorders.
authors.affiliation: Département Lipoprotéines and Médiateurs Lipidiques, CPTP, INSERM Unité 563, Bâtiment C, CHU Purpan, BP 3028, 31024 Toulouse Cedex 3, France.
authors.forename: Stéphanie
authors.initials: S
authors.lastname: Gayral
authors.forename: Paul
authors.initials: P
authors.lastname: Déléris
authors.forename: Karine
authors.initials: K
authors.lastname: Laulagnier
authors.forename: Muriel
authors.initials: M
authors.lastname: Laffargue
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: Salles
authors.forename: Bertrand
authors.initials: B
authors.lastname: Perret
authors.forename: Michel
authors.initials: M
authors.lastname: Record
authors.forename: Monique
authors.initials: M
authors.lastname: Breton-Douillon
mesh_terms: D000804:Angiotensin II / Q000494:pharmacology; D000818:Animals; D001011:Aorta / Q000166:cytology; D002478:Cells, Cultured; D004789:Enzyme Activation; D006801:Humans; D008246:Lysophospholipids / Q000494:pharmacology; D009131:Muscle, Smooth, Vascular / Q000166:cytology*; D032389:Myocytes, Smooth Muscle / Q000201:enzymology* / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism; D037342:Pertussis Toxin / Q000494:pharmacology; D010713:Phosphatidylcholines / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases; D010739:Phospholipase D / Q000378:metabolism*; D043562:Receptors, G-Protein-Coupled / Q000819:agonists*; D013552:Swine
chemical_list: D008246:Lysophospholipids; D009687:Nuclear Proteins; D010713:Phosphatidylcholines; D043562:Receptors, G-Protein-Coupled; D000804:Angiotensin II; D037342:Pertussis Toxin; D019869:Phosphatidylinositol 3-Kinases; D010739:Phospholipase D; C106801:phospholipase D1; C032881:lysophosphatidic acid