title: Ral isoforms are implicated in Fc gamma R-mediated phagocytosis: activation of phospholipase D by RalA.
abstract: Phagocytosis is an essential element of the immune response permitting the elimination of pathogens, cellular debris, apoptotic cells, and tumor cells. Recently, both phospholipase D (PLD) isoforms, PLD1 and PLD2, were shown to be necessary for efficient FcgammaR-mediated phagocytosis. In this study, we investigated the role of a potential PLD regulator, the Ral GTPases RalA and RalB, in murine RAW 264.7 macrophages. Both Ral isoforms are expressed in macrophages and are transiently activated following FcgammaR stimulation. When Ral expression levels were varied using Ral mutants or interference RNA, phagocytosis assays revealed that Ral isoforms have antagonistic effects; RalA is a positive modulator, whereas RalB plays a negative role. We then focused on RalA and its possible relationship with PLD. The increase in PLD activity that occurs when phagocytosis is stimulated was inhibited in cells with reduced RalA protein, but it was unaffected by reduced levels of RalB. Furthermore, in macrophages transfected with dsRed-RalA and GFP-PLD1 or GFP-PLD2, RalA colocalized with PLD1 and PLD2 at the phagocytic cup during phagosome formation. Additional results obtained from immunoprecipitation of PLD from macrophages transfected with myc-RalA and hemagglutinin-tagged PLD1 or PLD2 indicated an enhanced interaction of RalA with both PLD isoforms during phagocytic stimulation. The increase in RalA and PLD1 interaction was transient and correlated with the time course of RalA activation. These findings reveal a novel pathway involving RalA and PLD in the regulation of FcgammaR-mediated phagocytosis.
authors.affiliation: Département Neurotransmission et Sécrétion Neuroendocrine, Institut des Neurosciences Cellulaires et Intégratives, Centre National de la Recherche Scientifique, Unité Propre de Recherche 3212, Strasbourg, France.
authors.forename: Matthias
authors.initials: M
authors.lastname: Corrotte
authors.forename: An Phu Tran
authors.initials: AP
authors.lastname: Nyguyen
authors.forename: Marie Line
authors.initials: ML
authors.lastname: Harlay
authors.forename: Nicolas
authors.initials: N
authors.lastname: Vitale
authors.forename: Marie-France
authors.initials: MF
authors.lastname: Bader
authors.forename: Nancy J
authors.initials: NJ
authors.lastname: Grant
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D015536:Down-Regulation / Q000276:immunology; D008264:Macrophages / Q000201:enzymology / Q000276:immunology / Q000378:metabolism; D051379:Mice; D010587:Phagocytosis / Q000276:immunology*; D010588:Phagosomes / Q000201:enzymology / Q000276:immunology; D010712:Phosphatidic Acids / Q000502:physiology; D010739:Phospholipase D / Q000378:metabolism*; D020033:Protein Isoforms / Q000502:physiology; D017452:Receptors, IgG / Q000502:physiology*; D015398:Signal Transduction / Q000276:immunology; D015854:Up-Regulation / Q000276:immunology; D020725:ral GTP-Binding Proteins / Q000502:physiology*
chemical_list: C485701:Fcgr1 protein, mouse; D010712:Phosphatidic Acids; D020033:Protein Isoforms; D017452:Receptors, IgG; D010739:Phospholipase D; C118120:Rala protein, mouse; C535173:RalB protein, mouse; D020725:ral GTP-Binding Proteins