Activation of betagamma subunits of G(i2) and G(i3) proteins by basic secretagogues induces exocytosis through phospholipase Cbeta and arachidonate release through phospholipase Cgamma in mast cells.
Mast cells are activated by Ag-induced clustering of IgE bound to FcepsilonRI receptors or by basic secretagogues that stimulate pertussis toxin-sensitive heterotrimeric G proteins. The cell response includes the secretion of stored molecules, such as histamine, through exocytosis and of de novo synthesized mediators, such as arachidonate metabolites. The respective roles of G proteins alpha and betagamma subunits as well as various types of phospholipase C (PLC) in the signaling pathways elicited by basic secretagogues remain unknown. We show that a specific Ab produced against the C-terminus of Galpha(i3) and an anti-recombinant Galpha(i2) Ab inhibited, with additive effects, both exocytosis and arachidonate release from permeabilized rat peritoneal mast cells elicited by the basic secretagogues mastoparan and spermine. A specific Ab directed against Gbetagamma dimers prevented both secretions. Anti-PLCbeta Abs selectively prevented exocytosis. The selective phosphatidylinositol 3-kinase inhibitor LY 294002 prevented arachidonate release without modifying exocytosis. Gbetagamma coimmunoprecipitated with PLCbeta and phosphatidylinositol 3-kinase. The anti-PLCgamma1 and anti-phospholipase A(2) Abs selectively blocked arachidonate release. Protein tyrosine phosphorylation was inhibited by anti-Gbetagamma Abs, LY294002, and anti PLCgamma1 Abs. These data show that the early step of basic secretagogue transduction is common to both signaling pathways, involving betagamma subunits of G(i2) and G(i3) proteins. Activated Gbetagamma interacts, on one hand, with PLCbeta to elicit exocytosis and, on the other hand, with phosphatidylinositol 3-kinase to initiate the sequential activation of PLCgamma1, tyrosine kinases, and phospholipase A(2), leading to arachidonate release.
Ferry
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X
Laboratoire de Neuroimmunopharmacologie, Institut National de la Santé et de la Recherche Médicale, Unité 425, Université Louis Pasteur-Strasbourg I, Faculté de Pharmacie, Illkirch, France.
Eichwald
V
V
Daeffler
L
L
Landry
Y
Y
D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism*; D005089:Exocytosis; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000502:physiology*; D007527:Isoenzymes / Q000502:physiology*; D008297:Male; D008407:Mast Cells / Q000378:metabolism*; D054799:Phospholipase C beta; D051966:Phospholipase C gamma; D010741:Phospholipases A / Q000502:physiology; D021122:Protein Subunits; D011505:Protein-Tyrosine Kinases / Q000502:physiology; D051381:Rats; D017208:Rats, Wistar; D010738:Type C Phospholipases / Q000502:physiology*
D007527:Isoenzymes; D021122:Protein Subunits; D016718:Arachidonic Acid; D011505:Protein-Tyrosine Kinases; D010741:Phospholipases A; D010738:Type C Phospholipases; D054799:Phospholipase C beta; D051966:Phospholipase C gamma; D019206:GTP-Binding Protein alpha Subunits, Gi-Go