ADP-ribosylation factor-dependent phospholipase D activation by the M3 muscarinic receptor.
G protein-coupled receptors can potentially activate phospholipase D (PLD) by a number of routes. We show here that the native M3 muscarinic receptor in 1321N1 cells and an epitope-tagged M3 receptor expressed in COS7 cells substantially utilize an ADP-ribosylation factor (ARF)-dependent route of PLD activation. This pathway is activated at the plasma membrane but appears to be largely independent of G, phospholipase C, Ca2+ q/11, protein kinase C, tyrosine kinases, and phosphatidyl inositol 3-kinase. We report instead that it involves physical association of ARF with the M3 receptor as demonstrated by co-immunoprecipitation and by in vitro interaction with a glutathione S-transferase fusion protein of the receptor's third intracellular loop domain. Experiments with mutant constructs of ARF1/6 and PLD1/2 indicate that the M3 receptor displays a major ARF1-dependent route of PLD1 activation with an additional ARF6-dependent pathway to PLD1 or PLD2. Examples of other G protein-coupled receptors assessed in comparison display alternative pathways of protein kinase C- or ARF6-dependent activation of PLD2.
Pubmed ID: 12799371 RIS Download
ADP-Ribosylation Factor 1 | ADP-Ribosylation Factors | Animals | Biotinylation | Blotting, Western | Brefeldin A | COS Cells | Carbachol | Cell Line | Cell Membrane | Dose-Response Relationship, Drug | Enzyme Activation | Enzyme Inhibitors | Epitopes | Estrenes | Glutathione Transferase | Humans | Immunoblotting | Inhibitory Concentration 50 | Ligands | Models, Biological | Mutation | Phospholipase D | Precipitin Tests | Protein Binding | Protein Kinase C | Protein Structure, Tertiary | Protein Transport | Pyrrolidinones | Receptor, Muscarinic M3 | Receptors, Muscarinic | Signal Transduction | Subcellular Fractions | Time Factors | Transfection | Tumor Cells, Cultured