Regulation of phospholipase D by muscarinic receptors in rat submandibular ductal cells.
The muscarinic agonist carbachol stimulated phospholipase D (PLD) in rat submandibular gland (RSMG) ductal cells in a time and concentration-dependent manner. This effect was inhibited by chelation of extracellular calcium with ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA). PLD could also be activated by epinephrine and AlF(4)(-), two polyphosphoinositide-specific phospholipase C (PPI-PLC) activators, and by the phorbol ester o-tetradecanoylphorbol 13-acetate (TPA) which activates protein kinase C (PKC). Ionomycin and thapsigargin only slightly increased PLD activity. Ortho-vanadate, a tyrosine phosphatase inhibitor, also stimulated PLD activity. Both carbachol and o-vanadate increased the formation of inositol phosphates and the tyrosine phosphorylation of at least two proteins (55-60 and 120 kDa). Calphostin C (a PKC inhibitor), U73122 (a PPI-PLC inhibitor) and genistein (a tyrosine kinase inhibitor) blocked the activation of PLD, of PLC and the phosphorylation of tyrosyl residues in response to carbachol and vanadate. Taken together, these results suggest that rat submandibular gland ductal cells express a calcium-dependent PLD activity. This enzyme is regulated by carbachol via a PLC-PKC-tyrosine kinase pathway.
Pochet
Stéphanie
S
Laboratoire de Biochimie et de Biologie Cellulaire, Institut de Pharmacie CP 205/3, Campus Plaine, Université Libre de Bruxelles, Boulevard du Triomphe, B 1050 Brussels, Belgium. spochet@ulb.ac.be
Métioui
Mourad
M
Grosfils
Katrina
K
Gómez-Muñoz
Antonio
A
Marino
Aida
A
Dehaye
Jean-Paul
JP
D000818:Animals; D002118:Calcium / Q000502:physiology; D002217:Carbachol / Q000494:pharmacology; D002478:Cells, Cultured; D018679:Cholinergic Agonists / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007700:Kinetics; D008297:Male; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D010739:Phospholipase D / Q000378:metabolism*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000502:physiology; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000502:physiology; D051381:Rats; D017208:Rats, Wistar; D011976:Receptors, Muscarinic / Q000378:metabolism*; D015398:Signal Transduction; D013363:Submandibular Gland / Q000166:cytology* / Q000201:enzymology* / Q000378:metabolism; D010738:Type C Phospholipases / Q000502:physiology
D018679:Cholinergic Agonists; D004791:Enzyme Inhibitors; D011976:Receptors, Muscarinic; D002217:Carbachol; D011505:Protein-Tyrosine Kinases; D011493:Protein Kinase C; D010738:Type C Phospholipases; D010739:Phospholipase D; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D002118:Calcium