Activation of phospholipase D-2 by P2X(7) agonists in rat submandibular gland acini.
Exogenous ATP stimulated phospholipase D (PLD), but not sphingomyelinase in rat submandibular gland (SMG) acini. PLD activation was dependent upon extracellular Ca(2+) and did not involve intracellular Ca(2+) mobilization or phosphoinositide-specific phospholipase C activation. ATP-stimulated PLD was attenuated by inhibition or downregulation of protein kinase C (PKC). PLD activation was fully blocked by the cytosolic phospholipase A(2) (PLA(2)) inhibitor ONO-RS-082 and partially attenuated by the selective Ca(2+)-dependent cytosolic PLA(2) inhibitor, arachidonyl trifluoromethylketone (AACOCF(3)), or by bromoenol lactone, an inhibitor of Ca(2+)-independent cytosolic PLA(2). Magnesium, which decreases the concentration of ATP(4-), and nickel, which blocks nonspecific cation channels coupled to purinergic receptors, inhibited PLD activation by ATP. Using reverse transcription-polymerase chain reaction and Northern blotting techniques, we demonstrated that the PLD isoform stimulated by ATP was PLD-2. Among various ATP analogs, only the P2Z/P2X(7) purinergic receptor agonist benzoyl-benzoyl ATP stimulated PLD-2. The response to ATP was inhibited by the nonselective P2X purinergic antagonist suramin and by oxidized ATP, a potent P2Z/P2X(7) receptor antagonist. It is concluded that in rat SMG acinar cells, PLD-2 is upregulated by exogenous ATP through a mechanism involving Ca(2+) influx, cytosolic PLA(2), and PKC. Also, the data suggest an involvement of P2X(7) receptors in PLD-2 stimulation by ATP.
Pérez-Andrés
Encarnación
E
Department of Biochemistry and Molecular Biology, Faculty of Medicine, University of the Basque Country, P.O. Box 644, 48080 Bilbao, Spain.
Fernández-Rodriguez
María
M
González
Mónica
M
Zubiaga
Ana
A
Vallejo
Ainara
A
García
Itxaso
I
Matute
Carlos
C
Pochet
Stéphanie
S
Dehaye
Jean Paul
JP
Trueba
Miguel
M
Marino
Aida
A
Gómez-Muñoz
Antonio
A
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D017931:DNA Primers; D004789:Enzyme Activation; D008297:Male; D051379:Mice; D010739:Phospholipase D / Q000378:metabolism*; D058910:Purinergic P2 Receptor Agonists; D051381:Rats; D017207:Rats, Sprague-Dawley; D058486:Receptors, Purinergic P2X7; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013363:Submandibular Gland / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology
D017931:DNA Primers; C546469:P2rx7 protein, mouse; C546470:P2rx7 protein, rat; D058910:Purinergic P2 Receptor Agonists; D058486:Receptors, Purinergic P2X7; D000255:Adenosine Triphosphate; C105606:phospholipase D2; D010739:Phospholipase D