Cell signalling pathway involved in PACAP-induced AR4-2J cell proliferation.
The novel 38-amino acid neuropeptide PACAP (pituitary adenylate activating peptide) has recently been shown to induce the pancreatic acinar tumour AR4-2J cell growth. This growth promoting effect of PACAP was, however, independent of adenylate cyclase activation but suppressed by pertussis toxin and the somatostatin analog SMS 201-995. This study was undertaken to search for potential cell signalling pathways involved in the growth promoting effect of PACAP on AR4-2J cells. The AR4-2J cells were grown in Dulbecco's Modified Eagle's Medium containing 10% foetal calf serum. For studies on cell signalling pathways, all experiments were carried out on cells which have reached 50 to 75% confluency. At that point, they were transferred to serum free medium overnight with or without 1 microCi/ml myristic acid. The next morning, cells were harvested, washed and used for tyrosine kinase and phospholipase D (PLD) activities. For studies on growth, cells were grown for 2 days in the presence of 1 nM PACAP +/- the different inhibitors of tyrosine kinase and PLD. PACAP-38 and -27 caused a dose-dependent and parallel activation of tyrosine kinase and PLD an effect prevented by the antagonist PACAP 7-38. PACAP-38-stimulated tyrosine kinase and PLD activation are both dose-dependently inhibited by SMS 201-995. Finally, PACAP-stimulated tyrosine kinase and PLD activities are both inhibited by cell's preincubation with genistein and pertussis toxin. After 2 days, the PACAP-induced increase in AR4-2J cell growth was significantly inhibited by increasing concentrations of genistein and wortmannin, inhibitors of tyrosine kinase, PLD and phosphatidylinositol 3-kinase, respectively. PACAP can induce concomitant activation of tyrosine kinase and PLD; this finding and the observation that inhibition of these two enzymes inhibited PACAP-induced AR4-2J cell growth strongly suggests that they are intimately involved in the overall process of PACAP-induced AR4-2J cell proliferation.
Morisset
J
J
Départment de Biologie, Faculté des Sciences, Université de Sherbrooke, Québec, Canada.
Douziech
N
N
Rydzewska
G
G
Buscail
L
L
Rivard
N
N
D000730:Androstadienes / Q000494:pharmacology; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D002108:Ceruletide / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019833:Genistein; D006801:Humans; D007529:Isoflavones / Q000494:pharmacology; D008954:Models, Biological; D009479:Neuropeptides / Q000494:pharmacology*; D018377:Neurotransmitter Agents / Q000494:pharmacology; D010190:Pancreatic Neoplasms; D010446:Peptide Fragments / Q000494:pharmacology; D010739:Phospholipase D / Q000378:metabolism*; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D014407:Tumor Cells, Cultured; D000077191:Wortmannin
C495401:ADCYAP1 protein, human; D000730:Androstadienes; D004791:Enzyme Inhibitors; D007529:Isoflavones; D009479:Neuropeptides; D018377:Neurotransmitter Agents; D010446:Peptide Fragments; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide; D002108:Ceruletide; D019833:Genistein; D011505:Protein-Tyrosine Kinases; D010739:Phospholipase D; D000077191:Wortmannin