Association of Tyr phosphorylation of GTPase-activating protein with mitogenic action of serotonin.
We have previously shown that serotonin (5-HT) induces both hyperplasia and hypertrophy of pulmonary artery smooth muscle cells (SMC) but not of endothelial cells (EC) through its high-affinity uptake. The present studies demonstrate rapid enhancement by 5-HT of Tyr phosphorylation of proteins, including p120, which also occurs in SMC but not in EC. The p120 protein was identified as GTPase-activating protein (GAP) by immunoprecipitation. Its phosphorylation occurred within minutes and preceded other events associated with 5-HT-induced mitogenesis. Tyr kinase (TK) and 5-HT uptake inhibitors and 8-bromoadenosine 3',5'-cyclic monophosphate blocked both the 5-HT-induced DNA synthesis and Tyr phosphorylation of GAP. Vanadate elevated DNA synthesis and Tyr phosphorylation of GAP of both control and 5-HT-treated cells. 5-HT failed to alter Tyr phosphorylation of GAP in cellular homogenates, as opposed to intact cells. In the presence of 3-isobutyl-1-methylxanthine, 5-HT inhibited cellular growth, presumably through its action on 5-HT1A or 5-HT4 receptors and elevation of adenosine 3',5'-cyclic monophosphate, but this was not associated with an alteration of Tyr phosphorylation of GAP. Similarly, a 5-HT1 or 5-HT2 receptor agonist failed to stimulate Tyr phosphorylation or DNA synthesis of SMC. Stimulation of cellular proliferation and enlargement produced by 1 microM 5-HT were totally abolished by TK inhibitors that did not affect 5-HT uptake. These data indicate that Tyr phosphorylation of GAP may act as an intermediate signal in 5-HT-induced mitogenesis of SMC which requires cellular internalization of 5-HT rather than its action on a membrane receptor.
Lee
S L
SL
Department of Medicine, New England Medical Center/Tufts University School of Medicine, Boston, Massachusetts 02111, USA.
Wang
W W
WW
Fanburg
B L
BL
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology; D000818:Animals; D002417:Cattle; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D004247:DNA / Q000037:antagonists & inhibitors / Q000096:biosynthesis; D004791:Enzyme Inhibitors / Q000494:pharmacology; D020690:GTPase-Activating Proteins; D008934:Mitogens / Q000494:pharmacology*; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011233:Precipitin Tests; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000494:pharmacology; D011506:Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012701:Serotonin / Q000494:pharmacology*; D017366:Serotonin Receptor Agonists / Q000494:pharmacology; D017367:Selective Serotonin Reuptake Inhibitors / Q000494:pharmacology; D014443:Tyrosine / Q000378:metabolism*; D014638:Vanadates / Q000494:pharmacology
D015056:1-Methyl-3-isobutylxanthine; D015124:8-Bromo Cyclic Adenosine Monophosphate; D004247:DNA; D004791:Enzyme Inhibitors; D020690:GTPase-Activating Proteins; D008934:Mitogens; D011505:Protein-Tyrosine Kinases; D011506:Proteins; D012701:Serotonin; D017366:Serotonin Receptor Agonists; D017367:Selective Serotonin Reuptake Inhibitors; D014443:Tyrosine; D014638:Vanadates