Epinephrine suppresses rap1B.GAP-activated GTPase activity in human platelets.
Lysate from quiescent platelets promotes rapid hydrolysis of [gamma-32P]GTP bound to rap1B. Various platelet agonists, including platelet-activating factor, phorbol 12,13-dibutyrate, alpha-thrombin, epinephrine, ADP, and iloprost, that affect platelet metabolism by different signal transduction pathways were used to stimulate intact platelets and study their effects on rap1B.GAP-activated GTPase activity (GAP, GTPase-activating protein). Only epinephrine was found to dramatically decrease not only the rate but also the amount of hydrolysis of rap1B-bound GTP activated by rap1B.GAP. This effect was dose dependent and occurred rapidly. The suppression of GTPase activity was specific for rap1B.GAP in that ras.GAP- and rap2B.GAP-activated GTPase activity were not affected by epinephrine stimulation. This effect appears to be mediated by the alpha 2-adrenergic receptor, as evidenced by a similar suppression of GTPase activity by stimulating platelets with the synthetic alpha 2-adrenergic receptor agonist UK14304 (bromoxidine). Furthermore, the selective alpha 2-adrenergic receptor antagonist yohimbine blocked the suppression of GTPase activity expressed in epinephrine-stimulated cell lysates. No apparent changes in the patterns of protein expression or tyrosine phosphorylation were observed. Although the migration characteristics upon anion-exchange chromatography of rap1B.GAP and ras.GAP activities were unaffected by epinephrine stimulation, the specific activity of rap1B.GAP was noticeably decreased with 250 and 500 microM epinephrine. These results suggest a possible role for rap1B and rap1B.GAP in epinephrine-stimulated signal transduction.
Marti
K B
KB
Division of Cell Biology, Burroughs Wellcome Co., Research Triangle Park, NC 27709.
Lapetina
E G
EG
D001792:Blood Platelets / Q000201:enzymology*; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D004837:Epinephrine / Q000494:pharmacology*; D020558:GTP Phosphohydrolases / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000378:metabolism / Q000502:physiology*; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D016283:Proto-Oncogene Proteins p21(ras) / Q000378:metabolism; D011942:Receptors, Adrenergic, alpha / Q000502:physiology*; D020668:rap GTP-Binding Proteins
D011942:Receptors, Adrenergic, alpha; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins; C065562:RAP2B protein, human; C501009:HRAS protein, human; D016283:Proto-Oncogene Proteins p21(ras); D020668:rap GTP-Binding Proteins; D004837:Epinephrine