cAMP does not inhibit convulxin-induced tyrosyl-phosphorylation of human platelet proteins, including PLCgamma2, but completely blocks the integrin alphaIIb beta3-dependent dephosphorylation step: comparisons with RGDS peptide, cytochalasin D, and phenylarsine oxide.
Convulxin (Cvx) isolated from Crotalus durissus terrificus venom, induces platelet aggregation, phospholipase C (PLC) activation, and tyrosyl-phosphorylation (PTP) of multiple proteins, including PLCgamma2 by a mechanism independent of integrin alphaIIb beta3. However, PTP induced by Cvx is followed by a dephosphorylation step in a platelet aggregation-dependent manner. Here we show that increasing intraplatelet content of cAMP with forskolin is associated with the inhibition of Cvx-induced platelet aggregation, ATP secretion, and inositol-phosphates production. However, the early onset of Cvx-induced PTP is not sensitive to cAMP (including PLCgamma2), and it also occurs in the presence of integrin alphaIIb beta3-antagonist (RGDS peptide, RGDS) or inhibitors of actin polymerization (cytochalasin D, CD) and tyrosine-phosphatases (phenylarsine oxide, PAO). However, forskolin, RGDS, and CD prevented the dephosphorylation step together with inhibition of platelet aggregation, whereas in the presence of phenylarsine oxide (PAO) the dephosphorylation step was replaced by an increase in the number and intensity of tyrosyl-phosphorylated proteins. Our data provide evidence to conclude that (i) cAMP inhibits platelet aggregation at a downstream site to PLCgamma2 tyrosyl-phosphorylation; (ii) Cvx-induced PTP is independent on integrin alphaIIb beta3 engagement, actin polymerization, and tyrosine-phosphatases activation; (iii) integrin alphaIIb beta3 mediates the dephosphorylation step in a platelet aggregation-dependent manner; and (iv) Cvx and collagen stimulate platelets by a similar signal transduction pathway.
Francischetti
I M
IM
ICB/CCS, Cidade Universitária-Ilha do Fundão, Rio de Janeiro, CEP 21941-590, Brazil.
Carlini
C R
CR
Guimarães
J A
JA
D001152:Arsenicals / Q000494:pharmacology; D001798:Blood Proteins / Q000187:drug effects / Q000378:metabolism*; D005576:Colforsin / Q000494:pharmacology; D003435:Crotalid Venoms / Q000494:pharmacology*; D000242:Cyclic AMP / Q000494:pharmacology*; D015638:Cytochalasin D / Q000494:pharmacology; D004790:Enzyme Induction; D006801:Humans; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000187:drug effects / Q000378:metabolism*; D037181:Lectins, C-Type; D009842:Oligopeptides / Q000494:pharmacology; D051966:Phospholipase C gamma; D010766:Phosphorylation; D015539:Platelet Activation / Q000187:drug effects*; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000037:antagonists & inhibitors* / Q000378:metabolism; D010738:Type C Phospholipases / Q000187:drug effects / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism
D001152:Arsenicals; D001798:Blood Proteins; D003435:Crotalid Venoms; D007527:Isoenzymes; D037181:Lectins, C-Type; D009842:Oligopeptides; D010975:Platelet Aggregation Inhibitors; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; C029341:oxophenylarsine; D005576:Colforsin; D015638:Cytochalasin D; C028890:convulxin; D014443:Tyrosine; C047981:arginyl-glycyl-aspartic acid; D000242:Cyclic AMP; D010738:Type C Phospholipases; D051966:Phospholipase C gamma