Upon platelet activation, numerous proteins are known to be tyrosine phosphorylated. To investigate the mechanisms of the regulation of tyrosine phosphorylation and its physiological significance, the effects on tyrosine phosphorylation of agents that elevate the platelet level of the cyclic nucleotides cAMP and cGMP were examined in aspirin-treated gel-filtered platelets by Western blotting with a specific antiphosphotyrosine antibody. The effects of these agents on other aspects of platelet activation, i.e., aggregation, secretion, and elevation of the concentration of cytosolic ionized calcium ([Ca2+](i)), were also examined in parallel experiments. Tyrosine phosphorylation in platelets activated by α-thrombin (1 nM) was inhibited by prostaglandin (PG) E1 (2 μM) or by sodium nitroprusside (100 μM). Elevation of [Ca2+](i), aggregation, and serotonin secretion was also strongly inhibited. On the other hand, a higher concentration of α-thrombin (10 nM) induced tyrosine phosphorylation of the same proteins, elevation of [Ca2+](i), platelet aggregation, and serotonin secretion, irrespective of pretreatment of platelets by either PGE1 or sodium nitroprusside. Inhibition by sodium nitroprusside of tyrosine phosphorylation induced by α-thrombin (1 nM) was accompanied by an increased concentration of cGMP. 8-BrcGMP (2 mM) also inhibited tyrosine phosphorylation and aggregation, although less than sodium nitroprusside. ADP (20 μM) induced platelet shape change and tyrosine phosphorylation of only a few proteins; these effects were also inhibited by either PGE1 or sodium nitroprusside. Thus tyrosine phosphorylation in platelets can be inhibited by elevation of either cAMP or cGMP, an effect that is overcome by a high concentration of thrombin, resulting in granule secretion and aggregation. Some of the proteins that are tyrosine phosphorylated may be important in the regulation of platelet functions.
- guanine nucleotide-binding protein
ASJC Scopus subject areas
- Cell Biology