AGONIST-INDUCED PHOSPHORYLATION OF HUMAN PLATELET TXA(2)/PGH(2) RECEPTORS

Thromboxane A(2) (TXA(2)) and its precursor prostaglandin H-2 (PGH(2)) stimulate platelet activation through interaction with TXA(2)/PGH(2) receptors. We and others have shown that these receptors undergo homologous desensitization upon prolonged exposure to thromboxane A, mimetics. Phosphorylation of receptors has previously been reported to be an important mechanism for receptor desensitization. In the present study we examined the possibility that homologous desensitization of human platelet TXA(2)/PGH(2) receptors may involve phosphorylation. The ATP pool of human platelets was metabolically prelabeled with P-32(i) and the labeled platelets were subsequently exposed for 1 and 10 min to the stable TXA(2)/PGH(2) mimetic, U46619 (1 mu M). TXA(2)/PGH(2) receptors were purified approx. 2000-fold by affinity and wheatgerm lectin chromatography and subjected to SDS-PAGE followed by autoradiography. A phosphorylated plasma membrane glycoprotein (M(r) = 50-57 kDa) was detected with characteristics similar to the TXA(2)/PGH(2) receptor. This glycoprotein was found to be phosphorylated in the unstimulated state but phosphorylation was increased by exposure to U46619. Phosphorylation occurred rapidly and was inhibited when platelets were preincubated with the TXA(2)/PGH(2) receptor antagonist, SQ29548 (5 mu M), before being stimulated with U46619. These results suggest that human platelet TXA(2)/PGH(2) receptors are phosphoproteins and that the level of phosphorylation is increased during homologous desensitization.