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A pathway for the synthesis of membrane phosphatidylcholine involving the N-methylation of phosphatidylethanolamine has been detected in several types of mammalian cells. Furthermore, it has been implicated in the coupling of agonist binding to cell response. We examined whether human platelets exhibit this synthetic pathway and whether platelet agonists influence its activity. When washed platelets were incubated with 0.15 µMl-[methyl-3H]methionine at 37°C, they incorporated methyl–3H into their phospholipids linearly at the rate of 1 pmole/109platelets/hr. When incubated with 20 µMradiolabeled methionine, they incorporated about 15 pmole/109platelets/hr. The radioactivity was found predominantly in phosphatidyl-N-monomethylethanolamine, phosphatidyl-N,N-dimethylethanolamine, and phosphatidylcholine. Thrombin caused an immediate (within 15 sec) and sustained (up to 30 min) decrease in the rate and extent of N-methylation of platelet phospholipids. This was accounted for by a decrease in synthesis of methylated phospholipids rather than an increase in their degradation. This thrombin effect correlated with serotonin release and could be dissociated from platelet aggregation and prostaglandin synthesis. Thrombin also decreased the synthesis of phosphatidylcholine when choline was used as the radiolabeled substrate. Other agonists such as epinephrine, adenosine diphosphate (ADP), or A23187 also decreased phospholipid methylation under conditions in which they stimulated serotonin release. These data demonstrate that platelets are capable of synthesizing phosphatidylcholine from phosphatidylethanolamine by N-methylation and that agonists perturb this pathway as they induce platelet secretion. The precise role of phospholipid methylation in either resting or stimulated platelets remains to be established.

The platelet membrane glycoproteins lIb (Gpllb) and GpIIIa form calcium-dependent heterodimers containing binding sites for fibrinogen, von Willebrand factor, and fibronectin. Although Gpllb and GplIIa are distinct proteins, both Gpllb and Gpllla are deficient in platelets from individuals with the recessive disorder Glanzmann’s thrombasthenia. To gain a better understanding of the genetic basis for Gpllb and Gpllla synthesis, we studied their synthesis by two human leukemia cell lines, HEL and K562. HEL cells contained complexes of Gpllb and Gpllla, and K562 cells expressed Gpllla, but not Gpllb, when stimulated with phorbol-12-myristate-13-acetate (PMA). RNA from HEL cells directed the in vitro synthesis of a 110,000-Mrprecursor for Gpllb and a 92,000-Mrprecursor for Gpllla, which indicates that the synthesis of Gpllb and Gpllla by HEL cells is directed by separate mRNAs. In contrast, RNA from PMA-stimulated K562 cells only directed the synthesis of a 92,000-Mrprecursor for Gpllla. The dissociation of Gpllb and Gpllla synthesis in K562 cells suggests that Gpllb and Gpllla may be the products of separate genes.