G_p-regulated phosphoinositide hydrolysis in turkey and human erythrocytes exposed to fluoride ion: Relationship to calcium influx

Previous studies have demonstrated that although both mammalian and avion erythrocytes express an inducibie inositol bisphosphate-specific phospholipase C, only the latter possess the guanine nucleotide-binding protein (G_p) that regulates this activity. In confirmation of previous reports, turkey erythrocyte plasma membranes responded to guanosine 5′-0-(3-thio)triphosphcrte (GTP-γ-S) and fluoroaluminates with hydrolysis of phosphoinositides, release of inositol phosphates, and generation of diacylglycerol, whereas human erythrocyte plasma membranes exhibited no such changes when incubated with known activators of guanine nucleotide regulatory proteins. We next contrasted responses of intact turkey and human erythrocytes to fluoroaluminates to develop a model to investigate the cellular effects of G_p activation. When turkey erythrocytes were exposed to fluoroaluminates, cellular levels of diacylglycerol and phosphatidic acid rapidly increased as phosphoinosirides were hydrolyzed. The alterations in the lipid composition of turkey erythrocytes effected by fluoroaluminates were remarkable; phosphatidic acid levels increased over 30-fold, whereas levels of polyphosphoinositides were decreased to less than 10% of those present before stimulation. In contrast, fluoroaluminates caused only minor alterations in the diacylglycerol and phospholipid content of intact human erythrocytes. To define the role of inositol-specific phospholipase C activation in the transmembrane conveyance of extracellular Ca⁺⁺, we compared the influx of extracellular Ca⁺⁺ in human and turkey erythrocytes exposed to fluoroaluminates. Fluoroaluminates initiated a sustained influx of extracellular ⁴⁵Ca⁺⁺ into turkey, but not human, erythrocytes. These results provide strong support for the hypothesis that G_p activation results in an influx of calcium into stimulated cells. Moreover, the data demonstrate that comparison of responses of human and turkey erythrocytes to fluoroaluminates provides a well-defined method for investigating the mechanisms and consequences of G_p activation in intact cells.