PHOSPHORYLATION OF MEMBRANE-PROTEINS IN ERYTHROCYTES TREATED WITH LEAD

In immature rat microvessels, endothelial cells and glioma cells, expo sure to lead results in an increase in the level of protein kinase C i n membranes. In this paper we have extended these studies to human ery throcytes and, in addition, studied the phosphorylation of membrane pr oteins. A significant increase in the phosphorylation of membrane cyto skeletal proteins of molecular mass 120, 80, 52 and 45 kDa was observe d in human erythrocytes treated for 60 min with lead acetate at concen trations greater than 100 nM. These same proteins were phosphorylated when erythrocytes were treated for 10 min with 50 nM phorbol 12-myrist ate 13-acetate (PMA). Similarly, protein kinase C activity was elevate d and an increase in the amount of protein kinase C-alpha was observed in membranes from erythrocytes exposed to concentrations of lead acet ate above 100 nM. No changes, however, in the activities of cAMP-depen dent protein kinase, protein phosphatases I and IIA or casein kinase w ere observed. Phosphorylation of the membrane proteins stimulated by l ead acetate or by PMA was not observed in erythrocytes depleted of pro tein kinase C by a 72-h treatment with 500 nM phorbol 12,13-dibutyrate . Finally, no changes in the levels of calcium or diacylglycerol were observed in erythrocytes stimulated with 100 nM lead acetate. These re sults indicate that, in erythrocytes, lead acetate stimulates the phos phorylation of membrane cytoskeletal proteins by a mechanism dependent on protein kinase C. Since levels of calcium or diacylglycerols did n ot increase, it appears that lead may activate the enzyme by a direct interaction.