ERYTHROPOIETIN STIMULATES TYROSINE PHOSPHORYLATION AND TAURINE TRANSPORT IN SKATE ERYTHROCYTES

Taurine, a beta amino acid, is a primary osmolyte in nucleated skate e rythrocytes and is involved in the regulation of cell volume. Growth f actors may be involved in the regulation of cell volume which occurs d uring cell division. Erythropoietin (EPO) is the primary growth factor controlling erythropoiesis. To investigate its mechanism of action, w e used nucleated skate erythrocytes. EPO stimulates Na+-independent up take of taurine in a concentration-dependent manner. The uptake was in hibited by the tyrosine kinase inhibitor genistein. Concomitantly, EPO stimulates tyrosine phosphorylation of a number of proteins, particul arly ones of molecular masses 145, 120, 100, 80, 65, and 35 kDa. Using specific antibodies, the 145 kDa protein is identified as phospholipa se C gamma-1 (PLC gamma-1) and the 100 kDa protein as the skate homolo g of the anion exchanger band 3. Since PLC gamma-1 is activated, turno ver of membrane lipids was determined. EPO increased 1,2-diacylglycero l formation from phosphatidylinositols (phosphatidylinositol-4-monopho sphate and 4,5-biphosphate) during an early phase and later preferenti ally from phosphatidylcholine. The early hydrolysis of phosphoinositid es was confirmed measuring generation of inositol-1,4,5-trisphosphate, demonstrating an activation of PLC gamma-1 activity. To determine if phospholipase D (PLD) stimulation also occurred, ethanol was included in the reactions. Phosphatidylethanol, synthesized by PLD-mediated tra nsphosphatidylation, appeared at times longer than 5 min, suggesting d elayed activation of PLD. These results demonstrate that EPO, via simu lation of tyrosine phosphorylation, stimulates taurine transport in sk ate erythrocytes. (C) 1996 Wiley-Liss, Inc.