Erythropoietin stimulates tyrosine phosphorylation and taurine transport in skate erythrocytes.
Taurine, a beta amino acid, is a primary osmolyte in nucleated skate erythrocytes and is involved in the regulation of cell volume. Growth factors may be involved in the regulation of cell volume which occurs during cell division. Erythropoietin (EPO) is the primary growth factor controlling erythropoiesis. To investigate its mechanism of action, we used nucleated skate erythrocytes. EPO stimulates Na(+)-independent uptake of taurine in a concentration-dependent manner. The uptake was inhibited by the tyrosine kinase inhibitor genistein. Concomitantly, EPO stimulates tyrosine phosphorylation of a number of proteins, particularly ones of molecular masses 145, 120, 100, 80, 65, and 35 kDa. Using specific antibodies, the 145 kDa protein is identified as phospholipase C gamma-1 (PLC gamma-1) and the 100 kDa protein as the skate homolog of the anion exchanger band 3. Since PLC gamma-1 is activated, turnover of membrane lipids was determined. EPO increased 1,2-diacylglycerol formation from phosphatidylinositols (phosphatidylinositol-4-monophosphate and 4,5-biphosphate) during an early phase and later preferentially from phosphatidylcholine. The early hydrolysis of phosphoinositides 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 transphosphatidylation, appeared at times longer than 5 min, suggesting delayed activation of PLD. These results demonstrate that EPO, via simulation of tyrosine phosphorylation, stimulates taurine transport in skate erythrocytes.
Musch
M W
MW
Department of Medicine, University of Chicago, Illinois 60637, USA.
Davis-Amaral
E M
EM
Goldstein
L
L
D000818:Animals; D001457:Anion Exchange Protein 1, Erythrocyte / Q000378:metabolism; D001692:Biological Transport; D004789:Enzyme Activation; D004912:Erythrocytes / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D004921:Erythropoietin / Q000494:pharmacology*; D019833:Genistein; D006868:Hydrolysis; D007527:Isoenzymes / Q000378:metabolism; D007529:Isoflavones / Q000494:pharmacology; D010716:Phosphatidylinositols / Q000378:metabolism; D051966:Phospholipase C gamma; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D012860:Skates, Fish / Q000097:blood; D013654:Taurine / Q000097:blood*; D010738:Type C Phospholipases / Q000378:metabolism; D014443:Tyrosine / Q000097:blood*
D001457:Anion Exchange Protein 1, Erythrocyte; D007527:Isoenzymes; D007529:Isoflavones; D010716:Phosphatidylinositols; D004921:Erythropoietin; D013654:Taurine; D014443:Tyrosine; D019833:Genistein; D011505:Protein-Tyrosine Kinases; D010738:Type C Phospholipases; D051966:Phospholipase C gamma