Erythrocytic differentiation and glyceraldehyde-3-phosphate dehydrogenase expression are regulated by protein phosphorylation and cAMP in HD3 cells

Utilisation of glucose undergoes a marked decline during erythroblastic dif ferentiation in the chicken. Concomitantly there is a reduction in the expr ession of glucose transporter proteins and in the expression of the glycoly tic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAD). GAD activity dec lines, after an initial rise. while the level of GAD mRNA decreases rapidly after induction of differentiation, We have employed the temperature-sensi tive chicken erythroblast cell line HD3 that differentiates to the erythroc yte phenotype at 42 degrees C in the presence of inducers (hemin and butyri c acid). The role of tyrosine and serine/threonine phosphorylation pathways were evaluated with the phosphatase inhibitors sodium vanadate and okadaic acid, respectively. In the presence of phosphatase inhibitors, HD3 cells u nderwent differentiation and increased their synthesis of hemoglobin which is a marker protein for red blood cells differentiation. The levels of both GAD mRNA and enzymatic activity were increased by phosphatase inhibitors. The role of cAMP in differentiation was also assessed. Differentiation of H D3 cells was associated with an increase in cAMP. However the phosphodieste rase inhibitor IBMX was not a good inducer of hemoglobin synthesis but did induce GAD mRNA and enzymatic activity. Together these results suggest that multiple pathways (including serine/threonine phosphorylation, tyrosine ph osphorylation and elevated cAMP) are involved in the regulation of erythrob lastic differentiation, hemoglobin synthesis. GAD gene expression and GAD a ctivity in HD3 cells, (C) 2000 Elsevier Science Ltd. All rights reserved.