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 differentiation in the chicken. Concomitantly there is a reduction in the expression of glucose transporter proteins and in the expression of the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAD). GAD activity declines, after an initial rise, while the level of GAD mRNA decreases rapidly after induction of differentiation. We have employed the temperature-sensitive chicken erythroblast cell line HD3 that differentiates to the erythrocyte phenotype at 42 degrees C in the presence of inducers (hemin and butyric 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 underwent 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 HD3 cells was associated with an increase in cAMP. However the phosphodiesterase 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 phosphorylation and elevated cAMP) are involved in the regulation of erythroblastic differentiation, hemoglobin synthesis, GAD gene expression and GAD activity in HD3 cells.
Grdisa
M
M
Division of Molecular Medicine, "Rudjer Boskovic" Institute, Zagreb, Croatia. grdisa@rudjer.irb.hr
White
M K
MK
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D002645:Chickens; D000242:Cyclic AMP / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004912:Erythrocytes / Q000166:cytology / Q000201:enzymology* / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000235:genetics / Q000378:metabolism*; D019319:Okadaic Acid / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors; D010766:Phosphorylation; D012333:RNA, Messenger / Q000378:metabolism; D014638:Vanadates / Q000494:pharmacology
D004791:Enzyme Inhibitors; D012333:RNA, Messenger; D019319:Okadaic Acid; D014638:Vanadates; D000242:Cyclic AMP; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D010749:Phosphoprotein Phosphatases; D015056:1-Methyl-3-isobutylxanthine