Modulation of IRP Binding Activity by Oxygen in Primary Erythroid Cells.

Abstract

Iron regulatory proteins 1 and 2 (IRP1 and IRP2) are cytoplasmic RNA-binding proteins that target specific stem-loop RNA structures known as iron responsive elements (IRE). Binding of IRPs to IREs inhibits translation of ferritin mRNA and stabilizes transferrin receptor (TfR) mRNA. Various factors have been reported to regulate binding activity of IRPs, such as iron, phosphorylation, nitric oxide and hypoxia. While there is a consistent agreement on the negative effect of iron on the interaction between IRPs and IREs, reports regarding the influence of hypoxia on the IRE-binding activity of IRPs vary in a species and cell specific manner. It was the aim of this work to study the effect of hypoxic (3% oxygen) and normoxic (20% oxygen) conditions on IRP binding activity in primary erythroid cells. The cells were induced for differentiation and incubated under physiological, low (Desferrioxamine) and high (ferric ammonium citrate) iron conditions. Binding activity of IRPs and protein levels of ferritin and TfR as well as cell proliferation and differentiation parameters were determined to analyze the regulation of iron metabolism during terminal differentiation. The data show, that in developing red blood cells binding activities of IRP1 and IRP2 are reduced at 3% oxygen. This reduction correlates with increased ferritin protein levels and decreased TfR protein levels. Moreover, incubation under hypoxia strongly decreased cell expansion and reduces hemoglobinization. These results suggest that terminal erythroid differentiation in the bone marrow might occur under normoxic rather than hypoxic conditions.