Translation Initiation Control by Heme-Regulated Eukaryotic Initiation Factor 2α Kinase in Erythroid Cells under Cytoplasmic Stresses

Abstract

Cytoplasmic stresses, including heat shock, osmotic stress, and oxidative stress, cause rapid inhibition of protein synthesis in cells through phosphorylation of eukaryotic initiation factor 2␣ (eIF2␣) by eIF2␣ kinases. We have investigated the role of heme-regulated inhibitor (HRI), a heme-regulated eIF2␣ kinase, in stress responses of erythroid cells. We have demonstrated that HRI in reticulocytes and fetal liver nucleated erythroid progenitors is activated by oxidative stress induced by arsenite, heat shock, and osmotic stress but not by endoplasmic reticulum stress or nutrient starvation. While autophosphorylation is essential for the activation of HRI, the phosphorylation status of HRI activated by different stresses is different. The contributions of HRI in various stress responses were assessed with the aid of HRI-null reticulocytes and fetal liver erythroid cells. HRI is the only eIF2␣ kinase activated by arsenite in erythroid cells, since HRI-null cells do not induce eIF2␣ phosphorylation upon arsenite treatment. HRI is also the major eIF2␣ kinase responsible for the increased eIF2␣ phosphorylation upon heat shock in erythroid cells. Activation of HRI by these stresses is independent of heme and requires the presence of intact cells. Both hsp90 and hsc70 are necessary for all stress-induced HRI activation. However, reactive oxygen species are involved only in HRI activation by arsenite. Our results provide evidence for a novel function of HRI in stress responses other than heme deficiency.Protein synthesis in intact reticulocytes and their lysates is dependent on the availability of heme. In heme deficiency, protein synthesis is inhibited, with a marked decrease in the formation of 40S-eukaryotic initiation factor 2 (eIF2)-MettRNA f GTP (43S preinitiation complex) and with increased phosphorylation of the ␣ subunit of eIF2. eIF2 is an initiation factor which binds GTP and Met-tRNA f and is required for the formation of the 43S initiation complex. The phosphorylation of eIF2␣ in heme deficiency is the result of the activation of heme-regulated inhibitor (HRI), which is a heme-regulated eIF2␣ kinase (8, 9). The mechanism of inhibition of translation initiation by the phosphorylation of eIF2␣ has been studied extensively (18,19). In brief, the recycling of eIF2 in initiation requires the exchange of bound GDP for GTP. Under physiological conditions, eIF2 has a 400-fold-greater affinity for GDP than for GTP. The exchange of tightly bound GDP for GTP requires eIF2B, which is rate limiting and is present at 15 to 25% of the amount of eIF2. When eIF2␣ is phosphorylated, the binding of eIF2(␣P)-GDP to the regulatory subcomplex of eIF2B is much tighter than the binding of eIF2-GDP to eIF2B (25). This tighter interaction of phosphorylated eIF2 with eIF2B prevents the GDP-GTP exchange activity of eIF2B. In this manner, once the amount of phosphorylated eIF2 exceeds the amount of eIF2B, the shutoff of protein synthesis occurs.Phosphorylation of eIF2␣ was later found to be a common regulatory mechanism of protein tr...

miR-214 protects erythroid cells against oxidative stress by targeting ATF4 and EZH2

“…heme oxygenase-l (HO-1)) [72], the crosstalk between these two cytoprotective pathways remains less understood. Previous mechanistic studies demonstrated that ATF4 is predominantly activated through eIF2α phosphorylation by oxidative stress derived from unfolded proteins or As treatment, and its activation is necessary to antagonize cell death in erythroid cells upon these stresses [22,70]. In the current study, ATF4 was found to be activated by Nrf2 through repressing miR-214 expression.…”

“…Therefore, erythroid cells are highly sensitive to oxidative stress, and aberrant ROS accumulation incurs impaired erythropoiesis, shortened lifespan of RBCs and even hemolysis [66,69,70]. A comprehensive protective machinery is employed in erythroid cells during differentiation.…”

Rôle de l’heme regulated inhibitor(HRI) dans la résistance à l’apoptose

“…Dans la -thalassémie -due à une mutation qui bloque l'expression du gène codant la -globine -l'anémie est causée par une accumulation anormale d'-globine. Puisque HRI sous sa forme active inhibe la traduction protéique, sa déplétion ne fait qu'aggraver l'accumulation toxique d'-globine dans les pré-curseurs érythrocytaires [23,24]. L'anémie sévère qui en résulte explique la létalité des embryons homozygotes pour la mutation de la -globine.…”

“…D'autres études ont montré que cette kinase rédui-rait l'érythropoïèse inefficace causée par le stress oxydant induit par l'arsénite [24]. Dans les conditions de stress oxydant, la production de dérivés réactifs de l'oxygène (ROS, reactive oxygen species) est augmentée [25].…”

“…Dans les conditions de stress oxydant, la production de dérivés réactifs de l'oxygène (ROS, reactive oxygen species) est augmentée [25]. Or, ce taux élevé de ROS entraîne l'activation de HRI en induisant son autophosphorylation [24]. La kinase activée phosphoryle eIF2ce qui provoque une inhibition générale de l'initiation de la traduction, à l'exception toutefois de la traduction d'ARNm spécifiques ayant des petits cadres de lecture dans leur région 5'UTR (5' untranslated region) [26].…”