Translational control by heme-regulated eIF2α kinase during erythropoiesis

Abstract

Purpose of review: This review will provide an overview of the translational regulation of globin mRNAs and integrated stress response (ISR) during erythropoiesis by heme-regulated eIF2α kinase (HRI). HRI is an intracellular heme sensor that coordinates heme and globin synthesis in erythropoiesis by inhibiting protein synthesis of globins and heme biosynthetic enzymes during heme deficiency.

Recent findings: It has been demonstrated recently that HRI also activates the eIF2αP-activating transcription factor 4 (ATF4) ISR in primary erythroid precursors to combat oxidative stress. During chronic iron/heme deficiency in vivo, this HRI-eIF2αP-ATF4 signaling is necessary both to reduce oxidative stress and to promote erythroid differentiation. Augmenting eIF2αP signaling by the small molecule salubrinal, which inhibits dephosphorylation of eIF2αP, reduces excess α-globin synthesis and enhances translation of ATF4 mRNA in mouse β-thalassemic erythroid precursors. Intriguingly, salubrinal treatment of differentiating human CD34⁺ cells in culture increases fetal hemoglobin production with a concomitant decrease of adult hemoglobin by a posttranscriptional mechanism.

Summary: HRI-eIF2αP-ATF4 stress signaling is important not only to inhibit excess globin synthesis during erythropoiesis, but is also critical for adaptation to oxidative stress and for enhancing effective erythropoiesis. Modulation of this signaling pathway with small chemicals may provide a novel therapy for hemoglobinopathy.

Fig. 1. Family of eIF2α kinase and the integrated stress response of eIF2αP signaling

(A) Protein structure of HRI. There are two heme binding domains in HRI, the N-terminus and the kinase insertion region. The Roman numerals above the gray regions denote the conserved kinase domains of eIF2α kinases. (B) Integrated stress response. In mammalian cells, there are four eIF2α kinases that are activated in response to four major stress, nutrient starvation, viral infection, heme deficiency and ER stress. Activated eIF2α kinases phosphorylate eIF2α and inhibit protein synthesis to prevent proteotoxocity of unfolded protein occurs in ER stress and globin inclusions in heme deficiency. In addition, eIF2αP selectively enhances the translation of ATF4 mRNA. ATF4 then initiates an adaptive gene expression to mitigate stress.

Fig. 2. Up-regulation of the translation of ATF4 mRNA by eIF2αP upon stress

In the 5’UTR of ATF4 mRNA, there are two uORFs that are preferentially translated under non-stressed conditions and prevent the downstream translational initiation at the coding sequence of ATF4 mRNA. As initiating 40S ribosomal subunits scan from the cap structure, translation starts at the uORF1. After termination of translation, the 40S subunit remains associated with mRNA and reinitiates efficiently at uORF2 under non-stressed conditions. Upon stress, elevated eIF2αP impairs the reinitiation of 40S at uORF2 due to limiting functional eIF2. Thus, 40S continues to scan downstream and initiates at the AUG codon of coding sequence of ATF4 mRNA permitting the synthesis of ATF4 protein.

Fig. 3. The HRI-eIF2αP-ATF4 signaling during erythropoiesis

The expression of HRI increases during erythroid differentiation from BFU-E to reticulocytes. HRI is the major eIF2α kinase in the erythroid lineage, and is indispensable to coordinate heme and globin synthesis. Additionally, HRI activate ATF4 signaling pathway to mitigate oxidative stress in nucleated erythroblasts. This HRI-ATF4 pathway is activated during erythropoiesis and is necessary to promote erythroid differentiation. At the earlier stages of erythropoiesis before basophilic erythroblasts, other eIF2α kinases and HRI are both required for the proliferation of the erythroid precursors.

Fig. 4. Dephosphorylation of eIF2αP in the regulation of the HRI-eIF2αP-ATF4 signaling

The homeostasis of cellular eIF2αP level is controlled not only by the activation of HRI, but also by dephosphorylation of eIF2αP by PPase1 in order to regenerate active eIF2. Salubrinal, a small chemical molecule, is a selective inhibitor of dephosporylation of eIF2αP by interfering with the recruitment of eIF2αP to PPase1. Thus, treatment of cells with salubrinal results in increased eIF2αP level and the strength of its ISR resulting in reduction of α-globin protein synthesis and induction of ATF4 in mouse β-thalassemic erythroid precursors. In human erythroid cells, γ-globin synthesis is increased upon salubrinal treatment. These triple actions of salubrinal in erythroid precursors make it an extremely attractive candidate for treatment of β-thalassemia and other hemoglobinopathy.