Integrated protein quality-control pathways regulate free α-globin in murine β-thalassemia

Abstract

Cells remove unstable polypeptides through protein quality-control (PQC) pathways such as ubiquitin-mediated proteolysis and autophagy. In the present study, we investigated how these pathways are used in ␤-thalassemia, a common hemoglobinopathy in which ␤-globin gene mutations cause the accumulation and precipitation of cytotoxic ␣-globin subunits. In ␤-thalassemic erythrocyte precursors, free ␣-globin was polyubiquitinated and degraded by the proteasome. These cells exhibited enhanced proteasome activity, and transcriptional profiling revealed coordinated induction of most proteasome subunits that was mediated by the stress-response transcription factor Nrf1. In isolated thalassemic cells, short-term proteasome inhibition blocked the degradation of free ␣-globin. In contrast, prolonged in vivo treatment of ␤-thalassemic mice with the proteasome inhibitor bortezomib did not enhance the accumulation of free ␣-globin. Rather, systemic proteasome inhibition activated compensatory proteotoxic stress-response mechanisms, including autophagy, which cooperated with ubiquitin-mediated proteolysis to degrade free ␣-globin in erythroid cells. Our findings show that multiple interregulated PQC responses degrade excess ␣-globin. Therefore, ␤-thalassemia fits into the broader framework of protein-aggregation disorders that use PQC pathways as cellprotective mechanisms. (Blood. 2012; 119(22):5265-5275) IntroductionThe production of functional hemoglobin A (HbA) tetramers (␣ 2 ␤ 2 ) requires the coordinated synthesis and assembly of ␣-and ␤-globin protein chains and iron-containing heme groups. Individually, all HbA components are toxic to RBCs and their precursors, as illustrated by ␤-thalassemias, a common hemoglobinopathy in which ␤-globin gene (HBB) mutations cause the buildup of free ␣-globin. 1 These unpaired ␣ chains initiate an oxidative damage cascade and form damaging precipitates that contribute largely to the clinical problems associated with ␤-thalassemia.The pathophysiology of ␤-thalassemia bears similarities to a diverse group of protein-aggregation diseases affecting multiple organs (for review, see Khandros and Weiss 2 ). These disorders, which include Parkinson disease, Alzheimer disease, Huntington disease, amyotrophic lateral sclerosis, and ␣ 1 -antitrypsin deficiency, are caused by the accumulation of unstable, relatively insoluble proteins. It is believed that the affected cells can detoxify and remove these damaging proteins via multiple interacting biochemical pathways called protein quality-control (PQC) pathways, but that disease ensues when such compensatory mechanisms are overwhelmed (for review, see Ciechanover and Brundin,3 Ding and Yin, 4 and Jaeger and Wyss-Coray 5 ). Cellular PQC systems include molecular chaperones, ubiquitin-mediated proteolysis, and autophagy. Several lines of evidence suggest that ␤-thalassemic erythroid cells use PQC pathways to detoxify free ␣-globin (for review, see Khandros and Weiss 2 ): (1) the clinical severity of ␤-thalassemia is proportional to the degree of ␣-glo...

AG-348 enhances pyruvate kinase activity in red blood cells from patients with pyruvate kinase deficiency

“…For instance, b-thalassemia is a hemoglobinopathy characterized by the presence of excess a-hemoglobin chains, which are cleared by ATP-dependent proteolytic mechanisms. 38 It will be interesting to further investigate these possibilities in ex vivo studies on patient RBCs as well as in mouse models of disease.…”

Modified activin receptor IIB ligand trap mitigates ineffective erythropoiesis and disease complications in murine β-thalassemia

“…3 In the present study, we found that inhibition of Smad2/3 signaling reduces oxidative stress in erythrocytes, reduces accumulation of a-globin aggregates in erythrocyte membranes, improves erythrocyte morphology, and improves RBC life span. Although the mechanism responsible for this reduction in a-globin accumulation is unknown, potential explanations include an absolute decrease in a-globin chain production, enhancement of the known ability of erythroid cells to remove misfolded a-globin chains via proteosomes, 4 or hepcidin-mediated reduction in iron availability for synthesis of heme and globin. 22,42,43 Taken together, our results show that RAP-536, a modified ActRIIB ligand trap with Smad2/3 inhibitory activity, promotes terminal erythroid differentiation and mitigates IE in a mouse model of b-thalassemia.…”

“…3,4 Intracellular accumulation of free a-globin chains and precipitation of a-globin-heme complexes on red cell membranes in b-thalassemia generates proteotoxicity, inhibits late-stage erythroid differentiation, and is also thought to cause hemolysis of erythrocytes. 1,2,5,6 Ineffective erythropoiesis (IE) is a hallmark of b-thalassemia and promotes anemia, hypoxia, and elevated erythropoietin (EPO) levels.…”

α-Globin as a molecular target in the treatment of β-thalassemia

“…5,27,28 The a-globin monomers are degraded via several pathways including an adenosine triphosphate and ubiquitin-dependent proteolytic pathway, an autophagy pathway, and a nonenzymatic pathway triggered by ROS to result in the release of hemin (heme containing oxidized ferric iron) and free iron, which lodges in the cell membrane. [29][30][31][32] In addition, a-hemichromes are also found bound to the cytoskeleton. 27 In this unstable conformation, both the heme group and the iron are able to participate in redox reactions, leading to further generation of ROS, which damage cellular proteins, lipids, and nucleic acids.…”