Page 18 - 2021_06-Haematologica-web

Y. Mathangasinghe et al.
Formation of red blood cells
Erythropoiesis is a vital process throughout vertebrate life, which helps maintain adequate tissue oxygenation under physiological and nonphysiological states (e.g., hypoxia, hemorrhage or other anemic conditions). This cell differentiation event leads to the generation of highly specialized erythrocytes that function as dedicated oxy- gen and carbon dioxide transporting cells across the body. Erythrocytes have a finite lifespan (approximately 120 days in humans) in the circulatory system before they are recycled mainly in the spleen by macrophages.6 These cells, therefore, must be continuously and rapidly replaced in vertebrates. About two million new erythro- cytes per second are generated in adult humans7 via pro- liferation and differentiation of a self-renewing popula- tion of pluripotent hematopoietic stem cells (HSC) locat- ed in the yolk sac, liver, spleen (antenatal) or bone mar- row (postnatal) that give rise to early erythroid progeni- tors.6 During erythropoiesis, these progenitors undergo a red cell lineage specific terminal differentiation program to generate mature erythrocytes.
Erythrocyte production is tightly regulated by a set of hormones. For example, glucocorticoids regulate both the proliferation and differentiation of early erythroid pro- genitors known as early burst forming unit-erythroid (BFU-E) cells.8 The proliferation and differentiation of subsequent erythroid progenitors including late stage BFU-E and colony forming unit-erythroid (CFU-E) cells occur after the stimulation by erythropoietin (EPO), a gly- coprotein cytokine secreted by the kidneys.9 EPO stimu- lation is vital for the induction of GATA-binding factor 1 (GATA-1) transcription factor, the master regulator of ery- thropoiesis. GATA-1 together with the transcription fac- tor STAT5, promote further erythroblast proliferation10 and turn on the gene activation and repression program, which drives the multistep terminal differentiation process of these cells.11 In mammals, erythropoiesis can be resolved into six morphologically distinct cell stages that result from a series of mitotic cell divisions (Figure 1A). These stages include: (i) proerythroblast, (ii) basophilic erythroblast, (iii) polychromatophilic ery- throblast, (IV) orthochromatophilic erythroblast, (V) retic- ulocyte, and (VI) mature erythrocyte. During terminal dif- ferentiation, the erythroblasts decrease in cell size, con- dense chromatin, reorganize and reduce cellular pro- teome and membranes, and eliminate organelles, thereby making space for the rapidly increasing levels of Hb, the oxygen-trafficking protein complex. The most striking morphological change occurs in orthochromatophilic ery- throblasts that eject nuclei to form reticulocytes in the bone marrow (Figure 1A). These reticulocytes loose ribo- somes and the bulk of RNA molecules, and develop into Hb packed mature erythrocytes that have a characteristic biconcave disk-like shape with a flattened center.
The mature erythrocytes contain a remarkably high concentration of Hb molecules (approximately 29.5 pg/cell),12 which represents approximately 98% of the proteome.13 Hb is a tetrameric globular protein made up of four globin protein subunits/chains, each containing a heme group, which reversibly binds to oxygen and car- bon dioxide. The adult human hemoglobin (HbA) is made from two 141 amino acid-long α-globin chains and two 146 amino acid-long b-globin chains (α2b2) (Figure 1B). Under physiological conditions, HbA makes up
>97% of the Hb constituent in adult humans. The
remaining Hb contains fetal Hb (HbF; α γ ) and HbA2 22
(α d ), two isotypes generated by switching the b globin
22
chain with either γ or d globin chains.
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Proteotoxic stresses associated with erythropoiesis
Proteostasis is maintained by balancing the cellular pathways that facilitate protein synthesis, folding, assem- bly, trafficking, and degradation under varying environ- mental and metabolic conditions.1 Even under normal growth conditions, cells experience a continuous influx of misfolded proteins generated from various protein bio- genesis mistakes such as errors in transcription, transla- tion and folding. Additionally, most proteins are at risk of misfolding due to the marginal stability of their native conformations.14 Cells have evolved a set of intricate PQC pathways comprising of molecular chaperones and pro- tein degradation systems that operate constantly to decrease the levels of misfolded proteins that otherwise would easily form aggregates in crowded cellular envi- ronments. Protein aggregates typically show poor solubil- ity in aqueous cellular environments, have no physiolog- ical function per se and could instead elicit cytotoxicity.1 By untangling and unfolding such aberrant protein species, the ATP fueled chaperone machineries are able to “repair” and rescue proteins, which leads to a consider- able reduction in the risk of proteotoxicities in cells. Protein degradation pathways also represent an impor- tant line of defense by clearing misfolded proteins and preventing their accumulation.1 However, in aging and/or stressed cells, such defense mechanisms could become overwhelmed leading to the buildup of potentially toxic protein aggregate species. Many disease-linked mutant proteins also form such aggregates that are refractory to PQC systems, including degradation pathways.15
Due to various protein biosynthetic errors and consid- erable attenuation of basic PQC pathways, erythroid maturation is highly exposed to protein misfolding/aggre- gation. The first PQC challenge during erythropoiesis involves the folding and assembly of the α-globin chains that show a high degree of instability. In the absence of the partnering b-globin, the free α-chains with heme/iron could readily misfold to form protein aggregate deposits called Heinz bodies (for a review, see Voon et al.16). These highly toxic protein aggregates, when accumulated, could trigger the generation of reactive oxygen species (ROS)17 that damage cellular proteins, nucleic acids and lipids and induce oxidative stress in erythroblasts leading to prema- ture cell death (Figure 1B).18 This is circumvented to a cer- tain degree with the assistance of a dedicated chaperone named alpha hemoglobin stabilizing protein (AHSP). AHSP mimics the α-helix-loop-α-helix motif of b-globin and assists in the folding of α-chains in a “template” directed manner19,20 (for a review, see Weiss et al.)21. Apart from the α-chain instability, the generally high synthesis rates of globin proteins (300 mg of Hb per hour in healthy adult humans22) during this atypical state could also pro- portionally increase the level of intrinsic errors in folding and assembly23 of Hb. In particular, heme/iron imbalances could result in globin misfolding, which could induce severe oxidative stress in erythroblasts. A tightly regulat- ed supply of iron to support the production of heme is
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haematologica | 2021; 106(6)