Globin Types Research Articles

Synthesis of Adult-Type Hemoglobin in Human Erythremia Cell Line

KMOE -2/05 cells, derived from a patient with acute erythremia, became benzidine-positive after the addition of cytosine arabinoside (CA). Radioimmunoassays using antihuman hemoglobin antibodies revealed an elevated amount of hemoglobin in the CA-exposed cells over that in the control cells (without CA). Isoelectric focusing of the CA-exposed cell lysate formed benzidine-positive foci in the positions of human adult Hb (HbA) and human fetal Hb (HbF). To determine the types of globin synthesized in the CA-exposed cells, globin chains internally labeled with [3H] leucine were purified by carboxy-methyl (CM)-Sephadex column chromatography, immunoadsorption by Sepharose-coupled antihuman Hb antibodies and Sephadex G-100. The labeled globin chains were finally separated by CM-cellulose chromatography in urea. Two distinct peaks of radioactivity were shown in exactly the same fractions as carrier human globin alpha- and beta-chains. These observations indicate that these KMOE -2/05 cells synthesize HbA.

Genetic organization of the polymorphic equine alpha globin locus and sequence of the BII alpha 1 gene.

The equine alpha globin gene complex comprises two functional alpha genes and an alpha-like pseudogene arranged in the order 5'-alpha 2-(5kb)-alpha 1-(3kb)-psi alpha-3'. A single (embryonic) zeta-like sequence lies within a 12 kb region 5' to the alpha 2 gene. We have determined the sequence of the alpha 1 gene of the BII haplotype, one of two most common haplotypes (the other being BI) which encode alpha globins with either Tyr (BI) or Phe (BII) at codon 24 in both linked alpha genes. In BI and BII the non-allelic alpha 2 and alpha 1 genes respectively code for Gln or Lys at codon 60, thus accounting for the 4 alpha globin types seen in BI/BII heterozygotes. Genomic restriction enzyme maps of the BII alpha complex (24Phe/60Lys,Gln) and the allelic BI (24Tyr/60Lys,Gln) are identical to each other, and to those of a rarer normal haplotype, A, which encodes only alpha 24Tyr/60Gln globin, and a low expression mutant of BII which encodes only 24Phe/60Lys globin. These two latter haplotypes must therefore have a linked pair of alpha genes, as in BI and BII, but with identical coding properties, and it is suggested that this has arisen by gene conversion.

Hemin Preferentially Stimulates Synthesis of Alpha-Globin in K562 Human Erythroleukemia Cells

K562 human erythroleukemia cells are an established cell line derived from an adult with chronic myelogenous leukemia. Hemin stimulates their synthesis of embryonic and fetal hemoglobins. We have found that their globin synthetic pattern depends on the concentration of added hemin. Clone RA6 was cultured with 0--100 microM hemin and the globin synthetic pattern determined by 3H-leucine incorporation and analysis of 3H-globins by polyacrylamide gel electrophoresis in Triton X acid urea followed by fluorography and densitometry. The higher the hemin concentration, the greater the synthetic rate of each type of globin. However, the relative increase was greatest for alpha-globin. We propose that the differential dependence of alpha synthesis on added hemin is a reflection of translational inefficiency of alpha messenger RNA and that this property is exposed when the translational capacity of the cell is limited by hemin deficiency. We suggest that the differential dependence of alpha-chain synthesis on added hemin in clone RA6 is evidence of an intrinsic deficiency in heme synthesis.

Hemin preferentially stimulates synthesis of alpha-globin in K562 human erythroleukemia cells

K562 human erythroleukemia cells are an established cell line derived from an adult with chronic myelogenous leukemia. Hemin stimulates their synthesis of embryonic and fetal hemoglobins. We have found that their globin synthetic pattern depends on the concentration of added hemin. Clone RA6 was cultured with 0--100 microM hemin and the globin synthetic pattern determined by 3H-leucine incorporation and analysis of 3H-globins by polyacrylamide gel electrophoresis in Triton X acid urea followed by fluorography and densitometry. The higher the hemin concentration, the greater the synthetic rate of each type of globin. However, the relative increase was greatest for alpha-globin. We propose that the differential dependence of alpha synthesis on added hemin is a reflection of translational inefficiency of alpha messenger RNA and that this property is exposed when the translational capacity of the cell is limited by hemin deficiency. We suggest that the differential dependence of alpha-chain synthesis on added hemin in clone RA6 is evidence of an intrinsic deficiency in heme synthesis.

Control of gene expression during erythroid cell differentiation.

Erythroid cell differentiation has proved to be a useful system for exploring several aspects of gene expression during mammalian cell differentiation (1,2). In particular, fetal mouse erythropoiesis has been employed as a system to study: (a) the basis of changes in types of globins formed during fetal development; (b) mitotic activity of differentiating erythroblasts in relation to the synthesis of globins; (c) the number of structural genes coding for globins; (d) the relation of mRNA synthesis to globin formation during erythroid cell differentiation and (e) the mechanism of the erythropoietin effect in stimulating hemoglobin synthesis.

THE CONTROL OF HAEMOGLOBIN SYNTHESIS

Summary1. Haem and globin are synthesized separately by living cells before their conjugation to form haemoglobin.2. Protohaem is the form of haem utilized in the structure of haemoglobin, myo‐globin, erythrocruorin, catalases, some peroxidases and cytochromes of class B. All cells capable of aerobic respiration have the potential ability to synthesize haem.3. The main control mechanism in haem production is exercised by the enzyme ALA synthetase, and by cellular control over the level of activity of this enzyme.4. Haem probably affects the rate of globin synthesis but an effect of globin on haem synthesis is uncertain.5. Many organisms synthesize a variety of globins, some apparently exclusive to particular developmental stages. We suggest that the multiplicity of types of globin, whether changing during development, or present in balanced proportions in the adult, are not the result of evolutionary selection but of non‐adaptive changes in multiple copies of the primitive globin cistron.6. Ontogenic changes in the type of globin synthesized seem to be effected by differing mechanisms in different organisms. In some animals the changes seem to occur within one line of cells, whereas in others there is evidence for replacement of one cell population by another as in the switch from the embryonic to the adult or from tadpole to adult form of haemoglobin.7. Both transcriptional and post‐transcriptional control mechanisms are involved in globin synthesis, but the effects of the cell cycle and cell division on such regulation deserve intensive study in eukaryotic systems. We suggest that studies of globin synthesis provide evidence favouring some form of ‘linear reading’ of the genetic message.8. The hormones thyroxine and erythropoietin both affect haemoglobin synthesis in at least some organisms, and there are similarities in the pattern of action of the two hormones on erythropoietic tissues.9. All classes of RNA must affect globin synthesis and the isolation of globin mRNA has provided a particularly useful biochemical tool.10. Haemoglobin synthesis in artificial heterokaryons is of considerable interest but still poorly understood.11. The final assembly of tetrameric haemoglobin involves the conjugation of free haem with αβ globin dimers, the β globin chains apparently remain bound to ribo‐somes until they join with an α globin from the cellular pool of α globin. The mixed tetramer which is the most common form of haemoglobin possesses functional advantages over tetramers of only one globin type.

Myoglobin-facilitated oxygen diffusion: role of myoglobin in oxygen entry into muscle.

Myoglobin-facilitated oxygen diffusion: role of myoglobin in oxygen entry into muscle.