Translation of β-Globin m-RNA in β-Thalassemia and the S and C Hemoglobinopathies

Rieder, Ronald F.

doi:10.1172/jci106822

Cited by 36 publications

(5 citation statements)

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“…f3-Globin mRNAs in /3+-thalassemic and normal reticulocytes are translated with equal efficiencies (15)(16)(17), and the f3-globin genes in /3+-thalassemia do not contain detectable deletions (4,5).…”

mentioning

confidence: 99%

Processing of human beta-globin mRNA precursor to mRNA is defective in three patients with beta+-thalassemia.

Maquat¹,

Kinniburgh²,

Beach³

et al. 1980

Proc. Natl. Acad. Sci. U.S.A.

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Nucleated bone marrow cells from normal individuals and from three patients with homozygous -+ thalassemia were pulse-labeled with tritiated nucleosides. The processing of the newly synthesized globin mRNA precursors was monitored by inhibiting additional transcription with ac- The f3-thalassemias are a heterogeneous group of hereditary anemias in man in which /3-globin protein synthesis is decreased or absent and a-globin protein synthesis occurs at normal rates (1-6). Erythrocytes from patients with homozygous f30-thalassemia contain no detectable f3-globin protein, and their reticulocytes either lack f3-globin mRNA or contain f3-globin mRNA that is not translated into functional protein (7-9). The f3-globin genes are intact in most, but not all, #°-thalassemic patients (4, 5, 10). Erythroid cells from patients with f3+-thalassemia contain structurally normal f3-globin protein, but there is significantly less f3-globin than a-globin protein and the reticulocyte a/f3-globin mRNA ratio is increased (11-14).f3-Globin mRNAs in /3+-thalassemic and normal reticulocytes are translated with equal efficiencies (15-17), and the f3-globin genes in /3+-thalassemia do not contain detectable deletions (4, 5).Based on these data, the fl+-thalassemia phenotype must result from mutations that specifically reduce the expression of the f3-globin structural genes. Such mutations might affect gene transcription, processing of the mRNA precursor, transport of mRNA from the nucleus to the cytoplasm, or stability of mature mRNA in the cytoplasm. The ratios of a-globin to f3-globin mRNA sequences in nuclear RNA from normal individuals and from two patients with 3+-thalassemia were approximately equal (13). This result suggests that the transcription rates of the two genes are similar in some patients.We have investigated the possibility that the 3-globin mRNA precursor is processed to mRNA inefficiently in the f+-thalassemias. Nucleated bone marrow cells were pulse-labeled with 3H-labeled nucleosides and chased with actinomycin D, and globin-specific RNA was analyzed. The results indicate that the processing of f3-globin RNA, but not a-globin RNA, occurs inefficiently in three unrelated patients with #+-thalassemia. Nucleated cells were obtained from bone marrow aspirates, and all manipulations prior to cell culturing were performed at 240C. Each aspirate was passed through a series of progressively larger gauge needles and centrifuged for 10 min at 300 X g. The buffy coat was removed and brought to 5 ml with growth medium (RPMI-1640 containing 20% fetal calf serum and 50 units of penicillin and 50 ,g of streptomycin per ml).Cells were homogenized by gentle pipetting, transferred to five 1-ml Wintrobe tubes, and centrifuged for 10 min at 300 X g. The nucleated cells were pooled, brought to 10 ml with growth medium, and centrifuged for 12 min at 200 X g. The cells were resuspended in medium to 2-10 X 107 nucleated cells per ml and incubated in a humidified atmosphere of 5% C02/95% air at 370C for 1 hr, at which time [3H] .u...

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mentioning

confidence: 99%

Processing of human beta-globin mRNA precursor to mRNA is defective in three patients with beta+-thalassemia.

Maquat¹,

Kinniburgh²,

Beach³

et al. 1980

Proc. Natl. Acad. Sci. U.S.A.

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“…Among them was patient S. D., with mild homozygous #-thalassemia, in whom contamination of pT5 with 8T5 was minimal. The elongation time of P-chains is not prolonged in P-thalassemic reticulocytes (11) and may be even faster than normal (12). It could not be prolonged in this particular patient since otherwise we would have found less, rather than more, chain imbalance in his nascent compared with completed, chains (see below).…”

Section: Methodsmentioning

confidence: 72%

“…This has recently been confirmed by studies of formyl-methionyl-tRNAw binding to ribosomes and formation of the initial peptide bond (10). Simi-larly, direct measurements have not revealed any delay in elongation of the affected chain in thalassemic reticulocytes (11,12). All these data were, however, collected from the study of peripheral blood reticulocytes.…”

Section: Introductionmentioning

confidence: 78%

Translational Control of Hemoglobin Synthesis in Thalassemic Bone Marrow

Cividalli¹,

Nathan²,

Lodish³

1974

J. Clin. Invest.

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A B S T R A C T Previous studies of P-thalassemic reticulocytes have implied a decreased amount of functional P-mRNA but unimipaired translation of the P-mRNA present. However, the P/a synthetic ratios in P-thalassemic marrow are higher than those observed in reticulocytes of the same patients. This could imply that marrow cells contain an abnormally functioning 8-mRNA no longer active in reticulocytes. To test the function of mRNA found in marrow, intact cells were incubated with ['3S]methionine and the relative amounts of nascent a-and P-chains on polysomes of different sizes were measured by tryptic digestion and determination of the specific activities of the respective peptides. Results showed that in normal and P-thalassemic marrow, as well as in reticulocytes, P-chain production, though deficient, occurs predominantly on larger polysomes than the production of a-chains. In one patient with severe thalassemia and very little production of P-chains in marrow or reticulocytes, d-chain synthesis was found predominantly on larger polysomes than a-chain synthesis. These results indicate that in P-thalassemic as well as in nonthalassemic marrow and reticulocytes, each P-and 8-mRNA initiates protein synthesis at a rate faster than does each a-mRNA, and suggest that the P-mRNA in contact with polyribosomes is normally functioning but quantitatively deficient in P-thalassemic marrow as well as in reticulocytes. No translational defect was detected in a similar study performed in reticulocytes of a patient with hemoglobin H disease,

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“…Studies on various aspects of Iolypeptide chain initiation and translation in homozygous f thalassemia have revealed no abnormalities (10)(11)(12)(13)(14)(15). However, when globin mRNA isolated from fi thalassemia reticulocytes is added to a heterologous cell-free system, much less human fi chain is synthesized than a chain (16)(17)(18).…”

mentioning

confidence: 99%

Quantitative Deficiency of Chain-Specific Globin Messenger Ribonucleic Acids in the Thalassemia Syndromes

Housman

Forget²,

Skoultchi³

et al. 1973

Proc. Natl. Acad. Sci. U.S.A.

114

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A hybridization assay procedure was devised that makes possible quantitation of the ratio of mRNA of alpha to mRNA of beta globin chains in an RNA sample. The assay uses the radioactive synthetic DNA copies obtained by incubation of RNA-dependent DNA polymerase of avian myeloblastosis virus with rabbit globin mRNA that is 80-90%4 enriched in mRNA specific for synthesis of alpha or beta globin chains. The rabbit alpha-chain mRNA is obtained from the postribosomal supernatant of rabbit reticulocyte lysates; the rabbit beta-chain mRNA is obtained from the largest polysomes of rabbit reticulocytes treated with L-O-methylthreonine. Sufficient homology exists between rabbit and human globin chains and globin mRNAs that the synthetic DNA copies of chainspecific rabbit globin mRNA hybridize with human globin mRNA. Applied to the study of globin mRNA isolated from reticulocytes of humans with alpha and beta thalassemia, the technique revealed marked quantitative deficiency of alpha-chain mRNA relative to beta-chain mRNA in alpha thalassemia and similar deficiency of beta-chain mRNA relative to alpha-chain mRNA in beta thalassemia. The thalassemia' syndromes are therefore characterized by true quantitative deficiency of the mRNA specific for the affected globin chain.

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Translation of β-Globin m-RNA in β-Thalassemia and the S and C Hemoglobinopathies

Cited by 36 publications

References 20 publications

Processing of human beta-globin mRNA precursor to mRNA is defective in three patients with beta+-thalassemia.

Processing of human beta-globin mRNA precursor to mRNA is defective in three patients with beta+-thalassemia.

Translational Control of Hemoglobin Synthesis in Thalassemic Bone Marrow

Quantitative Deficiency of Chain-Specific Globin Messenger Ribonucleic Acids in the Thalassemia Syndromes

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