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Biomedical subjects

W F Anderson

Publications and source records attributed to W F Anderson.

At least 379 records · Page 21Linked to original sources
PubMed1973

Hemogloblin switching in sheep and goats: erythropoietin-dependent synthesis of hemoglobin C in goat bone-marrow cultures.

The anemia-induced switch from hemoglobin A (alpha(2)beta(2) (A)) to hemoglobin C (alpha(2)beta(2) (C)) synthesis occurring in vivo in sheep and goats has been reproduced in tissue culture of goat bone-marrow cells. Cultivation of primary cultures of goat bone marrow in the presence of erythropoietin results in the appearance of detectable amounts of beta(C) globin after 48-72 hr, as well as in a decrease in beta(A) globin. A population of proerythroblasts, as well as active heme and globin synthesis, are maintained for at least 3 days in erythropoietin-treated, but not in erythropoietin-deficient, cultures. These findings demonstrate (i) maintenance of erythropoietin-responsive cells from bone marrow in vitro, and (ii) switching in vitro from the synthesis of a globin chain coded by one gene to that coded by a different, nonallelic gene. Bone-marrow culture might be a useful model system for study of the mechanism of action of erythropoietin and for study of the activation (and inactivation) of specific genes in vitro.

Anemia↗
PubMed1973

A ribosome dissociation factor from rabbit reticulocytes distinct from initiation factor M3.

A ribosome dissociation factor (DF), from a 0.5 M KCl wash fraction of rabbit-reticulocyte-ribosomes, has been purified by Sephadex G-200, phosphocellulose, DEAE-cellulose, and hydroxyapatite chromatography. The most purified preparation displayed one major and several minor bands on 3.75% acrylamide gels.DF cannot replace IF-M(1), IF-M(2A), IF-M(2B), IF-M(2), EF-1, or EF-2 in poly(U)-directed polyphenylalanine synthesis at low Mg(++) concentrations or in endogenous mRNA-directed globin synthesis. Conversely, these initiation and elongation factors showed little or no dissociation activity, even when assayed at levels 5-10 times greater than those required to saturate a polypeptide synthesis assay. Reticulocyte DF thus appears to be a distinct factor.

Animals↗
PubMed1973

Hemoglobin messenger RNA from human bone marrow. Isolation and translation in homozygous and heterozygous beta-thalassemia.

A method for isolating human hemoglobin messenger RNA (mRNA) from bone marrow cells was developed to investigate the molecular basis for the defect in globin synthesis in beta thalassemia. Active mRNA was isolated from the bone marrow cells and peripheral reticulocytes of patients with homozygous beta thalassemia, heterozygous beta thalassemia, sickle cell trait, double heterozygosity for beta thalassemia and sickle cell trait, as well as from a patient with normal hemoglobin synthesis but with an elevated reticulocyte count secondary to hereditary spherocytosis. The mRNA was prepared for assay in an mRNA-dependent rabbit reticulocyte cell-free system and the amount of alpha and beta globin chains synthesized was determined by carboxymethylcellulose column chromatography. The relative synthesis of alpha to beta chains in response to normal hemoglobin mRNA was found to be a function of the amount of mRNA added to the assay system: increasing the amount of mRNA led to a decrease in the alpha-to-beta-chain synthetic ratio. Therefore, assays were carried out at limiting concentrations of mRNA. The molecular defect in homozygous beta thalassemia was shown to be carried in the mRNA of bone marrow cells as well as in the mRNA from peripheral reticulocytes, because much less beta than alpha globin was produced in the cell-free system in response to mRNA from either type of cell. In patients doubly heterozygous for beta thalassemia and sickle cell trait, little or no synthesis of beta(A) globin occurred in the bone marrow cells or the peripheral reticulocytes. The alpha to beta(S) synthetic ratio of the intact bone marrow cells was approximately 1, while the same ratio in the peripheral reticulocytes was between 1.5 and 2. The virtual absence of translatable beta globin mRNA in the mRNA prepared from the cells of these doubly heterozygous patients further demonstrates that the molecular defect produced by the beta thalassemia gene is in the beta globin mRNA.

Adolescent↗
PubMed1972

Initiation of hemoglobin synthesis: comparison of model reactions that use artificial templates with those using natural messenger RNA.

PARTIAL REACTIONS DESIGNED TO STUDY THE INDIVIDUAL STEPS IN THE INITIATION OF PROTEIN SYNTHESIS MAY BE DIVIDED INTO TWO GROUPS: artificial models using artificial mRNA templates and natural models using naturally occurring mRNA. The requirements for each of the reticulocyte initiation factors, IF-M(1), IF-M(2), and IF-M(3), and for GTP are examined using these models in order to determine if either type of model is a valid representation of the events occurring in natural initiation. Binding of the initiator tRNA, Met-tRNA(F), to endogenous mRNA requires IF-M(1), IF-M(2), and GTP. A requirement for hydrolysis of GTP is not found when the artificial template ApUpG is used since GDPCP may be substituted for GTP. Met-tRNA(F) bound to the template ApUpG by IF-M(1) + IF-M(2) can form a peptide bond with the aminoacyl-tRNA analog, puromycin. Met-tRNA(F) bound by IF-M(1) + IF-M(2) to the initiator codon of natural globin mRNA, however, cannot form a peptide bond with either puromycin or valine-tRNA unless IF-M(3) is also present. The requirements for Met(F)-valine synthesis on exogenous globin mRNA are the same as the requirements on endogenous mRNA. Synthesis of the initial tripeptides of the alpha and beta chains of rabbit globin, Met(F)-Val-Leu and Met(F)-Val-His, requires, in addition, leucyl-tRNA and histidyl-tRNA. It appears, therefore, that model systems that use natural messenger RNA can duplicate the factor and energy requirements of natural initiation, but that the model systems thus far studied that use artificial messengers as templates do not.

Animals↗
PubMed1972

Separation of reticulocyte initiation factor M 2 activity into two components.

IF-M(2), one of three initiation factors isolated by DEAE-cellulose chromatography from the 0.5 M KCl-wash fraction of rabbit reticulocyte ribosomes, has been separated by Sephadex G-200 chromatography into two components: IF-M(2A) and IF-M(2B). IF-M(2A) elutes near the void-volume, while IF-M(2B), which is much smaller in molecular weight than IF-M(2A), elutes slightly after a hemoglobin marker. In the presence of the other appropriate factors, both IF-M(2A) and IF-M(2B) are required to stimulate poly(U)-directed polyphenylalanine synthesis at low Mg(++) concentration, ApUpG-directed Met-tRNA(F) binding to washed reticulocyte ribosomes, and initiation of globin synthesis from endogenous mRNA. IF-M(2A) stimulates ribosome-dependent GTP hydrolysis, while IF-M(2B) does not; IF-M(2B) stimulates ApUpG-directed fMet-tRNA(F) binding in the presence of IF-M(1), while IF-M(2A) does not. Although IF-M(2A) and IF-M(2B) can be distinguished from each other by size and by activity, a distinct function for IF-M(2B) has not yet been found. Therefore, its precise role in the initiation process remains unclear.

Animals↗
PubMed1972

Hemoglobin switching in sheep and goats: change in functional globin messenger RNA in reticulocytes and bone marrow cells.

ANEMIA CAUSES A CHANGE IN THE TYPE OF CIRCULATING HEMOGLOBIN IN GOATS AND CERTAIN SHEEP: HbA (alpha(2)beta(2) (A)) is replaced by HbC (alpha(2)beta(2) (C)). We have isolated globin mRNA from erythroid cells of anemic and nonanemic animals to investigate the mechanism whereby anemia causes this switch. To study several stages in transition from beta(A) to beta(C) synthesis, active globin mRNA was isolated from bone marrow cells, as well as from reticulocytes. By assaying these globin mRNAs in a rabbit reticulocyte cell-free system, we have demonstrated that the switch from beta(A) to beta(C) globin synthesis is mediated via a change in functional globin mRNA.

Amino Acid Sequence↗
PubMed1971

Translation of exogenous messenger RNA for hemoglobin on reticulocyte and liver ribosomes.

The ribosome and initiation factor requirements for translation of rabbit-reticulocyte hemoglobin mRNA on rabbit reticulocyte ribosomes, reticulocyte ribosomal subunits, and liver ribosomes have been studied. Excellent synthesis of globin chains from exogenous mRNA in the fractionated cell-free system has been achieved. There is a near absolute requirement for each of the initiation factors, M(1), M(2), and M(3) (as well as for the supernatant proteins) for the translation of exogenous mRNA. Liver microsomal wash will partially replace reticulocyte factors M(1) and M(2), but will not replace the requirement for reticulocyte factor M(3). Rabbit liver ribosomes and rabbit reticulocyte ribosomes are equally active in their ability to support the translation of exogenous hemoglobin mRNA.

Animals↗