Regulation of the expression of murine adult B-Globin genes

Author

Cynthia Jo Wawrzyniak

Date of Award

8-1986

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Major

Biomedical Sciences

Major Professor

Raymond A. Popp

Committee Members

Julian Preston, Lee Rusell, Bruce Jacobson, Bob Fujimura

Abstract

Hemoglobin switching is ideal for studying the basic regulation of genes during development. Here, a mouse model was used to demonstrate independent expression of the hemoglobin genes β1^major and β2^minor during erythroid differentiation and in response to anemia. cp Mice homozygous for the Hbb^s2 haplotype code for two adult β-globins, β-s2major and β-sminor, which can be distinguished from other embryonic and adult hemoglobins by cellulose acetate electrophoresis and ion exchange chromatography. At 11.5 days of gestation, β-s2major hemoglobin comprises under 20 percent and β-sminor hemoglobin over 80 percent of the adult β-globin present in circulating erythrocytes. The relative level of β-sminor hemoglobin decreases through fetal development; at birth β-sminor represents 34 percent of the β-globin in circulation. The adult value of 70 percent β-s2major and 30 per cent β-sminor hemoglobin in circulation is already expressed in mice at 6 days after birth.

To investigate whether the rapid shift in the ratio of β-sminor to β-s2major hemoglobin observed during gestation was due to a change in erythrocyte populations, the larger primitive and the smaller definitive red cells were separated according to size via centrifugal elutriation. Analysis of fetal samples at daily intervals from 12.5 through 17.5 days of gestation revealed that the larger red cells, which are yolk sac-derived, have relatively greater amounts of β-sminor hemoglobin than the smaller red cells, which are derived from the fetal liver.

Dot blot hybridization was used to determine the relative amounts of the β-s2major and β-sminor globin RNAs present in circulating reticulocytes of homozygous Hbb^s2 mice at 14.5 and 17.5 days of gestation and at birth relative to the amount of these RNAs in induced reticulocytes of adults. The level of β-sminor RNA in reticulocytes at 14.5 and 17.5 days of gestation is nearly the same as that in induced reticulocytes of adult mice. In contrast, the level of β-s2major RNA in reticulocytes at 14.5 days of gestation is 23 percent, and at 17.5 days of gestation is 66 percent, of the amount found in induced reticulocytes of adult mice. These data indicate that the differential expression of β-sminor and β-s2major hemoglobins during development is regulated at the level of transcription.

Perturbation of the relative quantities of the two adult hemoglobins and the two adult globin RNAs by hematopoietic stress was evaluated in adults. β-sminor hemoglobin constitutes 29 percent of the total 6-globin of normal adults. In anemic mice, this proportion is raised to the following levels: 34 percent in β-thalassemic heterozygotes; 38 percent in α-thalassemic heterozygotes; and 41 per cent in doubly heterozygous α-thalassemic, β-thalassemic mice. The altered levels of globin RNAs in thalassemic mice demonstrate that the increase in the β-sminor hemoglobin is accompanied by an increase in β-sminor RNA. Thus, a level of murine adult β-globin gene expression can be regulated transcriptionaly, which differs in the subpopulations of erythrocytes.

Recommended Citation

Wawrzyniak, Cynthia Jo, "Regulation of the expression of murine adult B-Globin genes. " PhD diss., University of Tennessee, 1986.
https://trace.tennessee.edu/utk_graddiss/12492