Abstract
The two human alpha-globin genes, alpha 1 and alpha 2, are coexpressed in normal erythroid cells and encode identical alpha-globin protein products. Based upon genetic studies, it has been assumed that these two adjacent and highly homologous genes are equally expressed. In previous studies we have, however, demonstrated that the alpha 2 gene encodes a 2-3-fold higher steady state level of mRNA than the alpha 1 gene. In the present study, we monitor the relative levels of protein production from these two loci by quantitating the synthesis of specific alpha-globin structural mutants encoded by each alpha-globin gene. These values are then used to infer the relative contributions of the normal alpha 1 and alpha 2 loci to total alpha-globin production. The results of eight separate studies, each based upon a different alpha-globin structural mutant mapped to either the alpha 1 or the alpha 2 locus, are internally consistent. The data demonstrate that the alpha 2 gene encodes 2-3-fold more protein than the alpha 1 gene. These results suggest that the human alpha-globin gene cluster contains a major and a minor locus. The dominant expression of the alpha 2 gene predicts a greater impact of mutations at this locus, in comparison to mutations at the alpha 1 locus, in the generation of the alpha-thalassemia phenotype.
Highlights
The two human a-globin genes,a1 and a2, are coex- 16 [3,4,5,6,7]
We combine the modalities of DNA, mRNA,and impact of mutations atthis locus, in comparison to protein analysis to arrivaet a full description of a-globin gene mutations at the a1 locus, in the generation of the a- activity in these eight cases, and from this data we infer the thalassemia phenotype
Relative levels of activity of the normala1 and a2 genes. These eight studies yield a consistent finding: the a2 gene is functionally the major a-globin gene in humans,encoding two to three times as much a-globin protein as the adjaace1ngtene, a-Globin is an essential subuniotf the human hemoglobin This result predicts a significant difference in the impact of tetramer from the sixwtheek of development in utero through mutations at thetwo a-globin loci on a-globin synthesis and adult life [1].Fetal hemoglobin, a2y2, and adulthemoglobin, on the consequentseverity of a-thalassemia
Summary
The analysis demonstrates that the al-globin mRNA is uncontaminated with a2-globin mRNA. The analysoifs hybridselected&?-globinmRNA(lanes 5 and 6) demonstrates a blood samples were obtained from eight unrelated individuals,. Genetic profileof eight individuals with a-globinstructural mutants. J-Oxford, Queens, Hasharon, and Twin Peaks, were processed immediately after venipuncture while the remaining four samples, Russ, Winnipeg, Inkster, anGdPhiladelphia were processed after overnight delivery on wet Mutant am/a. Queens Hasharon Russ (15 Gly-Asp) 40 (34 Leu-Arg) 14 (47 Asp-H34is) 35 (51Gly-Arg) 17. Oxford mutationwas established by isolation of an abnormal G-Philadelphia (68Asn-Lys) 26 ND aa/aa 2.6 a-globin tryptic peptideby CMC chromatographyfollowed by Winnipeg (75 Asp-Tyr) 11 ND aa/aa 2.7 amino acid analysisof this peptide ona Beckman 118 amino Inkster (85 Asp-ValN) D 24 aa/aa 2.6 acid analyzer (data not shown). The level of mutant a-globin (am)chain as a percentage of a ND, not determined
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