Combined effect of two different polymorphic sequences within the β globin gene cluster on the level of HbF

Abstract

␤ thalassemia and Hb Lepore heterozygotes included in this study exhibit fetal hemoglobin levels varying from trace quantities to 14% (1.74 g/dl) of total hemoglobin in the adult. In this work, we have examined the correlation of DNA sequence polymorphisms with the observed HbF level. The analysis of polymorphic markers within the ␤ globin cluster in 39 individuals heterozygous for ␤ thalassemia or Hb Lepore confirms the previous findings for homozygous ␤ thalassemia: the presence of both an (AT) 9 T 5 sequence configuration at position −540 of the ␤ globin gene and a (C → T) variation at −158 of the G␥ globin gene is associated with elevated expression of HbF. However, at least one defective ␤ globin gene is required to reveal this association. The best evidence is from the study of individuals heterozygous for Hb Lepore with various levels of HbF. In these individuals it was possible to explore the effect of a single (AT) x T y motif (the other being absent from the rearranged Lepore chromosome) on HbF expression. The presence of the (AT) 9 T 5 configuration increases HbF level from a median of 0.515 g/dl observed in (AT) 7 T 7 subjects, to 1.39 g/dl.We confirm the existence of linkage disequilibrium between the (C → T) variation at −158 of G␥ gene and the (TG) 13 configuration at the second intervening sequence (IVS-2) of A␥ gene and identify two new polymorphisms in this region: (TG) 7 (CG) 5 (TG) 8 linked to haplotype V and (TG) 8 (CG) 5 (TG) 10 linked to haplotype II. This study suggests that two distinct regions of the ␤ cluster, whether in cis or in trans to each other, can interact to enhance HbF expression when a ␤ thalassemic determinant is present in heterozigosity.

Spectrum of β thalassemia mutations and HbF levels in the heterozygous Moroccan population

“…Haplotypes III, IV, and IX were found in 42 independent chromosomes, normal or ␤ thal, in linkage disequilibrium with XmnI (+). As the same association had been previously observed in normal and in ␤ thal chromosomes carrying the Mediterranean haplotype III, IV, or IX, the ␤ thal chromosomes were classified as having 5Ј subhaplotype class A [1]. The remaining chromosomes were associated with XmnI (−) and classified as having 5Ј subhaplotype class B. Gene sequencing of individual I-1 from family 19 revealed a polymorphism at position −369 C→G of A␥ globin gene (http://www.ncbi.nlm.…”

“…In the absence of experimental data on the regulatory function of the G␥ and A␥ polymorphisms per se or as linkage markers of HbF expression, one could speculate that the -369 A␥ polymorphism, in association with the reduction of ␤ gene transcription by the TATA box mutation [46], possibly creates an imbalance of transcription factors, favoring an HbF elevation under the mild erythropoietic stress caused by ␤ thalassemia heterozygous condition. Epidemiological studies of ␤ thalassemia homozygotes, heterozygotes, and Hb Lepore heterozygotes [1,10,11], show the importance, for the regulation of fetal genes, of effector(s) located in both the adult and fetal domains of the cluster. The hypothesis of a regulatory mechanism based on transcription factors competition for the LCR cannot be excluded.…”

“…The chromosomes were divided into two classes according to their extended 5Ј subhaplotype: class A, in which the XmnI site was present and class B, in which it was absent. AA T , BB T , and BA T refer to the genotypes of ␤ thalassemic carriers [1]. The configuration of (AT) x T y alleles was determined by direct sequencing using the ABI Prism Big Dye technology from Applied Biosystems.…”

“…The synthesis of HbF moderates the ␣/non-␣ globin chain imbalance and it attains, in the homozygote, levels of 85%. In the heterozygote, the HbF levels are above 2.5% in the adult, with a dispersion of values up to 14% and are hereditarily transmitted [1,2]. Determinants for this type of hereditary persistence of fetal hemoglobin (HPFH) in normal adults, in ␤ thalassemia and drepanocytosis are multiple and not yet clearly defined [3,4].…”

“…A study with twin pairs of European descent shows that the contribution of ␤ globin cluster elements to the variability of HbF level accounts for about 13% of that variation, and 2% is related to age and sex [6]. Close linkage to the 5Ј subhaplotype and to the (AT) x T y motif at −540 of the ␤ globin gene in ␤ thalassemia or Hb Lepore heterozygotes [1,10] or in homozygotes [11] of European descent, was also observed.…”