Hb Bronte or α93(FG5)Val→Gly: A New Unstable Variant of the α2‐Globin Gene, Associated with a Mild α⁺‐Thalassemia Phenotype

Abstract

We report a new unstable variant identified in three carriers of a family from East Sicily; it was named Hb Bronte after the place from which the family originated. DNA sequencing from nucleotides -181 to +894 (alpha1) and to +884 (alpha2) revealed a GTG-->GGG substitution at codon 93 of the alpha2-globin gene. The MCV and MCH values were at the lower end of the normal range in the carriers. On cation exchange high performance liquid chromatography (HPLC), the Hb A2 level was apparently increased to around 6%, and a small abnormal peak (0.3-0.4%) was detected after Hb A2. Two abnormal bands were detected by cellulose acetate electrophoresis: a major band (about 3-4%) migrated between Hb A and Hb F; a minor band (<1%) migrated between Hb A2 and carbonic anhydrase. Normal values of Hb A2 were detected by DEAE microchromatography. On reversed phase HPLC the variant chain was not detected, and most likely it was eluted with the alpha chain peak. The isopropanol stability test was very slightly positive in the carriers. Hemolytic symptoms were absent with the exception of indirect bilirubin, which was at high borderline in 2/3 carriers. In biosynthesis in vitro, the specific activity of the alpha chains was much higher than that of the beta-globin chains, and the alpha/beta biosynthetic ratio in the mother and proband was of the beta-thalassemia (thal) type (2.24 and 2.54, respectively). Time course experiments showed that the increase of the 3H-specific activity of the peak containing normal and variant alpha chains was not linear and was much higher than that of beta chains; moreover, the alpha/beta biosynthetic ratio varied during the 2 hours incubation.

Effect of Mutations on mRNA and Globin Stability: The Cases of Hb Bernalda/Groene Hart and Hb Southern Italy

“…In vitro biosynthesis of globin chain in reticulocytes was performed in a carrier of Hb Southern Italy and in a normal control, as already reported. Globin chains were separated by reversed-phase HPLC [ 19 ].…”

α-Thalassemia Associated with Hb Instability: A Tale of Two Features. The Case of Hb Rogliano or α1 Cod 108(G15)Thr→Asn and Hb Policoro or α2 Cod 124(H7)Ser→Pro.

“…The α-thalassemia deletions were tested by gap-PCR [ 14 ]; point mutations were analyzed by multiplex ARMS and DGGE [ 15 – 16 ]. The α1- or α2-globin genes were sequenced from -181 to +884 (α2) and +894 (α1) as previously reported [ 17 ]. An ARMS-PCR assay for the definition of the heterozygous Hb Policoro genotype was set up.…”

“…The analysis of the RFLP RsaI at 5’ of the α2-globin gene (rs2541669) was carried out on PCR fragments. The SNPs α2+14 (HBA2:c.-24C>G) and α2+861 (HBA2:c.565G>A, rs2685121) were determined by DNA sequencing or DGGE analysis [ 16 – 17 ]. RFPL and SNP association with the mutated α-globin allele was assembled by family segregation studies.…”

Identification and molecular characterization of a novel 163 kb deletion: The Italian (ϵγδβ)0-thalassemia

“…For the identification of alpha-or beta-thalassemia defects, molecular analyses have been conducted as already reported [10][11][12][13] (see Supplementary Data).…”