The Synthesis of Fetal Hemoglobin Types in red Blood Cells and in BFU-E Derived Colonies from Peripheral Blood of Patients with Sickle Cell Anemia, β⁺- and δβ-Thalassemia, Various forms of Hereditary Persistence of Fetal Hemoglobin, Normal Adults and Newborn

Abstract

The biosynthesis of two types of human fetal hemoglobin (Hb F), namely Hb F with ^Gγ chains having glycine in position 136 and Hb F with Ay chains having alanine in position 136, was studied in blood samples and in cultures of erythroid precursors from blood of patients with different hemoglobinopathies. High pressure liquid chromatography (HPLC) was adapted to allow the separation of the methionyl-containing tryptic peptides ^GγT-15 and ^AγT-15 (which include the Gly → Ala polymorphism at position 136) from a digest of microquantities of ³⁵S-methionyl labelled Hb F. This method was sensitive enough to quantitate the relative production of the ^Gγ and ^Aγ chains by erythroid colonies derived from cloned Burst Forming Units (BFU-E) which were cultured for 16 days on methylcellulose. The production of Hb F in these colonies was generally higher than the level of Hb F in blood except for subjects with the GγAγ-HPFH heterozygosity. The Gγ to Aγ ratio in the Hb F produced in cultures of cells from Gγδβ-thalassemia or Gγ-HPFH heterozygotes was lower and that from Aγ-HPFH heterozygotes was higher than the ratios in the Hb F of the corresponding peripheral blood cells. Mixtures of Gγ and Aγ chains were present in cell cultures of SS patients, β-thalassemia homozygotes and GγAγ-HPFH heterozygotes in a ratio similar to that in the Hb F of mature red cells. These data suggest that erythroblasts in BFU-E derived colonies reactivate all available γ chain structural genes, both in cis and in trans to the abnormal determinant.

Hb F biosynthesis by adult blood samples concerns primarily the Gγ chains. This was particularly striking for blood samples in which erythroblasts were absent and the biosynthesis took place in fetal reticulocytes. Thus, the F-reticuloytes in blood of Aγ-HPFH heterozygotes with about 5% Hb F of the Aγ type produced primarily Hb F with Gγ chains. Similar differences were observed for GγAγ-HPFH heterozygotes and, less strikingly, for SS patients. A satisfactory explanation for this observation has not yet been obtained.