2 α-Thalassaemia

α-Thalassaemias are genetic defects extremely frequent in some populations and are characterized by the decrease or complete suppression of α-globin polypeptide chains. The gene cluster, which codes for and controls the production of these polypeptides, maps near the telomere of the short arm of chromosome 16, within a G + C rich and early-replicating DNA region. The genes expressed during the embryonic (ζ) or fetal and adult stage (α₂ and α₁) can be modified by point mutations which affect either the processing-translation of mRNA or make the polypeptide chains extremely unstable. Much more frequent are the deletions of variable size (from ≈ 3 to more than 100 kb) which remove one or both α genes in cis or even the whole gene cluster. Deletions of a single gene are the result of unequal pairing during meiosis, followed by reciprocal recombination. These unequal cross-overs, which produce also α gene triplications and quadruplications, are made possible by the high degree of homology of the two α genes and of their flanking sequences. Other deletions involving one or more genes are due to recombinations which have taken place within non-homologous regions (illegitimate recombinations) or in DNA segments whose homology is limited to very short sequences. Particularly interesting are the deletions which eliminate large DNA areas 5′ of ζ or of both α genes. These deletions do not include the structural genes but, nevertheless, suppress completely their expression. Larger deletions involving the tip of the short arm of chromosome 16 by truncation, interstitial deletions or translocations result in the contiguous gene syndrome ATR-16. In this complex syndrome α-thalassaemia is accompanied by mental retardation and variable dismorphic features. The study of mutations of the 5′ upstream flanking region has led to the discovery of a DNA sequence, localized 40 kb upstream of the ζ-globin gene, which controls the expression of the α genes (α major regulatory element or HS-40). In the acquired variant of haemoglobin H (HbH) disease found in rare individuals with myelodysplastic disorders and in the X-linked mental retardation associated with α-thalassaemia, a profound reduction or absence of α gene expression has been observed, which is not accompanied by structural alterations of the coding or controlling regions of the α gene complex. Most probably the acquired α-thalassaemia is due to the lack of soluble activators (or presence of repressors) which act in trans and affect the expression of the homologous clusters and are coded by genes not (closely) linked to the α genes. The ATR-X syndrome results from mutations of the XH2 gene, located on the X chromosome (Xq13.3) and coding for a transacting factor which regulates gene expression. The interaction of the different α-thalassaemia determinants results in three phenotypes: the α-thalassaemic trait, clinically silent and presenting only limited alterations of haematological parameters, HbH disease, characterized by the development of a haemolytic anaemia of variable degree, and the (lethal) Hb Bart's hydrops fetalis syndrome. The diagnosis of α-thalassaemia due to deletions is implemented by the electrophoretic analysis of genomic DNA digested with restriction enzymes and hybridized with specific molecular probes. Recently polymerase chain reaction (PCR) based strategies have replaced the Southern blotting methodology. The straightforward identification of point mutations is carried out by the specific amplification of the α₂ or α₁ gene by PCR followed by the localization and identification of the mutation with a variety of screening systems (denaturing gradient gel electrophoresis (DGGE), single strand conformation polymorphisms (SSCP)) and direct sequencing.