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Disorder of thalassemias and hemoglobinopathies: A genetic overview

Osama Atif

By Osama Atif

Osama is a biotechnology undergraduate student at the Faculty of Science, Mansoura University and a student researcher at biotechnology field. He is interested in Medical Genetics and Molecular Pathology.


The thalassemia syndromes are genetic disorders characterized by absent or deficient synthesis of one or more of the normal globin chains. Absent globin synthesis is designated with a (°) superscript, e.g. β°-thalassemia, while the presence of some (but not enough) of the gene product is noted by a “+” superscript, e. g. β+-thalassemia.

When there is a partial synthesis of the affected globin chain, it is usually structurally normal, therefore, the defect is a quantitative one secondary to unbalanced globin synthesis. This contrasts with the hemoglobinopathies in which the variant haemoglobins are qualitatively or structurally abnormal.


The thalassemias are a heterogeneous group of disorders and are classified according to the particular globin chain or chains synthesized in reduced amounts, i.e. alpha, beta, or delta-beta thalassemia.


The pathophysiology is similar in all forms of thalassemia. An imbalance of globin chain production results in the accumulation of free globin chains in the red blood cell precursors. These are very insoluble and precipitate, resulting in the hemolysis of RBCs, i.e. Hemolytic anaemia, with consequent compensatory hyperplasia of the bone marrow.

Molecular basis

The thalassemia syndromes provide examples of defects at essentially all different steps of globin synthesis.

i) Deletion of the DNA sequences coding for the structural gene.
ii) Point mutations in the β-globin gene.
iii) Most deletions in β-globin cluster affect more than the β-globin gene, e.g. δβ-thalassemia, HPFH, and Hb-Lepore.

I- Deletion of the DNA Sequences Coding for the Structural Gene
(Occurs in most α-thalassemias and in certain rare types of β-thalassemia)

Alpha Thalassemia

Alpha thalassemia is due to an under-production of alpha chains. It occurs most commonly in persons from South-East Asia.

α-thalassemia genes affect synthesis of α-chains which are essential for embryonic, fetal and adult Hbs. Deficiency of α-chains causes excess of β, γ, δ chains in the fetal and neonatal Periods which cause the formation of Hb-Barts (γ4) and later of Hb-H (β4).

Types of α-thalassemia (Figure1)

Types of α-thalassemia: (Figure1)
Figure 1: Types of α-thalassemia

Four clinical types are seen, depending upon the number of α-genes affected:

1. α-thal1 homozygote (α°-thal): hydrops fetalis with Hb Barts (γ4).

• Genotype (–/–) with deletion of 4 α-genes. Both parents are α-thal1 (heterozygote) trait.
• Clinical and Hb-findings: Over 80% of the haemoglobin is Hb-Barts (γ4 tetramer), which has a very high O2  affinity (causing severe tissue hypoxia) and precipitate causing an inclusion body hemolytic anaemia cause severe in utero anaemia leads to hydrops fetalis.

2. Hb-H disease: Cooley’s anaemia with Hb-B (β4).

• Genotype (–/α-) with deletion of 3 α-genes. One parent is α-thal1 trait and the other is α-tl2 trait.
• Clinical and Hb-findings: Hb-H (β4 tetramer) has a very high oxygen affinity and precipitate causing an inclusion body hemolytic anaemia which called Cooley’s anaemia.

3. α-T thalassemia trait (minor):

• Genotype: Either    –/αα (Heterozygous α-thal1) : deletion of two α-genes or  -α/-α (Homozygous α-thal2): deletion of two α-genes.
• Clinical and Hb-findings: Thalassemia minor which is usually clinically normal.

4. Silent carries:

• Genotype: aa/-a (Heterozygous α-thal2): deletion of only one α-gene.
• Clinical and Hb-findings: Normal.

Hb constant spring

Molecular Diagnosis of α-Thalassemia:

1. Southern blot by specific gene probe

2. Recently, PCR : by direct visualization with ethidium bromide (absence α-gene amplification).

II- Point Mutations in the β-Globin Gene

>Less than 150 mutations which can produce β-thalassemia

Beta ThalassemiaBeta Thalassemia

Beta thalassemia is due to underproduction of the -chain of Hb. In thalassemia major, or Cooley’s anaemia as it was known, the child usually presents by 6-mnths of age with severe transfusion-dependent anaemia. Unless the child is adequately transfused, compensatory expansion of the bone marrow results in an unusually-shaped face and skull. (Figure 2)

Mutational basis: There are 4 main functional types of mutations:

1. Transcription mutations = Promotor mutations:

• Mutation in promotor site which decreases the transcription process
• Mild (β+)
• e.g. 87 from the cap site (β+-87).

2. mRNA modification mutations = capping / polyadenylationt mutations:

• Mutations in cap sile (5′) and polyadenylation site (3′) of mRNA cause abnormal transportation of mRNA to the cytoplasm with consequently reduced levels of translation.
• Results in β+ thalassemia.

3. mRNA splicing mutations

Mutations involving:

A. Splice Junctions :- GT or AG dinucleotides of the introns cause β° thalassemia. .g. intron 1-nucleotide 1 (β+IVSI-1), intron 2-nucleotide 1 (β+IVSII-1).

B. Consensus sequences:- (adjacent bases) around splice junction decrease the ability of this RNA to splice correctly but still result in the detectable amount of normal β-globin leading to β+ thalassemia e.g. Intron 1-nucleotide 6 (β+IVSI-6)

C. Introns to produce new acceptor AG dinucleotide splice site sequence (cryptic splice site). The cryptic splice site competes with the normal splice site leading to reduced levels of the normal beta-globin mRNA leading to β+ thalassemia e.g. intron 1-nucleotide 110 (β+IVSI-110), intron 2-nucleotide 754 (β+IVSII-754).

*Splicing mutation: abnormal splicing with consequently reduced levels of β-globin mRNA (β° or β+).

4. Chain termination mutations: (translation defects)

• Chain termination mutations lead to shortened β-globin mRNA, which is often unstable and rapidly degraded. This leads to reduced levels of translation of an abnormal β-globin leading to β° thalassemia (Figure 3).

Figure 3: Point mutations in β° thalassemia
Figure 3: Point mutations in β° thalassemia

Chain termination mutations may be:

A. Frame-shift mutations: deletion or additions of 1,2 or 4 nucleotides which change the ribosome rending frame and cause premature termination of translation of β-globin mRNA e.g. deletion of 1 base at position 6 (β°6-4bp).

B. Non-sense codon mutation: point mutation changes a codon into termination codon → premature termination of translation of β-globin mRNA. e.g. Non-sense mutation at codon 39 (β°39).

Clinical Aspects of β-Thalassemia:

  • Persons homozygous for β-thalassemia mutations have severe transfusion-dependent anaemia, especially in β° type.
  • Individuals heterozygous for β-thalassemia have thalassemia trait or thalassemia minor and usually experience no symptoms. They do have, however, a mild hypochromic, microcytic amenia.
  • Because of the marked mutational heterogeneity seen in β-thalassemia, affected individuals are often compound heterozygotes i.e. have different mutations in their β- globin genes, leading to a wide range of severity of the disease. One form of β-thalassemia of intermediate  severity requires less frequent transfusions and is known as thalassemia intermedia.
  • Affected children with β-thalassemia develop hepatosplenomegaly secondary to extramedullary hematopoiesis and a characteristic Oriental facial features due to
  • excessive intramedullary hematopoiesis. The major causes of mortality are hemochromatosis and overwhelming infections following splenectomy.

Diagnosis of β-Thalassemia

1. Hematologic Features: Hb electrophoresis

• β° homozygous: Hb-F > 90%, no Hb-A, Hb-A2 increased.
• β+ homozygous: Hb-A : 20-40%, Hb-F : 60-80%.
• β° and β+ heterozygous: increased Hb-A2 (≃ 5%), and slightly increase in Hb-F (≃ 5%).

2. Molecular Diagnosis: to confirm diagnosis, carrier detection and prenatal diagnosis.

I. PCR: direct detection of mutations:

a. Dot blot (allele-specific oligonucleotide) and reverse dot blot: probes complementary to most common mutations.
b. ARMS (amplification refractory mutation system allele-specific priming of the PCR.

II. PCR: RFLP linkage analysis.

III. Most deletions in β-globin cluster affect more than the β-globin gene (as in δβ-thalassemia, HPFH, and Hb-Lepore)

a. Delta-beta (δβ) thalassemia:

  • Mutational basis: δβ-Thalassemia is due to extensive deletions of delta and beta globin structural genes.
  • Clinically: Persons homozygous for δβ-Thalassemia produce no delta or beta globin chains.
  • Although one would except such persons to have a fairly profound illness, they are only mildly anaemic due to increased production of gamma globin chains.

b. Hereditary Persistence of Fetal Hemoglobin HPFH:

Mutational basis:

1. Deletion type: due to deletions of the delta and beta globin genes.

2. Non-deletion type: due to point mutations in the promoter region of either Gγ or Aγ globin genes which is involved in the control of expression of the haemoglobin genes.

– Clinically: persons with HPFH continue to produce significant amounts of fetal Hb after birth. This is not associated with any medical problems.

– Mutational basis: unequal crossing over between mis-paired δ and β globin genes leading to δ and β fusion with segments of δ, β lost.

– Clinically: Like β° thalassemia.

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