Molecular and hematologic characterization of Scottish-Irish type (epsilon gamma delta beta)zero thalassemia

The DNA deletion associated with an example of (epsilon gamma delta beta)zero thalassemia (Scottish-Irish type) was characterized. The deletion is approximately 205 kb in length and involves the epsilon, G gamma, A gamma, delta, and beta globin genes. The breakpoint is located 263 bp 3' to exon 3 of the beta globin gene. An LI (KpnI) repeat element approximately 320 bp in size is found at the 3' end of the novel DNA sequence. Different clinical phenotypes for three heterozygous neonates suggest that the deletion alone does not predict severity of (epsilon gamma delta beta)zero thalassemia at this age.     

A novel deletion causing (epsilon gamma delta beta) degrees thalassaemia in a Chilean family

We describe a novel deletion causing (epsilongammadeltabeta) degrees thalassaemia segregating in three generations of a Chilean family of Spanish descent. Heterozygotes for the deletion were all affected by neonatal haemolytic anaemia. The deletion of 152,569 bp extends from 77 kb upstream of the epsilon gene to 31 kb downstream of the beta gene, and includes the entire beta-globin gene cluster and two upstream olfactory receptor genes. Comparison of the sequences of the deletion junction with those of the flanking normal DNA suggests that the deletion results from a non-homologous recombination event. The insertion of 16 'orphan' nucleotides in the deletion junction creates a perfect inverted repeat of 12 nucleotides, forming a 12-bp stem with a four-nucleotide loop that could have contributed to the illegitimate recombination. The 3' breakpoint is located within an L1 family repeat that contains a perfect 160-bp palindrome, and is in close proximity to the 3' breakpoints of five other deletions in the beta cluster - Indian (HPFH-3), Italian (HPFH-4) and Vietnamese GgammaAgamma (deltabeta) degrees HPFH, German and Belgian Ggamma (Alphagammadeltabeta) degrees thalassaemia.     

A novel epsilon gamma delta beta thalassemia of 1.4 Mb deletion found in a Japanese patient

A novel large deletion, causing epsilon gamma delta beta thalassemia (here called, epsilon gamma delta beta thalassemia Jpn-I) was discovered in a 6-year-old Japanese boy. He was born uneventfully, but revealed thalassemia minor after birth. The mutation was inherited from his mother. The deletion, caused by an illegitimate recombination extended from 750 kb upstream to 660 kb downstream of e-globin gene, and removed about 1.4 Mb of DNA, the largest in epsilon gamma delta beta thalassemias. A 19-nucleotide orphan sequence and direct repeats were present at the junction. The deletion lost several functional genes, but no relevant symptoms manifested. The breakpoints were determined by relatively simple methods.     

Molecular characterization of a novel 27.6-kb deletion causing α<Superscript>+</Superscript> thalassemia in a Chinese family

Over 80% of the α-thalassemia cases in southern China are caused by large deletions involving the α-globin gene cluster on chromosome 16p13.3. Here, we characterized a novel 27.6-kb deletion on the α-globin gene cluster in a Chinese family. Its breakpoints were detected to lie between coordinates 9079 and 36718 of the α-globin gene cluster (NG_000006.1), with a total of 27,640 nucleotides deleted. It was designated as −α ^27.6 deletion. The proband is a compound heterozygote of −−^SEA and −α ^27.6 and he displayed very mild hemoglobin H disease phenotype with Hb 7.9–9.3 g/dl. Phenotypic analysis on heterozygote of this deletion revealed it as α⁺ mutation. It leads to a very mild phenotype as adult heterozygotes have normal hematological parameters with the values at the lower border of the normal range. RT-PCR analysis showed that the α-globin mRNA level of the heterozygotes was decreased when compared with that of normal people.     

A novel delta beta fusion gene expresses hemoglobin A (HbA) not Hb Lepore: Senegalese delta(0)beta(+) thalassemia

This study identified and characterized a novel delta beta fusion gene in which the delta-globin gene promoter is linked to intact beta-globin coding sequences in a Senegalese family. It results from a 7.4-kb deletion that removes the delta-globin coding sequences, the delta beta intergenic region as well as the beta-globin gene promoter and causes delta(0)beta(+) thalassemia with hemoglobin A expressed at the 11% to 15% range. The phenotype of this naturally occurring delta beta hybrid gene not only clarifies, in an in vivo context, the respective strength of delta- and beta-globin gene promoters, but also emphasizes the importance of beta-globin intragenic sequences in the expression of beta-globin chains. (Blood. 2001;98:1261-1263)     

Laotian (delta beta) degree-thalassemia: molecular characterization of a novel deletion associated with increased production of fetal hemoglobin

We have identified and molecularly characterized a novel deletion in the beta-globin gene cluster that increases fetal hemoglobin (HbF) synthesis in a 24-year-old Laotian man who is heterozygous for this mutation. The patient is asymptomatic with a mild anemia, hypochromia, and microcytosis (Ht = 39%, MCH = 22.8 pg, MCV = 71 fl), normal levels of HbA2 (3.0%) and 11.5% HbF (G gamma A gamma ratio 60 to 40), with heterocellular distribution (52% F cells). Extensive restriction endonuclease mapping defined the 5' breakpoint within the IVS II of the delta-globin gene, between positions 775 to 781 very similar to the 5' breakpoint of the Sicilian delta beta-thalassemia. However, the 3' breakpoint was localized between two Pst I sites 4.7 kb 3' of the beta- globin gene, thus ending about 0.7 kb upstream from the 3' breakpoint of the Sicilian delta beta-thalassemia. This results in a 12.5 kb deletion of DNA. It is of interest that the 5' breakpoint of the deletion residues within an AT-rich region which has been proposed as a specific recognition signal for recombination events, while the 3' breakpoint lies within a cluster of L1 repetitive sequences (formerly known as Kpn I family repeats). The presence of the 3' breakpoints of several other deletions within this region of L1 repeats also suggests that such sequences might serve as hot spots for recombination and eventually lead to thalassemia deletions. The similarity of the 5' and 3' breakpoints of these delta beta-thalassemias underscores the putative regulatory role of the deleted and juxtaposed sequences on the expression of the gamma-globin genes in adult life.     

A novel (g)gamma(a)gamma(deltabeta)O-thalassemia with a 27 kb deletion

A new (G)gamma(A)gamma(deltabeta)O-thalassemia (thal) was found in six unrelated Japanese individuals, and characterized by a method employing only polymerase chain reaction (PCR) and direct sequencing. This (G)gamma(A)gamma(deltabeta)O-thal mutation has removed a fragment of about 27 kb of DNA, that starts approximately 2.8 kb downstream of the Agamma-globin gene and ends in the L1 repeat sequence, 7.0 kb downstream of the beta-globin gene. The 5' breakpoint is similar to that of the previously reported Japanese (G)gamma(A)gamma(deltabeta)O-thal (called here Jpn type 1 for convenience). However, the 3' endpoint is quite different. This new Japanese deltabeta-thal, designated as Japanese type 2 (Jpn type 2), shows a deletion rather similar to Turkish type 3 deltabeta-thal but with 5' and 3' breakpoints located inside the deletion of Turkish type 3. A mutation-specific gap PCR was designed to diagnose patients with the Jpn type 2 (G)gamma(A)gamma(deltabeta)O-thal. The identified carriers exhibited a thalassemia minor.     

A novel basis for delta beta-thalassemia in a Chinese family

We have studied a Chinese family in which beta-thalassemia and delta beta-thalassemia were found in simple and compound heterozygous states. The delta beta-thalassemia heterozygote (the mother) had 22.3% hemoglobin F, of which 40% was G gamma and 60% A gamma; globin chain studies showed an alpha/beta + gamma ratio of 1.36. The compound heterozygote for delta beta-thalassemia and beta-thalassemia (the child) had the clinical picture of thalassemia intermedia and an alpha/beta + gamma ratio of 4.44. Gene mapping studies were performed using DNA from the affected child. Seventy kilobases of DNA in the beta- globin gene cluster starting upstream from the epsilon-globin gene and ending downstream from the beta-globin gene were mapped, and no detectable deletions or rearrangements were detected. In addition, heterozygosity was detected at multiple polymorphic restriction sites in and 3' to the beta-globin gene, which excludes the possibility of a deletion of the entire beta-globin gene cluster. This is the first example of a nondeletion delta beta-thalassemia associated with increased expression of both G gamma and A gamma genes.     

A novel 105 basepair deletion causing beta(0)-thalassemia in members of a Thai family.

We identified and characterized a novel beta(0)-thalassemia mutation due to partial deletion of the 5' end beta-globin gene including the mRNA cap site and a part of exon 1. The deletion was precisely 105 basepair (bp) in length extending from position -24 or -25 to +80 or +81 relative to the beta-globin gene mRNA cap site. This mutation was detected in three individuals from a family originating in the area of southern Thailand. The propositus was a 39-year-old female and noted to be heterozygous for beta-thalassemia with hemoglobin (Hb) level of 10.1 g/dl, MCV 70 fl, MCH 23.1 pg, HbA2 6.3%, and HbF 2.4%. Her son was 9 years of age and was also heterozygous for the mutation, having Hb level of 10.8 g/dl, MCV 58 fl, MCH 19.0 pg, HbA2 5.6%, and HbF 4.3%. Her 6-year-old daughter was affected, having a genotype of this mutation and a G-C transition at IVS 1 nt 5. Although the deletion does not include the beta-globin gene promoter sequences, the individuals heterozygous for this mutation have an elevated HbA2 level slightly higher than observed in most carriers of beta-thalassemia caused by point mutations.     

Different molecular defects of G gamma (A gamma delta beta)o-thalassaemia in Thailand

DNA from members of 2 Thai families with conditions considered to be delta beta-thalassaemia were studied by using restriction endonuclease DNA mapping. The propositus in family A is a double heterozygote for beta-thalassaemia and delta beta-thalassaemia. DNA analysis reveals a deletion of the beta-globin gene cluster starting at the area between the Sac I and Eco RI sites near the 3' end of the G gamma-gene and extending through the A gamma-, delta- and beta-genes to an unknown extent downstream. In family B, the propositus is delta beta-thalassaemia/Hb E. Deletion of the beta-globin gene cluster begins in the large intervening sequence of the A gamma-gene and removes both delta- and beta-genes downstream.     

Homozygosity for a new type of G gamma (A gamma delta beta)zero-thalassemia in a Malaysian male

Hematological and clinical data are presented for a young Malay patient with a homozygous (delta beta)zero-thalassemic condition. His red blood cells contained 100% fetal hemoglobin with alpha and G gamma chains only. Detailed gene mapping defined a large deletion with a 5' end between the Aha III and Apa I sites, some 200-400 bp 5' to the A gamma globin gene and a 3' end beyond sequences 17-18 kb 3' to the beta globin gene. This G gamma (A gamma delta beta)zero-type of thalassemia is different from all the other six types described before. Comparison of the hematological data of this patient with those of homozygotes for either the Sicilian or Spanish types of G gamma A gamma (delta beta)zero-thalassemia showed no differences; all homozygotes have a moderate anemia which is accentuated by the relatively high oxygen affinity of the Hb F containing erythrocytes.     

Thai G gamma (A gamma delta beta)zero-thalassemia and its interaction with a single gamma-globin gene on a chromosome carrying beta zero-thalassemia

Clinical manifestations and hematologic data of thalassemia intermedia were observed in three siblings of a Thai family. Analyses of the hemoglobin of their parents and other siblings indicated that they inherited a delta beta-thalassemia gene from the father and a beta zero thalassemia gene from the mother. Globin gene mapping confirmed that they carry two abnormal beta-globin gene complexes. On one chromosome more than 70 kb of DNA was removed which resulted in G gamma (A gamma delta beta)zero-thalassemia. The deletion started at the Hind III site located just 3' to the G gamma gene, and extended downstream to a region recognized by the p3'N 2.8R probe which is located more than 45 kb from the 3' end of the beta gene. The other chromosome carried a beta zero thalassemia gene, and a 5 kb deletion between the G gamma and A gamma genes which produced a hybrid -GA gamma- gene. A synthetic oligonucleotide probe showed that this beta zero thalassemia arose from a C----T mutation at position 654 of IVS-II in the beta-globin gene.     

Hemoglobin San Diego/beta zero thalassemia in a Greek adult

Interaction of beta zero thalassemia with Hb San Diego [a high affinity hemoglobin variant, alpha 2 beta (2)109(G11)Val----Met] in a 29-year-old Greek male is described. A marked polycythemia with low MCH and MCV, but minor clinical problems were observed. Functional properties of the isolated variant are described.     

Interaction of two different disorders in the beta-globin gene cluster associated with an increased hemoglobin F production: a novel deletion type of (G) gamma + ((A) gamma delta beta)(0)-thalassemia and a delta(0)-hereditary persistence of fetal hemoglobin determinant.

We report two different disorders of the beta-globin gene cluster segregating in a Belgian family: a novel deletion that results in (G) gamma + ((A) gamma delta beta)(0)-thalassemia (thal) and a heterocellular hereditary persistence of foetal hemoglobin of the Swiss type linked to a delta(0)-thal gene (delta (0)-HPFH). Heterozygosity for the heterocellular HPFH brings about a moderate (3.4% to 8.24%) increase of hemoglobin (Hb) F having a G gamma/A gamma ratio of 4:1, whereas carriers of the G gamma + ((A) gamma delta beta)(0)-thal deletion show in their peripheral blood a considerably higher (15%) percentage of Hb F. Both defects interact in the compound heterozygotes for G gamma + ((A) gamma delta beta)(0)-thal and delta(0)-HPFH producing a further increase (up to 24%) of fetal Hb consisting entirely of G gamma chains. Molecular characterization of the (G) gamma + ((A) gamma delta beta)(0)-thal by means of Southern analysis showed that the deletion spans about 50 kb, removing the 3' end of the A gamma-gene, the psi beta-, delta-, and beta-genes. A number of possible mechanisms leading to the overproduction of Hb F in HPFH and (G) gamma + ((A) gamma delta beta)(0)-thal will be discussed.