Genetic Mosaicism and Hemoglobinopathies; Uniparental Isodisomy

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 2258-2258
Author(s):  
Cornelis L Harteveld ◽  
Marion Phylipsen ◽  
Piero C Giordano
Keyword(s):  
Clonal Selection ◽  
Direct Sequencing ◽  
Globin Gene ◽  
Beta Thalassemia ◽  
Beta Globin ◽  
Chromosome 11 ◽  
Hematopoietic Stem ◽  
Beta Globin Gene ◽  
Progressive Development ◽  
Uniparental Isodisomy

Abstract Background Recently we discovered three independent cases of “severe late onset beta-thalassemia”, all presenting with the mild phenotype of beta thalassemia minor up to adult age and developing a severe transfusion dependent phenotype in the third and fourth decade of life when a presumed homozygosity for the beta-thalassemia mutation is observed. We demonstrate that uniparental isodisomy (UPD) of part of chromosome 11p, a mechanism also seen in 20% of Beckwith-Wiedemann syndrome patients, accounts for the observed mosaicism in all three independent cases. Clonal selection for hematopoietic stem cells containing the UPD for the mutant beta-globin gene during life may account for the progressive development of the disease. Recently we discovered another case showing a similar mosaic UPD of 11p, presenting as a regular beta-thalassemia carrier. Method Direct sequencing of the beta-globin genes. Multiplex Ligation dependent Probe Amplification (MLPA) analysis of the HBB (beta-globin) gene cluster. Affymetrix GeneChip Human Mapping 262K NspI array (Santa Clara, CA, USA). Illumina OmniExpress 730K SNP Beads array (San Diego, CA, USA). Results In all cases molecular analysis shows sequences in which a near to complete homozygosity for the beta-gene mutation in DNA extracted from leucocytes is seen. Loss of heterozygosity due to allele drop-out or a deletion of one allele was excluded by direct sequencing and MLPA analysis. Affymetrix and/or Illumina SNP-array analysis revealed incomplete homozygosity for SNP’s along almost the entire short arm of chromosome 11 containing the beta-globin gene, indicating mosaicism for a partial uniparental isodisomy of chromosome 11p. Three patients were born asymptomatic as beta-thal carriers and developed a severe blood-transfusion dependent beta-thalassemia major at different ages and with different percentages of mosaicism. The fourth patient however did not develop the clinical severity despite of an almost 50% mosaicism determined from the DNA isolated from leucocytes. Conclusion We demonstrate that uniparental isodisomy of part of chromosome 11 accounts for the observed mosaicism in all four independent cases. The most probable mechanism seems clonal selection for hematopoietic stem cells containing the uniparental isodisomy for the mutant beta-globin gene during life, this may account for the progressive development of the disease. However, there seems to be no correlation between the percentage of mosaicism measured in the DNA isolated from the white cells and the severity of the clinical phenotype related to the expression in red cells, which strongly suggests hematopoietic tissue heterogeneity for the observed UPD containing cell lineages. This may have serious consequences for disease prediction and counseling, as this is largely dependent upon DNA isolated from leucocytes. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Rapid detection of deletions causing delta beta thalassemia and hereditary persistence of fetal hemoglobin by enzymatic amplification

Blood ◽  
1994 ◽  
Vol 83 (6) ◽  
pp. 1673-1682 ◽  
Author(s):  
JE Craig ◽  
RA Barnetson ◽  
J Prior ◽  
JL Raven ◽  
SL Thein
Keyword(s):  
Asian Indian ◽  
Globin Gene ◽  
Fetal Hemoglobin ◽  
Beta Thalassemia ◽  
Beta Globin ◽  
Chromosome 11 ◽  
Gamma Delta ◽  
Beta Globin Gene ◽  
Enzymatic Amplification ◽  
Hereditary Persistence

Abstract A considerable number of deletions of variable size and position that involve the beta-globin gene complex on chromosome 11 are associated with the clinical entities of hereditary persistence of fetal hemoglobin (HPFH) and delta beta thalassemia. Specific deletions appear to be associated with consistent phenotypes and some are known to be recurrent. To facilitate the molecular diagnosis of uncharacterized patients with HPFH and delta beta thalassemia, oligonucleotide primers have been designed to enzymatically amplify deletion-specific products for nine known deletions, which include those responsible for HPFH-1, HPFH-2, HPFH-3, Spanish (delta beta)zero thalassemia, hemoglobin (Hb) Lepore, Sicilian (delta beta)zero thalassemia, Chinese G gamma(A gamma delta beta)zero thalassemia, Asian-Indian inversion-deletion G gamma(A gamma delta beta)zero thalassemia, and Turkish inversion-deletion (delta beta)zero thalassemia. Using this approach, we have successfully characterized the molecular basis for delta beta thalassemia in 23 individuals from 16 families of diverse ethnic origins. Thirteen individuals from this group were shown to be heterozygous for the 13.4- kb Sicilian deletion, two were heterozygous for the 100-kb Chinese G gamma(A gamma delta beta)zero deletion, four were heterozygous for the Turkish form of inversion-deletion delta beta thalassemia, and three were heterozygous for the Asian-Indian form of inversion-deletion G gamma(A gamma delta beta)zero thalassemia. One Vietnamese subject was heterozygous for a 12.6-kb deletion, which we have fully characterized at the molecular level. Sequence analysis of the breakpoint regions of the Chinese deletion and the Turkish rearrangement indicates that, in each case, the mutation is likely to have arisen from a single origin. This hypothesis is supported by the evident geographical clustering of the various deletions described here.

scholarly journals Characterization of a spontaneous mutation in beta-thalassemia associated with advanced paternal age

Blood ◽  
1989 ◽  
Vol 74 (2) ◽  
pp. 852-854 ◽  
Author(s):  
FF Chehab ◽  
KH Winterhalter ◽  
YW Kan
Keyword(s):  
Direct Sequencing ◽  
Globin Gene ◽  
Spontaneous Mutation ◽  
Paternal Age ◽  
Beta Thalassemia ◽  
Molecular Defect ◽  
Beta Globin ◽  
Beta Globin Gene ◽  
Gene Restriction

Abstract We characterized the molecular defect in a Swiss patient with a spontaneous beta-thalassemia mutation. Cloning and DNA sequencing of her beta-globin gene revealed a new frameshift mutation due to a single nucleotide deletion at codon 64 of the beta-globin gene. Restriction site polymorphism showed that the mutation arose on her paternal chromosome. Direct sequencing of a polymerase chain reaction amplified DNA segment showed absence of the lesion in both alleles of her father's beta-globin gene and confirmed the spontaneous nature of this mutation.

scholarly journals A beta-thalassemia gene caused by a 290-base pair deletion: analysis by direct sequencing of enzymatically amplified DNA

Blood ◽  
1989 ◽  
Vol 73 (6) ◽  
pp. 1695-1698 ◽  
Author(s):  
R Spiegelberg ◽  
C Aulehla-Scholz ◽  
H Erlich ◽  
J Horst
Keyword(s):  
Direct Sequencing ◽  
Globin Gene ◽  
Beta Thalassemia ◽  
Inverted Repeats ◽  
Beta Globin ◽  
Base Pairs ◽  
Direct Dna Sequencing ◽  
Beta Globin Gene ◽  
Base Pair Deletion ◽  
Exon 1

Abstract The base composition around a recently detected deletion in the human beta-globin gene was determined by direct DNA sequencing of an enzymatically amplified DNA segment. The deletion removes 290 base pairs (bp), including the entire exon 1 and the mRNA cap site. In the vicinity of the deletion endpoints, the normal beta-globin gene contains direct and inverted repeats which may have taken part in generation of this deletion.

scholarly journals Rapid detection of deletions causing delta beta thalassemia and hereditary persistence of fetal hemoglobin by enzymatic amplification

Blood ◽  
1994 ◽  
Vol 83 (6) ◽  
pp. 1673-1682
Author(s):  
JE Craig ◽  
RA Barnetson ◽  
J Prior ◽  
JL Raven ◽  
SL Thein
Keyword(s):  
Asian Indian ◽  
Globin Gene ◽  
Fetal Hemoglobin ◽  
Beta Thalassemia ◽  
Beta Globin ◽  
Chromosome 11 ◽  
Gamma Delta ◽  
Beta Globin Gene ◽  
Enzymatic Amplification ◽  
Hereditary Persistence

A considerable number of deletions of variable size and position that involve the beta-globin gene complex on chromosome 11 are associated with the clinical entities of hereditary persistence of fetal hemoglobin (HPFH) and delta beta thalassemia. Specific deletions appear to be associated with consistent phenotypes and some are known to be recurrent. To facilitate the molecular diagnosis of uncharacterized patients with HPFH and delta beta thalassemia, oligonucleotide primers have been designed to enzymatically amplify deletion-specific products for nine known deletions, which include those responsible for HPFH-1, HPFH-2, HPFH-3, Spanish (delta beta)zero thalassemia, hemoglobin (Hb) Lepore, Sicilian (delta beta)zero thalassemia, Chinese G gamma(A gamma delta beta)zero thalassemia, Asian-Indian inversion-deletion G gamma(A gamma delta beta)zero thalassemia, and Turkish inversion-deletion (delta beta)zero thalassemia. Using this approach, we have successfully characterized the molecular basis for delta beta thalassemia in 23 individuals from 16 families of diverse ethnic origins. Thirteen individuals from this group were shown to be heterozygous for the 13.4- kb Sicilian deletion, two were heterozygous for the 100-kb Chinese G gamma(A gamma delta beta)zero deletion, four were heterozygous for the Turkish form of inversion-deletion delta beta thalassemia, and three were heterozygous for the Asian-Indian form of inversion-deletion G gamma(A gamma delta beta)zero thalassemia. One Vietnamese subject was heterozygous for a 12.6-kb deletion, which we have fully characterized at the molecular level. Sequence analysis of the breakpoint regions of the Chinese deletion and the Turkish rearrangement indicates that, in each case, the mutation is likely to have arisen from a single origin. This hypothesis is supported by the evident geographical clustering of the various deletions described here.

scholarly journals Characterization of a spontaneous mutation in beta-thalassemia associated with advanced paternal age

Blood ◽  
1989 ◽  
Vol 74 (2) ◽  
pp. 852-854
Author(s):  
FF Chehab ◽  
KH Winterhalter ◽  
YW Kan
Keyword(s):  
Direct Sequencing ◽  
Globin Gene ◽  
Spontaneous Mutation ◽  
Paternal Age ◽  
Beta Thalassemia ◽  
Molecular Defect ◽  
Beta Globin ◽  
Beta Globin Gene ◽  
Gene Restriction

We characterized the molecular defect in a Swiss patient with a spontaneous beta-thalassemia mutation. Cloning and DNA sequencing of her beta-globin gene revealed a new frameshift mutation due to a single nucleotide deletion at codon 64 of the beta-globin gene. Restriction site polymorphism showed that the mutation arose on her paternal chromosome. Direct sequencing of a polymerase chain reaction amplified DNA segment showed absence of the lesion in both alleles of her father's beta-globin gene and confirmed the spontaneous nature of this mutation.

scholarly journals A beta-thalassemia gene caused by a 290-base pair deletion: analysis by direct sequencing of enzymatically amplified DNA

Blood ◽  
1989 ◽  
Vol 73 (6) ◽  
pp. 1695-1698
Author(s):  
R Spiegelberg ◽  
C Aulehla-Scholz ◽  
H Erlich ◽  
J Horst
Keyword(s):  
Direct Sequencing ◽  
Globin Gene ◽  
Beta Thalassemia ◽  
Inverted Repeats ◽  
Beta Globin ◽  
Base Pairs ◽  
Direct Dna Sequencing ◽  
Beta Globin Gene ◽  
Base Pair Deletion ◽  
Exon 1

The base composition around a recently detected deletion in the human beta-globin gene was determined by direct DNA sequencing of an enzymatically amplified DNA segment. The deletion removes 290 base pairs (bp), including the entire exon 1 and the mRNA cap site. In the vicinity of the deletion endpoints, the normal beta-globin gene contains direct and inverted repeats which may have taken part in generation of this deletion.

scholarly journals The inactive beta globin gene on a gamma delta beta thalassemia chromosome has a normal structure and functions normally in vitro

Blood ◽  
1988 ◽  
Vol 71 (3) ◽  
pp. 766-770
Author(s):  
PT Curtin ◽  
YW Kan
Keyword(s):  
Globin Gene ◽  
Large Deletion ◽  
Beta Thalassemia ◽  
Ribonuclease Protection Assay ◽  
Beta Globin ◽  
Gamma Delta ◽  
Beta Globin Gene ◽  
Thalassemia Minor

We have previously described an English family with gamma delta beta- thalassemia in which a large deletion stops 25 kilobases (kb) upstream from the beta-globin gene locus, and yet the beta-globin gene is inactive in vivo. Affected family members had a beta-thalassemia minor phenotype with a normal hemoglobin A2 level. Gene mapping showed that these subjects were heterozygous for a chromosome bearing a large deletion that began in the G gamma-globin gene, extended through the epsilon-globin gene, and continued upstream for at least 75 kb. The A gamma-, delta-, and beta-globin gene loci on this chromosome were intact. To examine the possibility that an additional defect was present in the beta-globin gene, we cloned, sequenced, and examined the expression of the beta-globin gene from the affected chromosome. No mutation was found in the beta-globin gene sequence from 990 base-pairs 5′ to the cap site to 350 basepairs 3′ to the polyadenylation signal. The gene was subcloned into an expression vector and introduced into HeLa cells. Analysis of RNA derived from these cells, using a ribonuclease protection assay, revealed qualitatively and quantitatively normal transcription. Thus a structurally and functionally normal beta-globin gene is inactive in the presence of a large deletion more than 25 kb upstream. The loss of beta-globin gene function may be due to disturbance of chromatin conformation caused by the deletion or may be the result of loss of upstream sequences that are necessary for beta-globin gene expression in vivo.

scholarly journals Molecular analysis of Japanese delta beta-thalassemia

Blood ◽  
1988 ◽  
Vol 72 (5) ◽  
pp. 1771-1776
Author(s):  
S Shiokawa ◽  
H Yamada ◽  
Y Takihara ◽  
E Matsunaga ◽  
Y Ohba ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Globin Gene ◽  
Fetal Hemoglobin ◽  
Beta Thalassemia ◽  
Globin Genes ◽  
Beta Globin ◽  
Beta Globin Gene ◽  
Gamma Globin ◽  
Large Deletions ◽  
Deletion Junction

A DNA fragment containing the deletion junction region from a Japanese individual with homozygous delta beta-thalassemia has been cloned. A clone containing the normal DNA surrounding the 3′ breakpoint of this deletion and a clone carrying the G gamma- and A gamma-globin genes of this patient were also isolated. Sequences of the deletion junction and both gamma-globin genes were determined. A comparison of these sequences with previously determined sequences of the normal counterparts revealed that the 5′ breakpoint is located between 2,134 and 2,137 base pairs (bp) 3′ to the polyA site of the A gamma-globin gene, the 5′ breakpoint is located just downstream of the 3′ border of the fetal gamma-globin duplication unit, and no molecular defects are evident within the gamma-globin gene region. A comparison between the sequences of the normal DNA surrounding the 3′ breakpoint and the normal DNA surrounding the 5′ breakpoint shows that deletion is the result of a nonhomologous recombination event. There are A+T-rich stretches near the 5′ and 3′ breakpoints in the normal DNA, and a portion of an Aly repeat is located in the region 3′ to the 3′ breakpoint. Southern blot analysis using probes 3′ to the beta-globin gene showed that the deletion extends in the 3′ direction further than any other deletions associated with delta beta-thalassemia and hereditary persistence of fetal hemoglobin (HPFH) heretofore reported. These results are discussed in terms of the mechanism generating large deletions in mammalian cells and three models for the regulation of gamma-globin and beta-globin gene expression in humans.

scholarly journals Current Perspective of Beta Thalassemia and Its Treatment Strategies

2019 ◽  
Author(s):  
Shaukat Ali ◽  
Shumaila Mumtaz ◽  
Hafiz Abdullah Shakir ◽  
Hafiz Muhammad Tahir ◽  
Tafail Akbar Mughal
Keyword(s):  
Genetic Testing ◽  
Blood Transfusion ◽  
Dna Analysis ◽  
Complete Blood Count ◽  
Globin Gene ◽  
Thalassemia Major ◽  
Beta Thalassemia ◽  
Beta Globin ◽  
Hematopoietic Stem ◽  
Thalassemia Minor

Thalassemia is genetic blood disease cause by absence or decrease of one or more of the globin chain synthesis. Beta thalassemia is characterized by one or more mutations in beta globin gene. Absence or reduced amount the of beta globin chains cause ineffective erythropoiesis which leads to anemia. Beta thalassemia has been further divided into three main forms: Thalassemia minor/silent carrier, major and intermedia. More severe form is thalassemia major in which patients depend upon blood transfusion for survival and high level of iron occur as a consequence of consistent blood transfusion. Over loaded iron invokes the synthesis of reactive oxygen species that are toxic in redundancy and triggering the impairment to vascular, endocrine and hepatic system. Thalassemia can be diagnosed and detected through various laboratory tests such as blood smear, prenatal testing (genetic testing of amniotic fluid), DNA analysis (genetic testing) and complete blood count. Treatment of thalassemia intermedia is symptomatic but it can also be managed by splenectomy and folic supplementation. While thalassemia major can be treated by transplantation of bone marrow, regular transfusion of blood and iron chelation treatment, stimulation of fetal hemoglobin production, hematopoietic stem cell transplantation and gene therapy.

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