Study of alpha-thalassemia in northeastern Thailand at the DNA level

The frequency of alpha-thalassemias in a general population sample from northeastern Thailand and in an Austroasiatic group with high frequencies of hemoglobin E and beta-thalassemia, the So, was estimated using DNA techniques. Among 64 healthy adult subjects from the Khonkaen and Ubol areas, the following haplotype frequencies were determined: alpha alpha, 0.742; -alpha 3.7 (subtype I), 0.148; -alpha 4.2, 0.016; -alpha del, 0.008; alpha Constant Spring alpha, 0.055; --SEA, 0.023, and alpha alpha alpha (triplicated alpha-globin gene), 0.008. In the So group, the combined frequency of alpha-thalassemia chromosomes was 0.525.     

Abnormal arrangements in the alpha- and gamma-globin gene clusters in a relatively large group of Japanese newborns.

Data were obtained on blood samples from a relatively large group (264) of healthy Japanese newborns, collected at hospitals in Tokyo, Kurashiki, and Ube. The studies included an evaluation of anomalies in alpha-globin gene and gamma-globin gene arrangements using gene mapping and gamma-chain composition analyses. The results confirmed the rarity of alpha-thalassemia among Japanese; only a few babies had alpha-thalassemia-2 trait (the -3.7-kilobase [kb] deletion), while others had alpha-globin gene triplications (both the alpha alpha alpha anti-3.7 and the alpha alpha alpha anti-4.2 types). Among the gamma-globin gene anomalies that were observed, a few babies had the -A gamma-A gamma- globin gene arrangement or the -G gamma A gamma- type of deletion. The gamma-chain triplication (-G gamma-A gamma G gamma-A gamma-) occurred in 10 out of 256 newborns, and its frequency exceeded that of its corresponding -G gamma A gamma- deletion by a factor of 5. The restriction endonuclease XmnI was a useful tool, in addition to the enzymes Bg1II and BclI, to evaluate and confirm the gamma-globin gene deletion and triplication. The A gamma T variant, which is the product of a mutant A gamma-globin gene, occurred at a frequency of 0.156. The chromosome carrying this mutant A gamma gene had a characteristic haplotype that was originally seen in black and Mediterranean patients with Hemoglobin (Hb) S or with beta-thalassemia.     

Deletional types of alpha-thalassaemia in central Java.

The frequency of deletional alpha-thalassaemia in a Javanese population sample (n = 103) was investigated at three restriction sites of the alpha-globin gene (BamHI, BglII and RsaI). The overall gene frequency of alpha+ deletional thalassaemia was found to be very low (0.03). Leftward (-alpha 4.2) and rightward (-alpha 3.7) deletions and triplicated genes were present in equal frequency (0.015 and 0.005, respectively).     

The mammalian alphaD-globin gene lineage and a new model for the molecular evolution of alpha-globin gene clusters at the stem of the mammalian radiation

We have explored the evolution of the alpha-globin gene family by comparative sequence and phylogenetic analyses of mammalian alpha-globin genes. Our analyses reveal the existence of a new alpha-globin gene lineage in mammals that is related to the alpha(D)-globin genes of birds, squamates and turtles. The gene is located in the middle of the alpha-globin gene cluster of a marsupial, Sminthopsis macroura and of humans. It exists in a wide variety of additional mammals, including pigs, cows, cats, and dogs, but is a pseudogene in American marsupials. Evolutionary analyses suggest that the gene has generally evolved under purifying selection, indicative of a functional gene. The presence of mRNA products in humans, pigs, and cows also suggest that the gene is expressed and likely to be functional. The analyses support the hypothesis that the alpha(D)-globin gene lineage has an ancient evolutionary origin that predates the divergence of amniotes. The structural similarity of alpha-globin gene clusters of marsupials and humans suggest that an eight gene cluster (5'-zeta2-zeta1-alpha(D)-alpha3-alpha2-alpha1-theta-omega-3'), including seven alpha-like genes and one beta-like globin gene (omega-globin) existed in the common ancestor of all marsupial and eutherian mammals. This basic structure has remained relatively stable in marsupials and in the lineage leading to humans, although omega-globin has been lost from the alpha-globin gene cluster of humans.     

Alpha-thalassemia in blacks is due to gene deletion.

We used molecular hybridization to test if alpha-thalassemia is due to gene deletion in the black. In 10 families with clinically well-defined alpha-thalassemia-1 (alpha-thal-1), hydribization of alpha-globin cDNA was reduced to the same level as that found in Asians with alpha-thal-1, where two of the four normally present alpha-globin genes are deleted. A black child with hemoglobin H (Hb H) disease also has three globin genes deleted, as do Asian patients with Hb H disease. We conclude that alpha-thalassemia in the black is most commonly due to gene deletion.     

中国人α珠蛋白基因-α3.7缺失亚型的研究

-α3.7是中国人常见的缺失型α-地中海贫血-2。根据重组位点的不同,-α3.7可分为-α3.7Ⅰ型、-α3.7Ⅱ型和-α3.7Ⅲ型,并且亚型的种类和频率具有种族差异性。本研究在中国人群中用PCR基因分析方法检出具有α珠蛋白基因-α3.7缺失的患者56例,然后用ApalⅠ和BalⅠ限制性内切酶进行分型。 结果表明,在这56例具有-α3.7缺失的患者中,有54例是-α3.7Ⅰ型,有2例是-α3.7Ⅱ型,尚未发现-α3.7Ⅲ型。此结果丰富了我国α地贫基因型谱的资料。 Abstract:-α3.7 is a common deletional α-thalassemia-2 in China.According to different recombination sites,-α3.7 can be divided into -α3.7Ⅰ、-α3.7Ⅱand -α3.7Ⅲ.The frequency and population distribution of these -α3.7 are quite different.In this study,we detected 56 patients among Chinese population of -α3.7 defect in alpha globin gene by PCR method,then the PCR product was digested by the restriction enzyme ApalⅠand BalⅠ.The sub-typing result shows that in the 56 cases of -α3.7 defect,54 out of 56 is -α3.7Ⅰ,2 out of 56 is -α3.7Ⅱ and none of -α3.7Ⅲ is detected.This result enriches the data about the alpha thalassemia genotypes of Chinese people.     

Two types of triplicated alpha-globin loci in humans.

DNA from healthy Malaysian newborns was studied on gene maps after digestion with different restriction endonucleases. Of 65 newborns, two were found to be carriers of two different variants of triplicated alpha-globin loci. In variant no. 1, found in an Malay, the three alpha-globin genes are in an elongated DNA fragment on digestion with Eco RI and Bam HI. The third alpha-globin gene was found in a additional 3.7-kb fragment on digestion with Hpa I, Bgl II and Hind III. In variant no. 2, a new type of triplicated alpha-globin loci, found in a Chinese, the three alpha-globin genes reside in an elongated DNA fragment longer than that of variant no. 1 on digestion with Eco RI and Bam HI. The third alpha-globin gene was found in an additional 4.2-kb fragment on digestion with Hpa I and Hind III. Digestion of this variant DNA with Bg1 II produced an abnormal 16.7-kb fragment in addition to the normal 7.0-kb Bgl-II fragment. The locations of the restriction sites in the two types of triplicated alpha-globin loci are compatible with a mechanism of unequal crossing over following two different modes of misalignment.     

Multiple alpha-globin genes in crab-eating macaques (Macaca fascicularis)

Alpha-globin genes in crab-eating macaques were found to be triplicated at high frequencies according to restriction-enzyme comparisons. The frequencies of triplicated alpha-globin genes in macaques originally from Malaysia and Indonesia were 0.432 and 0.275, respectively, while no triplication was found in individuals from either the Philippines or northern and central Thailand. Quadruplicated alpha-globin genes were also observed, at frequencies of 0.045 (Malaysia), 0.075 (Indonesia), and 0.021 (the Philippines). A single locus was detected in only one of 40 chromosomes from Indonesia (frequency 0.025).     

Beta-globin gene haplotypes and alpha-thalassemia analysis in Babinga pygmies from Congo-Brazzaville

We analyzed beta-globin gene cluster haplotypes and deletional alpha+-thalassemia (-alpha3.7kb) in 54 Babinga pygmy subjects from Congo-Brazzaville. The beta(S)-globin gene frequency was 0.065 and that of the deletional alpha-globin gene (-alpha3.7kb) was 0.29. Eighty-five percent of the beta(S) chromosomes and 13% of the beta(A) chromosomes were associated with the Bantu haplotype, 10% of beta(A) chromosomes with the Senegal haplotype, and the remaining beta chromosomes with atypical haplotypes. None of the chromosomes were of the Benin haplotype. These results are clearly of anthropological and evolutionary interest. They also support earlier observations that alpha+-thalassemia is prevalent at a high frequency in African populations.     

Molecular basis for HbH disease in Italy: geographical distribution of deletional and nondeletional alpha-thalassemia haplotypes.

We have investigated the molecular basis for HbH disease in 16 patients from Sardinia, and central and southern Italy. We have shown that HbH disease is produced by the interaction of at least 10 different deletional or nondeletional alpha-thalassemia haplotypes, some of which have been already described in the Mediterranean area (--Med,-(alpha)20.5,-alpha 3.7 type I,-alpha 3.7 type II, alpha 2 NcoI alpha 1, alpha 2 HphI alpha 1). Among the new mutations found in the course of our study, there is a complete deletion of the zeta-alpha cluster and three nondeletional determinants (alpha alpha T), affecting to various extents alpha-globin gene expression. The different alpha-thalassemia haplotypes are not evenly distributed throughout the country. Two alpha 0 determinants [-(alpha)20.5 and the complete deletion of the zeta-alpha cluster] and four alpha + determinants (-alpha 3.7 type II, three nondeletional alpha alpha T mutations) are found exclusively in southern Italy.     

Alpha-globin gene markers identify genetic differences between Australian aborigines and Melanesians.

Australian aborigines exhibit a number of alpha-globin cluster rearrangements involving both alpha- and zeta-globin genes. alpha+-Thalassemia (-alpha/) in this population is heterogeneous and includes the 3.7 types I, II, and III gene deletions. The alpha alpha alpha/ and zeta zeta zeta/ rearrangements are each found in association with two haplotypes, indicating origins from at least two separate DNA crossover events. Differences in alpha-globin cluster rearrangements and in haplotypes between Australian aborigines, Papua New Guinea highlanders and island Melanesians, are consistent with multiple colonizing events into Australia.     

Structural and evolutionary analysis of the two chimpanzee alpha-globin mRNAs.

Two distinct alpha-globin mRNAs were detected in chimpanzee reticulocyte mRNA using a primer extension assay. DNA copies of these two mRNAs were cloned in the bacterial plasmid pBR322, and their sequence was determined. The two alpha-globin mRNAs have obvious structural homology to the two human alpha-globin mRNAs, alpha 1 and alpha 2. Comparison of the two chimpanzee alpha-globin mRNAs to each other and to their corresponding human counterparts revealed evidence of a recent gene conversion in the human alpha-globin complex and a marked heterogeneity in the rate of structural divergence within the alpha-globin gene.     

Molecular analysis of hemoglobin H disease in Taiwan

The molecular basis of seven Chinese patients in Taiwan with hemoglobin H disease was investigated and was found to be heterogeneous in the mutation type. They were alpha-thalassemia-1 mutation combined with hemoglobin Constant Spring, an undetermined nondeletion form of alpha-thalassemia and a deletion form of alpha-thalassemia-2 mutations. The alpha-thalassemia-1 mutation was shown to be the --SEA type I haplotype.     

Triple alpha-genes (alpha alpha alpha anti3.7) in a patient with sickle cell anaemia.

This paper reports the case of a 17-year-old male student from the Jaizan area in south-western Saudi Arabia who had sickle cell anaemia and possessed three alpha-genes on one chromosome (alpha alpha alpha anti3.7) and two on the other. The clinical manifestations were severe, with frequent blood transfusion requirements and frequent episodes of painful crises, severe anaemia and tissue involvement. In comparison with age and sex-matched sickle cell anaemia patients with one alpha-gene deletion (-alpha/alpha alpha), or a normal alpha-gene arrangement (alpha alpha/alpha alpha), a more severe disease presentation was obvious in the propositus. It is suggested that with the surplus alpha-globin chains, more severe haematological and clinical abnormalities occur, these influence the phenotypic expression of sickle cell anaemia. However, more patients with this type of gene arrangement must be studied before a definite conclusion can be reached regarding the influence of excess alpha-globin chains on the presentation of sickle cell anaemia.     

Recombinational resolution in primate cells of two homologous human DNA segments with a gradient of sequence divergence.

Human alpha-thalassemia-2 genotype -alpha 4.2 is the result of meiotic recombination between two 1.3 kb long, homologous DNA segments, X(alpha 2) and X(alpha 1), located in the adult alpha globin locus. The two segments can also undergo intramolecular recombination on extrachromosomal vectors transfected into mitotically dividing primate cells (COS 7). The existence of a gradient of sequence divergence between X(alpha 2) and X(alpha 1) makes them an interesting system to study the relationship between efficiencies of homologous DNA recombination and the extent of dispersed and localized base mismatches. By partial restriction mapping and DNA sequencing of plasmids recombined in COS 7 cells and rescued from bacteria HB 101, we have determined the distribution of recombinational resolution sites along the two X blocks. Most, if not all, of the homologous recombination events between the two X blocks appear to be single crossing-over without efficient gene correction or repair of base mismatches. The distribution of the sites of recombinational resolution is inversely correlated with that of the gradient of sequence divergence, with only approximately 7% of the X recombinants resolved within the 3' third of the X blocks where two diverged Alu family repeats reside. The Alu sequence within which one of the X recombinants resolved is homologous to a previously characterized alpha thalassemia deletion point.