The evolution of the α- and β-globin gene clusters in human populations

Summary DNA analysis of the - and -globin gene clusters has revealed substantial variability between individuals and populations. As well as restriction enzyme site and length polymorphisms, variation in gene copy number and type is observed. Because of this extensive polymorphism DNA analysis offers a highly informative method of studying genetic affinities between human populations. Haplotypes, consisting of a set of restriction enzyme polymorphisms distributed along the cluster, have been developed for both loci. Analysis of the molecular basis of numerous -thalassaemia alleles has revealed, in general, different sets of mutations in different populations, indicating that these postdate the racial divergence. Recent microepidemiological studies on the distribution of -thalassaemia support the hypothesis that this condition, like the {ie16-1}, has been selected because it confers protection against malaria. Population-specific DNA polymorphisms at these and other loci promise to be of considerable value to genetic anthropology.     

Knockdown of mouse adult β-globin gene expression in MEL cells by retrovirus vector-mediated RNA interference

RNA interference (RNAi) efficiently induces sequence-specific gene silencing in mammalian cells through short interfering RNA (siRNA) of 21–23 nucleotides synthesized in vitro or expressed by DNA-based vector. However, introduction of siRNA into mammalian cells by transfection limits the application of RNAi, especially when it is necessary to generate long-term gene silencing in vivo. Virus vector-mediated RNAi provides an alternative to transfection. In the present study, we investigated such transduction system and showed that retrovirus vector-mediated RNAi can substantially down-regulate expression of mouse adult β-globin gene in MEL cells. The results suggest that retrovirus vector-delivered RNAi may find its use in functional genomics and in gene therapy.     

Sequence analysis of the upstream regions of Xenopus laevis β-globin genes and arrangement of repetitive elements within the globin gene clusters

The globin gene clusters of Xenopus laevis are interspersed by various different repetitive DNA elements. A specific repeat, the JH12 element, has been mapped by Southern analysis and some of its locations have been subsequently confirmed by nucleotide sequencing. JH12 family members seem to represent mobile genetic elements and display a high degree of divergence. The nucleotide sequences upstream to the adult _I-globin gene and to the two coordinately expressed larval _I- and _II genes have been determined and compared to those of the adult -genes. Besides some repetitive DNA elements and a short sequence of rather weak homology we have found no characteristic sequence motifs to be common to the adult - and -genes. The two larval -genes share one short sequence element being absent from the adult genes. This might reflect completely different sequence requirements for protein interactions and for the regulation of adult and larval globin gene expression.     

Equilibrium frequencies of α-globin genes

A mathematical model has been developed to study the mechanism of the maintenance of variable numbers of α-globin genes in human populations. The model incorporates both selection and unequal crossing-over. The selection is formulated so that a homozygous individual with a double deletion is lethal and a heterozygous individual with a deletion or addition of an α-globin gene in a chromosome has decreased fitness. This differs from the previous models of stabilizing selection studied by Ohta (1981) and Takahata (1981). The effect of random genetic drift on the α-thalassemia polymorphims has also been studied.It has been shown that, although the results obtained are compatible with the observation of the low frequency of triple α-globin loci, it cannot explain the high frequency of single and double deletions in Asian populations. For the latter case, some type of heterozygote advantage may be operating.     

Primary structure and evolutionary relationship between the adult α-globin genes and their 5′-flanking regions ofXenopus laevis andXenopus tropicalis

Summary To investigate the evolution of globin genes in the genusXenopus, we have determined the primary structure of the related adult ₁- and _II genes ofX. laevis and of the adult -globin gene ofX. tropicalis, including their 5-flanking regions. All three genes are comprised of three exons and two introns at homologous positions. The exons are highly conserved and code for 141 amino acids. By contrast, the corresponding introns vary in length and show considerable divergence. Comparison of 900 bp of the 5-flanking region revealed that theX. tropicalis gene contains a conserved proximal 310-bp promoter sequence, comprised of the canonical TATA and CCAAT motifs at homologous positions, and five conserved elements in the same order and at similar positions as previously shown for the corresponding genes ofX. laevis. We therefore conclude that these conserved upstream elements may represent regulatory sequences for cell-specific regulation of the adultXenopus globin genes.     

ααααanti-3.7 type II: a new α-globin gene rearrangement suggesting that the α-globin gene duplication could be caused by intrachromosomal recombination

Summary We report here a new human -globin gene rearrangement carrying the two normal, 2 and 1, and two hybrid, 1/2, globin genes in the order 5-2-1/2-1/2-1-3. Both the hybrid genes, subtyped with ApaI and RsaI restriction enzymes, were found to be of the uncommon anti 3.7 type II. The hybrid genes were expressed at the biosynthetic level and their interaction with the -thalassaemia IVS 1 nt 1 GA mutation caused thalassaemia intermedia. We also report a case of an -globin gene rearrangement in the twin of one of the -globin gene carriers; the duplicated gene was of the anti 4.2 type and was associated with the absence of RsaI polymorphism. The singular finding of an -anti 3.7 cluster with two identical rare hybrid genes suggests that the reciprocal unequal recombination causing the -globin gene rearrangements could be of the intra-chromosomal rather than the interchromosomal type.     

Evolution of the β-globin gene cluster in man and the primates

The arrangement of primate β-related globin genes has been determined by restriction endonuclease mapping of genomic DNA from species ranging from prosimians to man. The arrangement of the entire ?γγδβ-globin gene cluster in the gorilla and the yellow baboon is indistinguishable from that of man. Restriction site differences between these species are consistent with a surprisingly low overall rate of intergenic DNA sequence divergence of approximately 1% in 5 million years. A new world monkey (owl monkey) has a single γ-globin gene, suggesting that the ^Gγ-^Aγ-globin gene duplication in man is ancient, and occurred about 20 to 40 million years ago. The β-globin gene cluster in the brown lemur, a prosimian, is remarkably short (about 20,000 base-pairs) and contains single ?-, γ- and β-globin genes. The γ- and β-globin genes in this animal are separated by a curious gene containing the 3′ end of a β-globin gene preceded by sequences related to the 5′ end of the ?-globin gene.     

DNA polymorphisms in the ψβ1- and β-globin gene regions in asian macaques

DNA polymorphisms in the ₁--globin gene region in nine Asian macaques(Macaca fuscata, M. mulatta, M. nemestrina, M. cyclopis, M. fascicularis, M. arctoides, M. radiata, M. maura, andM. assamensis) were examined using several restriction endonucleases and the human ₁, IVS2, and IVS2 probes. TheBamHI site 3 to the -globin gene was polymorphic inM. fuscata andM. mulatta, while the HincII site and the EcoRI site in the ₁-globin gene region was highly polymorphic inM. fuscata andM. mulatta, respectively. These polymorphic sites also seem to be present in other Asian macaques. The present study of the polymorphism at theBamHI site 3 to the -globin gene in Asian macaques supports, at the nuclear DNA level, the idea that thefascicularis group includingM. fuscata, M. mulatta, M. cyclopis, andM. fascicularis is different from other Asian macaque groups.This study was supported in part by the Cooperation Research Program of the Primate Research Institute, Kyoto University.     

Heterogeneity of the α-globin gene defects in German α-thalassemia affected families

Summary Analysis of -thalassemia syndromes in several German families revealed DNA deletion as well as nondeletion forms as the molecular basis for the defects. Thus, the -thalassemia haplotype was identified as the (–)^3.7 rightward deletion form, and the region of the putative recombination process generating such a deletion was further characterized. In addition three different ° haplotypes, (--)^MED, (--)^>26, and ()^T, could be detected using -and -globin gene-specific probes.     

Altitudinal variation at duplicated β-globin genes in deer mice: effects of selection, recombination, and gene conversion

Spatially varying selection on a given polymorphism is expected to produce a localized peak in the between-population component of nucleotide diversity, and theory suggests that the chromosomal extent of elevated differentiation may be enhanced in cases where tandemly linked genes contribute to fitness variation. An intriguing example is provided by the tandemly duplicated β-globin genes of deer mice (Peromyscus maniculatus), which contribute to adaptive differentiation in blood-oxygen affinity between high- and low-altitude populations. Remarkably, the two β-globin genes segregate the same pair of functionally distinct alleles due to a history of interparalog gene conversion and alleles of the same functional type are in perfect coupling-phase linkage disequilibrium (LD). Here we report a multilocus analysis of nucleotide polymorphism and LD in highland and lowland mice with different genetic backgrounds at the β-globin genes. The analysis of haplotype structure revealed a paradoxical pattern whereby perfect LD between the two β-globin paralogs (which are separated by 16.2 kb) is maintained in spite of the fact that LD within both paralogs decays to background levels over physical distances of less than 1 kb. The survey of nucleotide polymorphism revealed that elevated levels of altitudinal differentiation at each of the β-globin genes drop away quite rapidly in the external flanking regions (upstream of the 5' paralog and downstream of the 3' paralog), but the level of differentiation remains unexpectedly high across the intergenic region. Observed patterns of diversity and haplotype structure are difficult to reconcile with expectations of a two-locus selection model with multiplicative fitness.     

Locus assignment of human α-globin structural mutants by selective enzymatic amplification of α1 and α2-globin cDNAs

Summary We have used the powerful methodology of DNA enzymatic amplification in order to assign human -globin structural mutants to one of the two highly homologous -globin genes. Selectively amplified 1 and 2-globin cDNAs were dot-blotted and further hybridized to synthetic oligonucleotides encompassing either the normal or the mutated sequences. The generated signals corresponded specifically to one of the two -globin genes. Using this approach the -globin structural mutants J-Buda and G-Pest were found to be encoded by the 2 and the 1-globin genes, respectively. Furthermore, the exact nucleotide changes were determined. We propose this technique to serve as a simple and definitive method for assigning -globin structural mutants.     

Linkage mapping of α-2 adrenergic receptor genes to mouse Chromosomes 2 and 5

-2 adrenergic receptors can be subdivided into three related subtypes which are conserved in humans, rats, and mice. In the mouse, these receptors are encoded by three genes (Adra-2a, Adra-2b, Adra-2c). To gain insight into the evolution of this multigene family and to investigate whether these genes are candidates for previously identified mouse mutations, we have determined the map positions of the Adra-2b and Adra-2c genes. The Adra-2a gene has been previously mapped to mouse Chromosome (Chr) 19 (Oakey et al. Genomics 10, 338–334, 1991). Using segregation among recombinant inbred strains of a single-stranded conformational polymorphism specific for alleles of Adra-2b and Adra-2c, we present map positions for these genes on mouse Chrs 2 and 5, respectively. In the case of Adra-2b, these results have been confirmed by an analysis of somatic cell hybrids. In addition, we generate AKXD recombinant inbred strain distribution patterns for 11 previously defined SSLP microsatellite markers, further refining the haplotype maps for these chromosomes. Finally, several candidate mouse mutations that map close to Adra-2b and Adra-2c are discussed.