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Monophyletic units thus far identified - be it Y SNPs, mtDNA lineages, JC virus genes , or insulin haplotypes, and so forth, tell the same story…either showing dispersal patterns that hint on anatomically modern humans originating in the African continent, and/or that Africans sport the highest biological diversity, of which non-Africans are derived from a subset…
Global Haplotype Diversity in the Human Insulin Gene Region John D.H. Stead1,3,4, Matthew E. Hurles2 and Alec J. Jeffreys1
The insulin minisatellite (INS VNTR) has been intensively analyzed due to its associations with diseases including diabetes. We have previously used patterns of variant repeat distribution in the minisatellite to demonstrate that genetic diversity is unusually great in Africans compared to non-Africans. Here we analyzed variation at 56 single nucleotide polymorphisms (SNPs) flanking the minisatellite in individuals from six populations, and we show that over 40% of the total genetic variance near the minisatellite is due to differences between Africans and non-Africans, far higher than seen in most genomic regions and consistent with differential selection acting on the insulin gene region, most likely in the non-African ancestral population. Linkage disequilibrium was lower in African populations, with evidence of clustering of historical recombination events. Analysis of haplotypes from the relatively nonrecombining region around the minisatellite revealed a star-shaped phylogeny with lineages radiating from an ancestral African-specific haplotype. These haplotypes confirmed that minisatellite lineages defined by variant repeat distributions are monophyletic in origin. These analyses provide a framework for a cladistic approach to future disease association studies of the insulin region within both African and non-African populations, and they identify SNPs which can be rapidly analyzed as surrogate markers for minisatellite lineage.
The insulin minisatellite, located within the promoter of the human insulin gene, has been intensely investigated for nearly two decades due to its associations with diseases such as diabetes (Bell et al. 1984; Bennett and Todd 1996). Most studies have analyzed populations of European descent where low diversity at the minisatellite combined with strong linkage disequilibria in flanking regions make it difficult to distinguish between etiological and associated variants (Bennett and Todd 1996; Doria et al. 1996). The identification of etiological polymorphisms in the insulin region may therefore require analysis of a range of different populations, in particular those showing a greater range of haplotype diversity than that seen in Europeans.
As a first step in these studies, we used a system of minisatellite variant repeat mapping by PCR (MVR-PCR; Jeffreys et al. 1991) to analyze variation at the insulin minisatellite both in allele size (number of tandem repeats) and structure (distribution of different variant repeats) in a range of populations from Africa, Asia, and Europe (Stead and Jeffreys 2000, 2002). Pairwise comparisons between all alleles showed that structures were either very different from each other, indicating that they had diverged through multiple complex mutational rearrangements, or displayed very similar patterns of variant repeat distribution, resulting in these closely related structures having similar overall sizes. In this way, alleles could be readily divided into lineage groups with low levels of variation within each lineage both in allele size and structure (Stead and Jeffreys 2000, 2002). Analysis of three African populations from the Ivory Coast (156 alleles), Zimbabwe (138), and Kenya (84) revealed substantial structural diversity with minisatellite alleles assigned to one of 22 different lineages. In contrast, all alleles within non-African populations from the U.K. (1080 alleles), Japan (118), and Kazakhstan (80) belong to just three lineages. These lineages, termed I, IIIA, and IIIB, are all present in Africa. This reduction in diversity from 22 lineages in Africans to a subset of just three lineages in non-Africans is consistent with a wealth of genetic evidence from mtDNA, Y chromosome polymorphisms, and autosomal markers which demonstrates a greater diversity in Africans than non-Africans and supports an African origin for anatomically modern humans (for review, see Tishkoff and Williams 2002). Although other explanations are possible, an African origin is further supported by the presence at multiple loci of ancestral sequences seen exclusively in Africa (Takahata et al. 2001).
Although most studies of genetic variation have identified greater diversity in Africans compared with non-Africans, the difference in lineage frequencies at the insulin minisatellite between these population groups is unusually large, with 28% of the total genetic variance being due to differences between Africans and non-Africans…
We now extend these minisatellite structural studies by analyzing SNP variation flanking the minisatellite in the three African and three non-African populations previously analyzed. If the 22 minisatellite lineages are truly monophyletic and deeply diverged from each other, then the unusually great differentiation between Africans and non-Africans should be reflected by elevated haplotype differentiation. SNP-based haplotypes could also give further clues about the nature, timing, and intensity of selective pressures in this region of DNA…
Sequence comparisons between humans and chimpanzees generated a sequence divergence estimate of 2.11%. By random concatenation of human and chimpanzee sequence data from 53 noncoding autosomal loci described by Chen and Li (2001), we generated a distribution of divergences between these sequences. Divergence estimates as high as 2.11% were not observed in this distribution, allowing us to show that sequence divergence at the insulin region was significantly (P less than 0.05) higher than at most other loci (see Methods). Furthermore, sequence comparisons with other great apes revealed high divergence between all species analyzed (Supplemental Information Table S1, available online at www.genome.org Chen and Li 2001). This elevated divergence may reflect the high GC content near the insulin gene (65%) and the abundance of segregating sites at CpG dinucleotides. A relative rates test performed on these sequence data showed that the insulin gene region was evolving in a clock-like fashion (P > 0.05)…
Patterns of Linkage Disequilibrium D' is a measure of complete disequilibrium where |D' | = 1 if at most three of the four possible haplotypes are present (Lewontin 1964). To investigate patterns of linkage disequilibrium (LD) at the insulin gene region, D' was estimated between all pairwise combinations of markers in each of the six populations tested (Fig. 2). High LD was observed in each of the three non-African populations across the 7.8-kb region. The only exception is the 5' most distal SNP INS-1, though LD has been reported to extend over 6 kb 5' of this position (Doria et al. 1996), suggesting that LD breakdown at INS-1 may instead have resulted from recurrent mutation or localized gene conversion at this marker rather than from crossover. The main difference between the non-African populations was reduced statistical power supporting LD in the Japanese, a consequence of 95% of chromosomes within this population belonging to lineage I…
The degree of LD in the insulin gene region is lower in Africans compared to non-Africans (Fig. 2). Patterns of complete disequilibrium (D' ) do show evidence for possible LD blocks that broadly correspond to the two domains of absolute disequilibrium (delta symbol) identified in non-Africans (Table 2B). Application of the four-gamete test to these populations found evidence of recombination between the two blocks of disequilibrium. Furthermore, there was evidence of homoplasy at four markers within the INS LD block (INS-21, INS-24, INS-69, and INS-28) which cluster in an interval from 50 bp 5' of the minisatellite to 800 bp 3' of the minisatellite (Fig. 2B, Table 2B). This clustering, plus the observation that two of the four markers are not located at sites prone to increased mutation, argue against recurrent mutation and instead provide evidence of a minisatellite-associated increase in local recombination frequency. Interestingly there is little evidence that linkage phase has been disrupted between markers distal to these apparent recombinants, suggesting that either crossing-over occurred between similar haplotypes or that recombination may be generating conversions in this region. The lack of observable recombinants in this region in non-Africans may simply reflect the fact that these markers are monomorphic in Europeans and Asians.
Historical recombination rates in the insulin gene region were investigated by analyzing the rate of decay of absolute disequilibrium with distance (Fig. 2C). LD decays more rapidly in Africans than non-Africans, with extended LD greatest in the Japanese population, followed by Kazakhstan, then the U.K….
Ps - In this study, one of the interesting points, is that of the seeming correlation between level of linkage disequilibrium and level of diversity: the lower the diversity across the insulin DNA segment, the stronger the linkage disequilibrium, was observed. With higher diversity of minisatellite variation, relatively lower linkage disequilibrium was observed.
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