Blue eye color in humans may be caused by a perfectly associated founder mutation in a regulatory element located within the <Emphasis Type="Italic">HERC2</Emphasis> gene inhibiting <Emphasis Type="Italic">OCA2</Emphasis> expression

The human eye color is a quantitative trait displaying multifactorial inheritance. Several studies have shown that the OCA2 locus is the major contributor to the human eye color variation. By linkage analysis of a large Danish family, we finemapped the blue eye color locus to a 166 Kbp region within the HERC2 gene. By association analyses, we identified two SNPs within this region that were perfectly associated with the blue and brown eye colors: rs12913832 and rs1129038. Of these, rs12913832 is located 21.152 bp upstream from the OCA2 promoter in a highly conserved sequence in intron 86 of HERC2. The brown eye color allele of rs12913832 is highly conserved throughout a number of species. As shown by a Luciferase assays in cell cultures, the element significantly reduces the activity of the OCA2 promoter and electrophoretic mobility shift assays demonstrate that the two alleles bind different subsets of nuclear extracts. One single haplotype, represented by six polymorphic SNPs covering half of the 3′ end of the HERC2 gene, was found in 155 blue-eyed individuals from Denmark, and in 5 and 2 blue-eyed individuals from Turkey and Jordan, respectively. Hence, our data suggest a common founder mutation in an OCA2 inhibiting regulatory element as the cause of blue eye color in humans. In addition, an LOD score of Z = 4.21 between hair color and D14S72 was obtained in the large family, indicating that RABGGTA is a candidate gene for hair color. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Genotyping of five single nucleotide polymorphisms in the OCA2 and HERC2 genes associated with blue‐brown eye color in the Japanese population

Human eye color is a polymorphic phenotype influenced by multiple genes. It has recently been reported that three single nucleotide polymorphisms (SNPs) within intron 1 of the OCA2 gene (rs7495174, rs4778241, rs4778138) and two SNPs in intron 86 (rs12913832) and the 3′ UTR region (rs1129038) of the HERC2 gene—located in the upstream of the OCA2 locus —have a high statistical association with human eye color. The present study is the first to examine in detail the genotype and haplotype frequencies for these five SNPs in an Asian (Japanese) population (n = 523) comprising solely brown‐eyed individuals. Comparison of the genotype and haplotype distributions in Japanese with those in African and European subjects revealed significant differences between Japanese and other populations. Analysis of haplotypes consisting of four SNPs at the HERC2‐OCA2 locus (rs12913832/rs7495174/rs4778241/rs4778138) showed that the most frequent haplotype in the Japanese population is A‐GAG (0.568), while the frequency of this haplotype is rather low in the European population, even in the brown‐eyed group (0.167). The haplotype distribution in the Japanese population was significantly different from that in the brown‐eyed European group (F_ST = 0.18915). Copyright © 2009 John Wiley & Sons, Ltd.     

Genome-Wide Association Studies of Quantitatively Measured Skin,Hair, and Eye Pigmentation in Four European Populations

Pigmentation of the skin, hair, and eyes varies both within and between human populations. Identifying the genes and alleles underlying this variation has been the goal of many candidate gene and several genome-wide association studies (GWAS). Most GWAS for pigmentary traits to date have been based on subjective phenotypes using categorical scales. But skin, hair, and eye pigmentation vary continuously. Here, we seek to characterize quantitative variation in these traits objectively and accurately and to determine their genetic basis. Objective and quantitative measures of skin, hair, and eye color were made using reflectance or digital spectroscopy in Europeans from Ireland, Poland, Italy, and Portugal. A GWAS was conducted for the three quantitative pigmentation phenotypes in 176 women across 313,763 SNP loci, and replication of the most significant associations was attempted in a sample of 294 European men and women from the same countries. We find that the pigmentation phenotypes are highly stratified along axes of European genetic differentiation. The country of sampling explains approximately 35% of the variation in skin pigmentation, 31% of the variation in hair pigmentation, and 40% of the variation in eye pigmentation. All three quantitative phenotypes are correlated with each other. In our two-stage association study, we reproduce the association of rs1667394 at the OCA2/HERC2 locus with eye color but we do not identify new genetic determinants of skin and hair pigmentation supporting the lack of major genes affecting skin and hair color variation within Europe and suggesting that not only careful phenotyping but also larger cohorts are required to understand the genetic architecture of these complex quantitative traits. Interestingly, we also see that in each of these four populations, men are more lightly pigmented in the unexposed skin of the inner arm than women, a fact that is underappreciated and may vary across the world.     

Iris pigmentation as a quantitative trait: variation in populations of European,East Asian and South Asian ancestry and association with candidate gene polymorphisms

In this study, we present a new quantitative method to measure iris colour based on high‐resolution photographs. We applied this method to analyse iris colour variation in a sample of individuals of East Asian, European and South Asian ancestry. We show that measuring iris colour using the coordinates of the CIELAB colour space uncovers a significant amount of variation that is not captured using conventional categorical classifications, such as ‘brown’, ‘blue’ or ‘green’. We tested the association of a selected panel of polymorphisms with iris colour in each population group. Six markers showed significant associations with iris colour in the European sample, three in the South Asian sample and two in the East Asian sample. We also observed that the marker HERC2 rs12913832, which is the main determinant of ‘blue’ versus ‘brown’ iris colour in European populations, is also significantly associated with central heterochromia in the European sample.     

Technical note: quantitative measures of iris color using high resolution photographs

Our understanding of the genetic architecture of iris color is still limited. This is partly related to difficulties associated with obtaining quantitative measurements of eye color. Here we introduce a new automated method for measuring iris color using high resolution photographs. This method extracts color measurements in the CIE 1976 L*a*b* (CIELAB) color space from a 256 by 256 pixel square sampled from the 9:00 meridian of the iris. Color is defined across three dimensions: L* (the lightness coordinate), a* (the red-green coordinate), and b* (the blue-yellow coordinate). We applied this method to a sample of individuals of diverse ancestry (East Asian, European and South Asian) that was genotyped for the HERC2 rs12913832 polymorphism, which is strongly associated with blue eye color. We identified substantial variation in the CIELAB color space, not only in the European sample, but also in the East Asian and South Asian samples. As expected, rs12913832 was significantly associated with quantitative iris color measurements in subjects of European ancestry. However, this SNP was also strongly associated with iris color in the South Asian sample, although there were no participants with blue irides in this sample. The usefulness of this method is not restricted only to the study of iris pigmentation. High-resolution pictures of the iris will also make it possible to study the genetic variation involved in iris textural patterns, which show substantial heritability in human populations.     

Sequences associated with human iris pigmentation

To determine whether and how common polymorphisms are associated with natural distributions of iris colors, we surveyed 851 individuals of mainly European descent at 335 SNP loci in 13 pigmentation genes and 419 other SNPs distributed throughout the genome and known or thought to be informative for certain elements of population structure. We identified numerous SNPs, haplotypes, and diplotypes (diploid pairs of haplotypes) within the OCA2, MYO5A, TYRP1, AIM, DCT, and TYR genes and the CYP1A2-15q22-ter, CYP1B1-2p21, CYP2C8-10q23, CYP2C9-10q24, and MAOA-Xp11.4 regions as significantly associated with iris colors. Half of the associated SNPs were located on chromosome 15, which corresponds with results that others have previously obtained from linkage analysis. We identified 5 additional genes (ASIP, MC1R, POMC, and SILV) and one additional region (GSTT2-22q11.23) with haplotype and/or diplotypes, but not individual SNP alleles associated with iris colors. For most of the genes, multilocus gene-wise genotype sequences were more strongly associated with iris colors than were haplotypes or SNP alleles. Diplotypes for these genes explain 15% of iris color variation. Apart from representing the first comprehensive candidate gene study for variable iris pigmentation and constituting a first step toward developing a classification model for the inference of iris color from DNA, our results suggest that cryptic population structure might serve as a leverage tool for complex trait gene mapping if genomes are screened with the appropriate ancestry informative markers.     

Comprehensive candidate gene study highlights UGT1A and BNC2 as new genes determining continuous skin color variation in Europeans

Natural variation in human skin pigmentation is primarily due to genetic causes rooted in recent evolutionary history. Genetic variants associated with human skin pigmentation confer risk of skin cancer and may provide useful information in forensic investigations. Almost all previous gene-mapping studies of human skin pigmentation were based on categorical skin color information known to oversimplify the continuous nature of human skin coloration. We digitally quantified skin color into hue and saturation dimensions for 5,860 Dutch Europeans based on high-resolution skin photographs. We then tested an extensive list of 14,185 single nucleotide polymorphisms in 281 candidate genes potentially involved in human skin pigmentation for association with quantitative skin color phenotypes. Confirmatory association was revealed for several known skin color genes including HERC2, MC1R, IRF4, TYR, OCA2, and ASIP. We identified two new skin color genes: genetic variants in UGT1A were significantly associated with hue and variants in BNC2 were significantly associated with saturation. Overall, digital quantification of human skin color allowed detecting new skin color genes. The variants identified in this study may also contribute to the risk of skin cancer. Our findings are also important for predicting skin color in forensic investigations.     

Genetics of human iris colour and patterns

The presence of melanin pigment within the iris is responsible for the visual impression of human eye colouration with complex patterns also evident in this tissue, including Fuchs’ crypts, nevi, Wolfflin nodules and contraction furrows. The genetic basis underlying the determination and inheritance of these traits has been the subject of debate and research from the very beginning of quantitative trait studies in humans. Although segregation of blue‐brown eye colour has been described using a simple Mendelian dominant‐recessive gene model this is too simplistic, and a new molecular genetic perspective is needed to fully understand the biological complexities of this process as a polygenic trait. Nevertheless, it has been estimated that 74% of the variance in human eye colour can be explained by one interval on chromosome 15 that contains the OCA2 gene. Fine mapping of this region has identified a single base change rs12913832 T/C within intron 86 of the upstream HERC2 locus that explains almost all of this association with blue‐brown eye colour. A model is presented whereby this SNP, serving as a target site for the SWI/SNF family member HLTF, acts as part of a highly evolutionary conserved regulatory element required for OCA2 gene activation through chromatin remodelling. Major candidate genes possibly effecting iris patterns are also discussed, including MITF and PAX6.     

A genome scan for eye color in 502 twin families: most variation is due to a QTL on chromosome 15q.

We have rated eye color on a 3-point scale (1 = blue/grey, 2 = hazel/green, 3 = brown) in 502 twin families and carried out a 5-10 cM genome scan (400-757 markers). We analyzed eye color as a threshold trait and performed multipoint sib pair linkage analysis using variance components analysis in Mx. A lod of 19.2 was found at the marker D15S1002, less than 1 cM from OCA2, which has been previously implicated in eye color variation. We estimate that 74% of variance in eye color liability is due to this QTL and a further 18% due to polygenic effects. However, a large shoulder on this peak suggests that other loci affecting eye color may be telomeric of OCA2 and inflating the QTL estimate. No other peaks reached genome-wide significance, although lods > 2 were seen on 5p and 14q and lods >1 were additionally seen on chromosomes 2, 3, 6, 7, 8, 9, 17 and 18. Most of these secondary peaks were reduced or eliminated when we repeated the scan as a two locus analysis with the 15q linkage included, although this does not necessarily exclude them as false positives. We also estimated the interaction between the 15q QTL and the other marker locus but there was only minor evidence for additive x additive epistasis. Elaborating the analysis to the full two-locus model including non-additive main effects and interactions did not strengthen the evidence for epistasis. We conclude that most variation in eye color in Europeans is due to polymorphism in OCA2 but that there may be modifiers at several other loci.     

Genetic Architecture of Skin and Eye Color in an African-European Admixed Population

Variation in human skin and eye color is substantial and especially apparent in admixed populations, yet the underlying genetic architecture is poorly understood because most genome-wide studies are based on individuals of European ancestry. We study pigmentary variation in 699 individuals from Cape Verde, where extensive West African/European admixture has given rise to a broad range in trait values and genomic ancestry proportions. We develop and apply a new approach for measuring eye color, and identify two major loci (HERC2[OCA2] P = 2.3×10⁻⁶², SLC24A5 P = 9.6×10⁻⁹) that account for both blue versus brown eye color and varying intensities of brown eye color. We identify four major loci (SLC24A5 P = 5.4×10⁻²⁷, TYR P = 1.1×10⁻⁹, APBA2[OCA2] P = 1.5×10⁻⁸, SLC45A2 P = 6×10⁻⁹) for skin color that together account for 35% of the total variance, but the genetic component with the largest effect (∼44%) is average genomic ancestry. Our results suggest that adjacent cis-acting regulatory loci for OCA2 explain the relationship between skin and eye color, and point to an underlying genetic architecture in which several genes of moderate effect act together with many genes of small effect to explain ∼70% of the estimated heritability.     

Multilocus OCA2 genotypes specify human iris colors

Human iris color is a quantitative, multifactorial phenotype that exhibits quasi-Mendelian inheritance. Recent studies have shown that OCA2 polymorphism underlies most of the natural variability in human iris pigmentation but to date, only a few associated polymorphisms in this gene have been described. Herein, we describe an iris color score (C) for quantifying iris melanin content in-silico and undertake a more detailed survey of the OCA2 locus (n = 271 SNPs). In 1,317 subjects, we confirmed six previously described associations and identified another 27 strongly associated with C that were not explained by continental population stratification (OR 1.5–17.9, P = 0.03 to <0.001). Haplotype analysis with respect to these 33 SNPs revealed six haplotype blocks and 11 hap-tags within these blocks. To identify genetic features for best-predicting iris color, we selected sets of SNPs by parsing P values among possible combinations and identified four discontinuous and non-overlapping sets across the LD blocks (p-Selected SNP sets). In a second, partially overlapping sample of 1,072, samples with matching diplotypes comprised of these p-Selected OCA2 SNPs exhibited a rate of C concordance of 96.3% (n = 82), which was significantly greater than that obtained from randomly selected samples (62.6%, n = 246, P<0.0001). In contrast, the rate of C concordance using diplotypes comprised of the 11 identified hap-tags was only 83.7%, and that obtained using diplotypes comprised of all 33 SNPs organized as contiguous sets along the locus (defined by the LD block structure) was only 93.3%. These results confirm that OCA2 is the major human iris color gene and suggest that using an empirical database-driven system, genotypes from a modest number of SNPs within this gene can be used to accurately predict iris melanin content from DNA. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Oculocutaneous albinism with TYRP1 gene mutations in a Caucasian patient

Non-syndromic oculocutaneous albinism (OCA) is a clinically and genetically heterogeneous autosomal recessive disorder with mutations identified in several genes: OCA1 (tyrosinase, TYR), OCA2 (OCA2), OCA3 (tyrosinase-related protein 1, TYRP1), and OCA4 (membrane-associated transporter protein, MATP). OCA3 was thought to be restricted to black populations, where it was clinically described as rufous or brown albinism, until the recent report of a homozygous TYRP1 mutation in Caucasian patients from a consanguineous Pakistani family. Here, we describe a German patient of Caucasian origin, with a light-yellow skin, yellow-gold hair with orange highlights, fair eyelashes, several pigmented naevi, and no tendency to tan, only to burn. Eye-colour is blue-green with substance defects of the iris. Molecular analysis did not reveal any mutation in the TYR and OCA2 genes. Two mutations were found in the TYRP1 gene: a missense mutation (c.1066G>A/p.Arg356Glu) that was inherited from the mother, and a de novo single-base deletion (c.106delT/p.Leu36X). This finding suggests that mutation screening should be extended to the TYRP1 gene in patients from all ethnic origins, at least in cases where no mutations have been identified in the other OCA genes.     

Characterization of OCA2 cDNA in different porcine breeds and analysis of its potential effect on skin pigmentation in a red Iberian strain

Although the function of the OCA2 gene product has not been totally clarified, variation in OCA2 has been associated with skin and hair pigmentation in human and mouse. However, its contribution to skin colour in domestic species has not been reported. In this study, cDNA and intron 9 sequences of the porcine OCA2 gene have been characterized in several pig populations. The cDNA sequence alignment of 20 animals from eight porcine populations allowed the identification of 10 single nucleotide polymorphisms (SNPs); five of the 10 SNPs were non-synonymous. The intron 9 sequence alignment of 12 animals belonging to four pig populations revealed four additional SNPs. Skin colour variation was analysed in a red strain of Iberian pigs with segregation of three SNPs forming two OCA2 intragenic haplotypes. Results from this study provide evidence of a suggestive dominant effect of haplotypes on colour intensity and indicate an important contribution of additive polygenic effects (h2 = 0.56 +/- 0.21) to the variance of this trait.     

Implications of the Admixture Process in Skin Color Molecular Assessment

The understanding of the complex genotype-phenotype architecture of human pigmentation has clear implications for the evolutionary history of humans, as well as for medical and forensic practices. Although dozens of genes have previously been associated with human skin color, knowledge about this trait remains incomplete. In particular, studies focusing on populations outside the European-North American axis are rare, and, until now, admixed populations have seldom been considered. The present study was designed to help fill this gap. Our objective was to evaluate possible associations of 18 single nucleotide polymorphisms (SNPs), located within nine genes, and one pseudogene with the Melanin Index (MI) in two admixed Brazilian populations (Gaucho, N = 352; Baiano, N = 148) with different histories of geographic and ethnic colonization. Of the total sample, four markers were found to be significantly associated with skin color, but only two (SLC24A5 rs1426654, and SLC45A2 rs16891982) were consistently associated with MI in both samples (Gaucho and Baiano). Therefore, only these 2 SNPs should be preliminarily considered to have forensic significance because they consistently showed the association independently of the admixture level of the populations studied. We do not discard that the other two markers (HERC2 rs1129038 and TYR rs1126809) might be also relevant to admixed samples, but additional studies are necessary to confirm the real importance of these markers for skin pigmentation. Finally, our study shows associations of some SNPs with MI in a modern Brazilian admixed sample, with possible applications in forensic genetics. Some classical genetic markers in Euro-North American populations are not associated with MI in our sample. Our results point out the relevance of considering population differences in selecting an appropriate set of SNPs as phenotype predictors in forensic practice.     

Transferability of type 2 diabetes implicated loci in multi-ethnic cohorts from Southeast Asia

Recent large genome-wide association studies (GWAS) have identified multiple loci which harbor genetic variants associated with type 2 diabetes mellitus (T2D), many of which encode proteins not previously suspected to be involved in the pathogenesis of T2D. Most GWAS for T2D have focused on populations of European descent, and GWAS conducted in other populations with different ancestry offer a unique opportunity to study the genetic architecture of T2D. We performed genome-wide association scans for T2D in 3,955 Chinese (2,010 cases, 1,945 controls), 2,034 Malays (794 cases, 1,240 controls), and 2,146 Asian Indians (977 cases, 1,169 controls). In addition to the search for novel variants implicated in T2D, these multi-ethnic cohorts serve to assess the transferability and relevance of the previous findings from European descent populations in the three major ethnic populations of Asia, comprising half of the world's population. Of the SNPs associated with T2D in previous GWAS, only variants at CDKAL1 and HHEX/IDE/KIF11 showed the strongest association with T2D in the meta-analysis including all three ethnic groups. However, consistent direction of effect was observed for many of the other SNPs in our study and in those carried out in European populations. Close examination of the associations at both the CDKAL1 and HHEX/IDE/KIF11 loci provided some evidence of locus and allelic heterogeneity in relation to the associations with T2D. We also detected variation in linkage disequilibrium between populations for most of these loci that have been previously identified. These factors, combined with limited statistical power, may contribute to the failure to detect associations across populations of diverse ethnicity. These findings highlight the value of surveying across diverse racial/ethnic groups towards the fine-mapping efforts for the casual variants and also of the search for variants, which may be population-specific.     

Association of the OCA2 Polymorphism His615Arg with Melanin Content in East Asian Populations: Further Evidence of Convergent Evolution of Skin Pigmentation

The last decade has witnessed important advances in our understanding of the genetics of pigmentation in European populations, but very little is known about the genes involved in skin pigmentation variation in East Asian populations. Here, we present the results of a study evaluating the association of 10 Single Nucleotide Polymorphisms (SNPs) located within 5 pigmentation candidate genes (OCA2, DCT, ADAM17, ADAMTS20, and TYRP1) with skin pigmentation measured quantitatively in a sample of individuals of East Asian ancestry living in Canada. We show that the non-synonymous polymorphism rs1800414 (His615Arg) located within the OCA2 gene is significantly associated with skin pigmentation in this sample. We replicated this result in an independent sample of Chinese individuals of Han ancestry. This polymorphism is characterized by a derived allele that is present at a high frequency in East Asian populations, but is absent in other population groups. In both samples, individuals with the derived G allele, which codes for the amino acid arginine, show lower melanin levels than those with the ancestral A allele, which codes for the amino acid histidine. An analysis of this non-synonymous polymorphism using several programs to predict potential functional effects provides additional support for the role of this SNP in skin pigmentation variation in East Asian populations. Our results are consistent with previous research indicating that evolution to lightly-pigmented skin occurred, at least in part, independently in Europe and East Asia.     

Two Genomic Loci Control Three Eye Colors in the Domestic Pigeon (Columba livia)

The iris of the eye shows striking color variation across vertebrate species, and may play important roles in crypsis and communication. The domestic pigeon (Columba livia) has three common iris colors, orange, pearl (white), and bull (dark brown), segregating in a single species, thereby providing a unique opportunity to identify the genetic basis of iris coloration. We used comparative genomics and genetic mapping in laboratory crosses to identify two candidate genes that control variation in iris color in domestic pigeons. We identified a nonsense mutation in the solute carrier SLC2A11B that is shared among all pigeons with pearl eye color, and a locus associated with bull eye color that includes EDNRB2, a gene involved in neural crest migration and pigment development. However, bull eye is likely controlled by a heterogeneous collection of alleles across pigeon breeds. We also found that the EDNRB2 region is associated with regionalized plumage depigmentation (piebalding). Our study identifies two candidate genes for eye colors variation, and establishes a genetic link between iris and plumage color, two traits that vary widely in the evolution of birds and other vertebrates.