Saioa López 33

Rev. Esp. Antrop. Fís. (2012) Vol. 33: 7-19

Evolution of human skin pigmentation. Genetic factors underlying variability and association with eye and hair color

LÓPEZ S.1, SMITH-ZUBIAGA I.2, IZAGIRRE N.1 Y ALONSO S.1

1 Dpto. Genética, Antropología Física y Fisiología Animal. UPV/EHU 2 Dpto. Zoología y Biología Celular Animal. UPV/EHU

Corresponding Author: Saioa López Dpto. Genética, Antropología Física y Fisiología Animal Universidad del País Vasco / Euskal Herriko Unibertsitatea Barrio Sarriena s/n 48940. Leioa (Vizcaya) [email protected] +34 946015430

ABSTRACT

Keywords: In this review we present an updated overview of the main current hypotheses for the evolution of skin color and the genetic factors underlying its variability, as well as a brief remark of the Skin Pigmentation relationship of skin pigmentation with other pigmentary phenotypes, such as eye and hair color. Genetics Pigmentation has also important biomedical implications, for instance as regards skin cancer Evolution development. The continuous nature of skin color in humans seems likely to be the result of Melanogenesis adaptive evolution; however, it is currently unknown how selection has affected the genetic architecture of pigmentation loci in different populations. Many hypotheses have been proposed to explain this issue: protection from skin cancer and sunburn, protection against folate deficiency, protection against DNA damage and oxidative stress, permeability barrier or sexual selection. Human pigmentation is presumed to be under the control of an undetermined number of genes, Recibido: 28-10-2012 which may act at different stages of melanogenesis. To date, however, only a few genes have Aceptado: 12-12-2012 been shown to have effects on normal variation in pigmentation; the strongest evidences are found in the melanogenic genes MC1R, ASIP, SLC24A5, MATP, TYR, TYRP1, DCT, OCA2 and KITLG. There are also other non-melanogenic genes with a putative role in skin pigmentation, such as VDR or beta defensins.

RESUMEN

Palabras clave: En esta revisión presentamos una visión general actualizada de las principales hipótesis sobre la evolución del color de la piel y los factores genéticos responsables de la variabilidad, así como Pigmentación de la piel una breve descripción de la relación de la pigmentación de la piel con otros fenotipos pigmenta- Genética rios, como el color de los ojos o del pelo. El estudio de la pigmentación tiene también importan- Evolución Melanogénesis tes implicaciones biomédicas, como en lo que respecta al desarrollo de cáncer de piel. La natura- leza continua del color de la piel en humanos parece ser el resultado de una evolución adaptati- va; sin embargo, actualmente se desconoce cómo la selección ha afectado a la estructura genética de los loci pigmentarios en las diferentes poblaciones. Se han propuesto varias hipótesis para explicar esto: la protección frente a cáncer de piel y quemadura solar, protección contra la defi- ciencia de folato, protección contra daño del DNA y estrés oxidativo, una barrera de permeabili- dad o la selección sexual. La pigmentación en humanos está controlada por la acción de numero- sos genes, que actúan en diferentes etapas de la melanogénesis. No obstante, hasta hoy, solo se han identificado unos pocos genes que tengan efectos en la variabilidad normal de la pigmenta- ción; las mayores evidencias se encuentran en los genes melanogénicos MC1R, ASIP, SLC24A5, MATP, TYR, TYRP1, DCT, OCA2 y KITLG. También existen otros genes no-melanogénicos con un posible papel en la pigmentación, como VDR o las beta defensinas.

Introduction protective barrier to the environment, regulating the passage of water and electrolytes while simultaneously Skin color is one of the most conspicuous mor- providing protection against damaging agents, such as phological traits in humans and its continuous nature microorganisms, ultraviolet radiation or toxic agents. It seems likely to be the result of adaptive evolution. The seems also to be involved in thermoregulation and in association of this human trait with the environment the immune system (Menon & Kligman, 2009; Ban- roots in the mid-18th century when naturalists such as gert et al. 2011) constituting thus a further layer of John Mitchell first related human skin pigmentation protection against disease. The skin is made up of with latitude and solar radiation (Mitchell & Collinson, three main layers: epidermis, dermis and subcutaneous 1744). Further support for the adaptive nature of pig- tissue. The epidermis is the outer layer, serving as the mentation comes from Relethford et al. (2002) who physical and chemical barrier between the interior bo- observed that, contrary to other neutral genetic mar- dy and environment; the dermis is the deeper layer, kers and DNA polymorphisms (which show most of providing the structural support of the skin. Below the their diversity within local populations), in skin color dermis lies the subcutaneous tissue, an important depot 88% of total variation is due to differences among ma- of fat (Menon, 2002). As regards pigmentation, the jor geographic groups. epidermis is the most important layer, as it is where melanin, the pigmentary polymer, is synthesized and The study of pigmentation is not only a major distributed. Nevertheless, skin pigmentation is also anthropological issue, but it also has important biome- influenced, although to a minor extent, by other pig- dical implications. For instance, exposure to UVB ra- ments such as carotene, reduced hemoglobin and diation from the sun increases the risk of developing oxyhemoglobin. skin cancer (Ravanat et al. 2001), so, individuals with more pigmented or darker skin are very much less li- Melanin is a complex mixture of biopolymers, kely to suffer damage in their DNA by UV radiation exclusively synthesized by melanocytes of the skin than those with pale skin. Therefore, the study of the and the retinal pigment epithelium. Melanocytes depo- genetic patterns that shape the distribution of pigmen- sit granules of melanin into cellular organelles called tation among humans can shed some light on the evo- melanosomes. Then melanosomes are transferred to lutionary forces that have molded human genetic di- keratinocytes, the cells that constitute most of the epi- versity protecting against pigmentation and ultraviolet dermis, through a process that has not been definiti- radiation related diseases in populations such as cuta- vely unraveled yet. Recent works suggest that this may neous malignant melanoma. An insufficient UVB ex- occur via the shedding vesicle system -packaging, re- posure is also related to other diseases due to the diffi- lease, uptake, and dispersion- (Ando et al. 2012; Scott, culty to maintain adequate vitamin D levels, including 2012). Melanosomes can be of two basic types depen- rickets or multiple sclerosis (Jablonski & Chaplin, ding on their shape and the melanin they contain: eu- 2012). Our understanding of the genes involved in melanosomes, ellipsoid and with black-brown eumela- these processes can be of the highest relevance as re- nin, and pheomelanosomes, spherical, with yellow- gards prevention and development of targeted treat- reddish-brown melanin (pheomelanin). These two ba- ments for specific populations. Here, we aim to review sic types of melanosomes can be produced by the me- the main current hypothesis for the evolution of skin lanocyte at different times, switching from one to the color and the genetic factors underlying its variability, other (Marks & Seabra, 2001). Rather than the number as well as to summarize the relationship of skin pig- of melanocytes in the skin, it is the number and size of mentation with other pigmentary phenotypes, such as melanosomes, the amount, type, internal arrangement eye and hair color. of melanin and the efficiency and characteristics of melanosome transfer to keratinocytes, which seems to Biology of skin pigmentation determine the high degree of variation in skin colour between individuals, within and among geographical The skin is the largest organ of the body, consti- groups. Thus, within skin keratinocytes, dark Africans tuting 16% of body weight. It functions as a dynamic and native Australians show large and singly dispersed

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melanosomes, whereas in Caucasians melanosomes lanogenesis stimulated by UVR (see Jablonsky, 2012 are smaller and aggregated into membrane-bound me- and references therein). Thus, the subsequent evolution lanosome compelexes. Interestingly, keratinocytes of of dark skin was likely an adaptive process that occu- all geographical groups exist in the single form in hair rred parallel to or immediately after the loss of hair, in bulb keratinocytes (Robins, 1991). order to provide protection against the noxious effects of UV radiation. Afterwards, as modern humans mi- The process by which melanin is synthesized in grated out of Africa towards northern latitudes where melanosomes is called melanogenesis (reviewed by solar radiation was less intense, it is speculated that Lin & Fisher, 2007; Simon et al. 2009; Park et al. they suffered again an adaptive skin depigmentation 2009). Briefly, in the absence of alpha-melanocortin process, whose nature has been associated to the need (α-MSH), pheomelanin is synthesized by default. to synthesize vitamin D (Murray, 1934; Loomis, 1967; ACTH and α-MSH, subproducts of the pro-opiomela- see also Robins, 1991 or Jablonsky, 2012 and referen- nocortin gene (POMC), act as ligands of MC1R. The ces therein). This process has been suggested to occur union of α-MSH and ACTH to MC1R leads to an in- independently in Asia and Europe (Norton et al. 2007) crease in cellular cAMP, which in its turn, leads to an prompting to the existence of a convergent evolutio- activation of protein kinase A (PKA). This leads to nary force. increased levels of microphtalmia transcription factor (MITF) expression, which upregulates the transcrip- In this regard, although it is strongly assumed tion of tyrosinase (TYR) and tyrosine related proteins that the evolution of skin pigmentation has been adap- (TYRP1 and DCT/TYRP2). Synthesis of melanin be- tive, it is currently unknown how selection has affec- gins with the hydroxylation of tyrosine to form 3,4- ted the genetic architecture of pigmentation loci in dihydroxyphenylalanine (DOPA), a step catalyzed by different populations. Many hypotheses have been the enzymes TYR and TYRP1. DOPA is then oxidized proposed to explain this issue; most of them however to DOPAquinone. In the presence of cysteine, DOPA- are non-exclusive. quinone is reduced to cysteinylDOPA, to form phaeo- melanin. In the absence of cysteine, DOPAquinone is Protection from skin cancer and sunburn converted to DOPAchrome and then to eumelanin by DCT. Basal pigmentation seems to be mostly genetica- UV radiation that reaches the Earth surface is lly determined, but skin melanocytes can also adapt composed of a mixture of UVA (320-400 nm), which their melanogenic abilities to extracellular stimuli, can penetrate deeply into the dermis of skin as well as either generated by the organism as paracrine and en- UVB (280-320 nm), which is more energetic and da- docrine factors, or by the external environment, like maging, but is absorbed and scattered in the atmosphe- ultraviolet radiation (Schiaffino, 2010). re and generally does not penetrate the dermis. The third type of UV radiation, UVC (100-280 nm), is al- Hypotheses for the evolution of skin colour most completely absorbed by the ozone layer and therefore, it does not affect the skin. Levels of UVA are It is speculated that first hominids that appeared considerable higher than UVB, being the tropical and in Africa probably had a lightly pigmented skin cove- subtropical areas those that receive the highest intensi- red with hair (Jablonski & Chaplin, 2000). Hot envi- ties. Levels of UVB are also highest in the tropics, ronmental conditions and higher activity levels asso- near the equator and in high altitude areas (Jablonski ciated with the limb proportions and bipedalism made & Chaplin, 2010). It has been proposed that darker the loss of hair necessary for the thermoregulation of skins would have been favored in regions of high ul- the body, leading to an unpigmented skin, unprotected traviolet radiation, where heavily pigmented mela- against high intensity solar radiation (Montagna, 1981; nocytes would minimize the damaging effects of UV Chaplin et al. 1994). Others suggest that the main rea- radiation, hence protecting against skin cancer (Ro- son for the evolution of dark pigmentation was to pro- bins, 1991) and sunburn (Thomson, 1951). Povey et al. tect against folate deficiency caused by elevated de- (2007) discovered that XPD, ERCC1 and XPF, which mands for folate in cell division, DNA repair, and me- play an important role in the nucleotide excision repair

9 Evolution of human skin pigmentation

(NER) pathway that deals with UV-induced damage, biologically meaningful from an evolutionary pers- show polymorphisms related to the susceptibility to pective, but to our knowledge, it has not been tested so cutaneous melanoma. From an evolutionary point of far. view this hypothesis has been prone to some objec- tions, as skin cancers generally do not develop in deve- Protection against low vitamin D levels loped societies until late in the third decade, and therefore it is expected that it should have a minimal effect Vitamin D is essential to promote a good mine- on reproductive success or fitness (Johnson et al. ralization of the bones by stimulating the absorption of 1998). However, there is evidence that supports the calcium. Precursors of vitamin D are synthesized in idea of protection. For instance, squamous cell carci- the skin from 7-dehydrocholesterol in a process ca- noma in African albinos has been reported to be the talyzed by UVB. These precursors are taken up by the commonest skin malignancy, which usually appears in liver and converted into 25-hydroxyvitamin D (25- young adults by the age of 20 (Okoro, 1975; Opara & OHD), which is carried to the kidney and metabolized Jiburum, 2010). Furthermore, the incidence of mela- into 1,25-hydroxivitamin D (1,25-(OH)2D), the active noma in the USA is nowadays around ten times higher form of vitamin D. Thus, it is speculated that a dark in Caucasians compared to African Americans (Ameri- skin would have been disadvantageous at high latitu- can Cancer Society, 2012). It also seems that death des because it would prevent the synthesis of enough rates from melanoma are not negligible in the age vitamin D. Instead, the lighter tones would have resul- group in reproductive age. Thus death rates in young ted beneficial in the less radiation-intensive regions people (aged 25-49) are increased 2-3% per year in situated at high latitudes, as low amounts of eumelanin some north and west European countries and up to 8% would allow UV radiation to penetrate the skin and in Spain (de Vries & Coebergh, 2005). Also, Robins catalyze the synthesis of the right levels of vitamin D (1991) points out that studies done in 1975 and 1985 (Holick, 2003). It was also speculated that dark skins indicated that in Nigeria and Tanzania all the “Ne- could have evolved in order to avoid intoxication cau- groid” albinos studied exhibited skin cancers or pre- sed by an excess of vitamin D synthesis. However, it malignant lesions by the age of 20. Only a 6% of Nige- has already been demonstrated that high UVB irradia- rian albinos were in the age-range of 31-60 years com- tion does not lead to vitamin D over-synthesis. Instead, pared to 20% of the non-albinos. In any case, it is im- when previtamin D reaches a critical level, it is con- portant to consider the selective relevance in evolutio- verted into two inert compounds avoiding the synthe- nary terms, not in present-day terms only (i.e. from a sis of more vitamin D (Holick et al. 1981). clinical perspective). We, in fact, ignore what was the effect of prolonged sun exposure, say, 20,000 year ago, Protection against oxidative stress in prehistoric human hunter-gatherer societies. It has also been suggested that the higher sus- Protection against folate deficiency ceptibility to skin cancer in fair-skinned Caucasians could be due to a reduced ability to detoxify the Folate is an essential vitamin to fetal deve- oxygen radicals that are produced in the cell after UV lopment and male fertility. It has been shown that defi- exposure. It has been shown that mutations in glu- ciency of this vitamin during pregnancy can result in tathione S-transferases, such as GSTT1, that contribute congenital neural tube anomalies and reduced sperma- to the defense against oxidative stress, are associated togenesis (Jablonski & Chaplin, 2000). The levels of with increased sunburn sensitivity and predisposition folate are influenced both by dietary intake and by to non-melanoma skin cancer (Kerb et al. 2002). destructive factors such as UVB radiation; therefore, the high concentration of melanin in dark skins would Permeability barrier provide protection against folate photolysis due to UV radiation. Given that there seems to be an obvious rela- Other authors suggest that the development of tionship between maintaining adequate folate levels epidermal pigmentation in UV-enriched environments and fitness (or reproductive success) this hypothesis is could have provided a superior permeability barrier,

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not only to prevent desiccation of the organism, but Initial candidate genes for skin pigmentation also to defend against microbial pathogens or allergens have been identified from mutations that lead to pig- (Elias et al. 2010). mentation disorders, such as oculocutaneous albinism, linked to OCA2 (Lee et al. 1994), Xeroderma pigmen- Sexual selection tosum, caused by a mutation in XPA gene that encodes a protein involved in DNA excision repair (Tanaka et It was Darwin who first linked variation in skin al. 1990) or Waardenburg Syndrome type 2, associated pigmentation with sexual selection. More recently, with mutations in the MITF gene (Tassabehji et al. some authors still maintain the existence of a unidirec- 1994). To date, however, only a few genes have been tional sexual selection in women’s skin color favoring shown to have effects on normal variation in pigmen- lighter pigmentation (Aoki, 2002). However, the role tation (Table 1). The strongest evidences are found in of sexual selection might have been secondary, having the melanogenic genes MC1R, ASIP, SLC24A5, MATP, women’s lighter skin first evolved for other reasons. TYR, TYRP1, DCT, OCA2 and KITLG. Due to the pro- For instance, van der Berghe and Frost (1986) specula- ved relevance of these genes in pigmentation variation, ted that differences in skin tones between males and we are going to review them in more detail; however it females would have arisen as a hormonal consequence, must be taken into consideration that there are other and then males would have preferred lighter colors as genes that have also been reported to play a role in a measure of hormonal status and reproductive poten- pigmentation, like PMEL, which encodes a melanoso- tial. Others suggested that a fairer skin color was mal matrix protein, SHROOM2 and GPR143, impor- acquired by women as a childlike trait that reduced tant in melanosomal biogenesis or ATRN, implicated in aggressiveness in men (Guthrie, 1970). And thirdly, it the eumelanin/pheomelanin switch. For further infor- was proposed that the acquisition of a lighter skin by mation (Oetting & Bennett, 2012), a database contai- women would facilitate vitamin D synthesis in order to ning all coat color genes described so far in mice and obtain adequate quantities of calcium for pregnancy their human homologues is available at and lactation (Jablonski & Chaplin, 2000). www.espcr.org/micemut.

In summary, many hypotheses have been pro- MC1R posed to explain differences in pigmentation between populations, but none has been fully tested yet. In this MC1R is a single-exon gene that codes for a regard, analyzing the diversity patterns of pigmenta- seven pass transmembrane G protein coupled receptor located in the cell membrane of the melanocytes. This tion genes can offer some insights into the evolution of skin pigmentation. gene is highly polymorphic in European populations and most of the polymorphisms correspond to non- synonym nucleotidic positions. Interestingly, non- Genetics of skin pigmentation variability synonymous variants are nearly absent in the African samples analyzed so far (Rana et al. 1999; Harding et Initially, studies carried out in mice allowed the al. 2000). Many of these variants have been correlated identification of at least 150 loci that regulate eye, skin to a fair skin and red-hair phenotype, as well as to a and hair colour, and that follow a Mendelian pattern of low tanning ability and a higher skin cancer risk (Val- inheritance (Silvers, 1979). It is clear that constitutive verde et al. 1995; Box et al. 1997). The alleles associa- pigmentation, that is, the pigmentation that is genetica- ted to the reddish-hair phenotype have been classified lly determined and is not influenced by environmental into two groups, depending on their penetrance: high- conditions, is a complex phenomenon that follows a penetrance “R” alleles (Asp84Glu, Arg151Cys, polygenic model with a few major genes (Sturm et al. Arg160Trp and Asp294His) and low-penetrance “r” 2001) and that varies in its heritability between diffe- alleles (Val60Leu, Val92Met and Arg163Gln). The rent populations (Byard, 1981). It is obvious too that functionality of these mutations has been studied in pigmentation is not influenced only by genes, but also vitro, showing that many of them lead to a reduced by other factors such as age, sex, social habits or the ability to bind either α-MSH or activate adenylyl cy- effect of environment (Robins, 1991). clase, which results in an increased production of

11 Evolution of human skin pigmentation

Table 1: SNPs known to have an effect on normal pigmentation variation

SNP Aminoacidic change Gene Pigmentary phenotype

rs1805006 Asp84Glu MC1R fair skin and red hair (R variant) rs1805007 Arg151Cys MC1R fair skin and red hair (R variant) rs1805008 Arg160Trp MC1R fair skin and red hair (R variant) rs1805009 Asp294His MC1R fair skin and red hair (R variant) rs1805006 Val60Leu MC1R fair skin and red hair (r variant) rs2228479 Val92Met MC1R fair skin and red hair (r variant) rs885479 Arg163Gln MC1R fair skin and red hair (r variant) rs6058017 g.8818A>G ASIP brown eyes and dark hair and skin rs1426654 Ala111Thr SLC24A5 light skin in Europeans rs16891982 Leu374Phe MATP dark hair and skin in Europeans rs1042602 Ser192Tyr TYR light skin in Europeans rs1800414 His615Arg OCA2 light skin in East Asia pheomelanin, instead of eumelanin, in melanocytes leading to the production of eumelanin instead of that confers a red hair and light skin phenotype pheomelanin (Kanetsky et al. 2002). (Schiöth et al. 1999; Ringholm et al. 2004). SLC24A5 Interestingly, MC1R is also located in the brain, where it acts as a receptor of the endorphins, or pain SLC24A5 (Solute carrier family 24, member 5), relieving hormones. Thus, a mutation in MC1R would encodes the NCKX5 protein. NCKX5 is a member of lead to the production of the reddish pheomelanin in the potassium-dependent sodium calcium exchanger melanocytes, and also to a different tolerance or res- protein family that locates in normal human epidermal ponse to pain in the brain. This is why it has been melanocytes. SLC24A5 seems to be one of the major hypothesized that red-haired individuals have a diffe- human pigmentation genes, as it appears to be crucial rent sensitivity to pain compared to individuals with for melanin synthesis. In particular, Lamason et al. other hair phenotype (Liem et al. 2004; Liem et al. (2005) showed a polymorphism in SLC24A5, 2005, Mogil et al. 2005). rs1426654 (Ala111Thr), which accounts for a high percentage of normal variation in pigmentation among ASIP individuals. This SNP is located in the third exon of SLC24A5. Whereas alanine form is found in 93 to ASIP is a homologue of the mouse agouti signa- 100% of samples of Africans, East Asians and Indige- ling protein gene (ASP), which has an essential role in nous Americans, the threonine variant is nearly fixed determining coat color in mice. In humans, the protein in Europeans. It has been estimated that around 25- codified by ASIP (Agouti signaling peptide) is an in- 38% variation in skin pigmentation can be explained verse agonist of α-MSH that hampers its union with by this mutation. MC1R, promoting the synthesis of pheomelanin. A SNP in the 3’ untranslated region of this gene MATP (g.8818A→G) has been strongly associated with dark hair, brown eyes and dark skin in African Americans Mutations in MATP (human membrane associa- (Kanetsky et al. 2002; Bonilla et al. 2005). It has been ted transporter gene), also known as SLC45A2, can suggested that the G allele could lead to mRNA insta- cause oculocutaneous albinism type 4 (Newton et al, bility and premature degradation of the protein, thus 2001) and are also associated with pigmentary phe- favoring the union of MC1R to its antagonist α-MSH, notypes. Two polymorphisms in this gene, Leu374Phe

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and Glu272Lys have been strongly associated with al. 2007), thus suggesting that rs1800414 in OCA2 fits dark hair, skin and eye pigmentation and shown to be a codominant model of inheritance. distinctively distributed among the different population groups (Graf et al. 2005). These non-synonymous KITLG SNPs lead to aminoacidic substitutions that could affect the gene’s function. It has been proposed that This gene encodes the ligand of the tyrosine-ki- 374Leu allele contributes to a melanosomal environ- nase receptor encoded by the KIT locus. It plays an ment favoring optimal eumelanin production, while important role in the localization of melanocytes, and 374Phe allele would originate an acidic environment, in the regulation of survival and proliferation of fully due to an alteration in the transport of protons that differentiated melanocytes in adults (Wehrle-Haller, negatively affects tyrosinase activity. 2003). Genetic variations in KITLG are associated with variation in skin, hair and eye pigmentation. Recent TYR, TYRP1 and DCT DNA sequence-based evidence has revealed a signifi- cant departure from neutrality for this gene in the Eu- Tyrosinase (TYR) and tyrosinase-related pro- ropean population (de Gruijter, 2011). teins 1 and 2 (TYRP1 and DCT) coordinate the production of melanin from tyrosine. The rate-limiting enzy- me in melanogenesis is tyrosinase, which catalyses the Other non-melanogenic genes with a role in skin conversion of tyrosine into dopaquinone (Cooksey, pigmentation 1997). This activity is required for the synthesis of the two types of melanins: pheomelanins or the more pho- The vitamin D receptor gene, VDR toprotective eumelanins. Mutations in these genes result in different types of oculocutaneous albinism, but Given the major role of UV radiation in vitamin nonpathologic polymorphisms have been also shown D synthesis, it has been speculated that polymorphisms to result in skin pigmentation variation. For instance, in VDR could also interact epistatically with skin co- two nonsynonymous SNPs in TYR, rs1042602 lor-determining genes (Hochberg & Templeton, 2010). (Ser192Tyr) (Shriver et al. 2003; Stokowski et al. The VDR gene has four major polymorphic SNPs in 2007) and rs1126809 (Arg402Gln) (Nan et al. 2009) humans and several other rare ones. Li et al. (2008) have been significantly associated with skin color and showed that polymorphisms in this gene do affect me- tanning ability. lanoma risk, although there are not strong evidences that correlate directly these polymorphisms with skin OCA2 color.

This gene encodes the human protein P, an in- Beta defensins tegral membrane protein involved in the transport of tyrosine. Mutations in OCA2 result in type II oculocu- Defensins are short cationic and cysteine rich taneous albinism, but polymorphisms in this gene have peptides that play an essential role in innate immune also been associated with skin and eye pigment varia- system, the first line of host defense against many tion. A missense mutation (rs1800414) is a candidate common microorganisms. They are classified into variant for light skin pigmentation in East Asia, where three main classes, α-, β- and θ-defensins, (Yang et al. it is found at high frequencies but absent in European 2002). It has been demonstrated that antimicrobial and West African populations (Yuasa et al. 2007; Ed- activities of defensins protect the host against bacteria, wards et al. 2010; Donnelly et al. 2012). It has been fungi, viruses and parasites. However, they have in- estimated that each copy of the derived G allele de- creasingly been observed to exhibit numerous other creases skin pigmentation by 0.85–1.3 melanin units activities, both in vitro and in vivo, that do not always (Edwards et al. 2010), contrary to polymorphisms in relate directly to host defense. Beta defensins are other genes that have an effect of more than 3 melanin highly polymorphic in sequence and copy number units per allele copy (Lamason et al. 2005; Norton et (Hollox et al. 2003). It has been demonstrated that

13 Evolution of human skin pigmentation

higher copy numbers of beta defensin genes are signi- colour can be expressed independently with different ficantly associated with the risk of psoriasis (Hollox et combinations in humans (Sturm et al. 2001). al. 2008), while lower copy numbers are correlated with predisposition to Crohn’s disease (Fellerman et al. 2006). Interestingly, polymorphisms in these genes Genetics of eye color variation have also recently been associated with pigmentation. In fact, Candille et al. (2007) identified a mutation in a Eye color is determined by two different fac- β-defensin, CBD103, which correlates with black coat tors: the pigmentation if the iris (Prota, 1998) and the color in dogs by acting as an alternative ligand for scattering of light in the stroma of the iris (Fox, 1979). MC1R. They suggest that its ability to modulate mela- It is categorized into blue, grey, green, yellow, hazel, nocortin receptor signaling (MC1R and others) may light brown and dark brown, while the shades of hair have been selected during vertebrate evolution to pro- color range from light blond, blond, dark blond, red, vide camouflage and adaptive visual cues. Soon after- red blond to brown and dark brown/black. Eye colour wards, Anderson et al. (2009) demonstrated the exis- is determined by the distribution and content of the tence of this mutation in the grey wolf, which exhibits melanocyte cells in the uveal tract of the eye. It has a molecular signature of positive selection, and in the been suggested that the number of melanocytes does coyote. Following this thread, researchers are now not differ between eye colours, being the melanin (in focusing on studying the role of β-defensins in human the form of eumelanin or pheomelanin) quantity, pigmentation (Gläser et al. 2009; Beaumont et al. packaging and quality responsible for the range of eye 2012). shades (Prota, 1998). Although many genes responsible for eye color have been so far identified, it is High-throughput sequencing methods have mostly attributed to two adjacent genes located on suggested many other candidate genes with a role in chromosome 15: OCA2 and HERC2, which explain pigmentation variability, such as SLC24A4, TPCN2 or 74% of the variability (Lehman et al. 1998; Eiberg et IRF4. SLC24A4 encodes a member of the potassium- al. 2008; Rebbeck, 2002). Nevertheless, polymor- dependent sodium/calcium exchanger protein family phisms in other genes such as TYR, TYRP1, DCT, and has been reported to contain three SNPs in linkage SLC45A2, ASIP, MYO5A, IRF4, SLC24A4 and SILV disequilibrium associated with blond hair color (Sulem are also of relative importance in determining eye co- et al. 2007). For TPCN2, two identified coding va- lor (Reviewed in Sturm & Larsson, 2009). riants have also been associated with hair color (Sulem et al. 2007). The protein encoded by IRF4 is a member of the interferon regulatory factor family of transcrip- Genetics of hair color variation tion factors that has been proposed as specific marker for melanoma and benign melanocytic nevi. A SNP in Red hair colour (RHC) phenotype, characteri- intron 4 of IRF4 has been proposed to be associated zed by high pheomelanin content, is caused by with hair color and tanning ability (Han et al. 2008). polymorphisms in MC1R which also lead to light skin, as explained above. This phenotype has been widely studied due to its relationship with propensity to sun- A brief remark on hair and eye colour burn and risk of skin cancer. SNPs in other pigmentary genes have also been signaled as candidates leading to Skin color is correlated with other pigmentary red hair (SLC45A2, SLC24A5, OCA2, HERC2 and phenotypes, such as eye and hair color. However, it ASIP) (Valenzuela et al. 2010). Quite surprisingly, should be highlighted that apart from some alleles for blond hair contains mostly eumelanin, being the con- fair skin, red hair and blue eyes (described for instance tribution of pheomelanin less than 20% (Ito & Waka- in MC1R) skin color is weakly influenced by the diffe- matsu, 2011a). Although the genetic basis of blonde rent alleles for hair or eye color. We have to bear in hair are not as clear as in the red hair colour, this phe- mind that although hair and skin melanocytes arise notype is known to be in association with SNPs in from the same embryonic source, the genes affecting pigmentary genes ASIP, TPCN2, SLC24A4, MC1R and

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TYRP1 (Valenzuela et al. 2010; Sulem et al. 2008). A of natural selection for fairer skins. However, in con- genome-wide association study has recently revealed a trast to skin pigmentation where selective advantages SNP in TYRP1, rs13289810, which causes an arginine- for lighter tones have been described (protection to-cysteine mutation that is strongly associated to against low vitamin D levels), the adaptive significan- blond hair in a population from the Solomon Islands ce of a loss of eye or hair colour pigmentation is less (Kenny et al. 2012) which proves that blond hair is not certain. an exclusive trait from nor-European populations and that it can be adquired by different mutations. Finally, brown or black hair, which contains mostly eumelanin, has been shown to be associated with SNPs in SLC45A2, HERC2 and SLC24A5. Other pigmentary Conclusions genes are also likely related to this phenotype, such as DCT, TYR, TYRP1, MC1R, OCA2, TPCN2 or CTNS Some SNPs in pigmentary genes have already (Ito & Wakamatsu, 2011b). been identified as an important source of variability; nevertheless, a high percentage of this variation cannot still be explained. Being a polygenic trait it becomes a Hair and eye color variability difficult task to associate a particular polymorphism with a concrete phenotype, thus, it can generally only While variation in skin pigmentation is notable be regarded as a predisposition or susceptibility that between populations, hair and eye colour variation is might be interacting at the same time with other most notable within European populations, being dark polymorphisms in other genes. Epistasis, that is, the brown eyes and hair dominant in African and Asian phenomenon by which the effects of one gene are mo- populations. Many hypotheses have been proposed to dified by one or several other genes, is also of the explain the variability of eye and hair color among upmost importance in pigmentation. Albinism, which Europeans: relaxation of selection, admixture with affects hair, skin, and eye color in humans, is an Neanderthals, and natural or sexual selection. It has example of epistasis. In its recessive form it has the been rejected that these high levels of diversity are due ability to mask a substantial amount of genotypic in- to relaxation of selection, as it has been estimated that formation, thus drastically altering the human's phe- it would have taken more than 850,000 years to deve- notypes (Brilliant, 2001). Red hair is another example lop, and humans have just been in Europe for 35,000 of epistasis, in which an individual that is homozygous years (Harding et al. 2000; Templeton, 2002). It has for red hair alleles has the expression of other brown/ also been speculated that this diversity could be due to blonde hair loci overridden, and therefore resulting in admixture with Neanderthals, an older European popu- red hair phenotype (Valverde, 1995). lation. Although first mtDNA comparative studies showed lack of inbreeding with Neanderthals (Currat Furthermore, the study of diverse populations is & Excoffier, 2004; Serre et al. 2004), comparisons of essential to unraveling the genetic basis of pigmenta- the Neanderthal genome with the complete genomes of tion, as it can vary among ethnic groups. A good humans from different parts of the world showed that example of convergent evolution, that is, groups that Europeans and Asians share 1% to 4% of their nuclear independently evolve similar traits, is the recent paper DNA with Neanderthals, contrary to Africans that do by Kenny et al. (2012). Here, they described a new not share any genetic variants with them (Green et al. mutation that is responsible for blond hair in Melane- 2010). However, a study by Lalueza-Fox (2007) revea- sians, but that is not seen in other human groups led a Neanderthal-specific MC1R sequence that led to worldwide. light skin color and/or red hair, supporting the hypothesis of convergent evolution between humans and Finally, the identification in beta defensins of a Neanderthals, at least with regard to this pigmentary mutation responsible for pigmentation, first in dogs, gene. It is more likely that some selective force might and now also in humans is a rather promising new be responsible for high levels of diversity in eye and direction for future research. It is a good example that hair color among Europeans. It could be a side effect our efforts to understand the phenomenon of skin color

15 Evolution of human skin pigmentation

cannot be focused only on candidate pigmentary genes Brilliant M.H. (2001) The mouse p (pink-eyed dilution) and human and pathways; rather, we must investigate alternative P genes, oculocutaneous albinism type 2 (OCA2), and melanosomal pH. Pigm. Cell Res. 14:86-93. hypotheses that complement current knowledge. Byard P.J. (1981) Quantitative genetics of human skin colour. Am. J. Phys. Anthrop. 24: 123-137. Candille S.I., Kaelin C.B., Cattanach B.M., Yu B., Thompson D.A., Nix M.A., Kerns J.A., Schmutz S.M., Millhauser G.L. & Barsh Acknowledgements G.S. (2007). A β-defensin mutation causes black coat color in domestic dogs. Science. 318: 1418–1423. Chaplin G., Jablonski N.G. & Cable N.T. (1994) Physiology, ther- This work was partly supported by grants CGL- moregulation and bipedalism. J. Hum. 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