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A molecular basis for classic blond hair color in Europeans

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Abstract

Hair color differences are among the most obvious examples of phenotypic variation in humans. Although genome-wide association studies (GWAS) have implicated multiple loci in human pigment variation, the causative base-pair changes are still largely unknown1. Here we dissect a regulatory region of the KITLG gene (encoding KIT ligand) that is significantly associated with common blond hair color in northern Europeans2. Functional tests demonstrate that the region contains a regulatory enhancer that drives expression in developing hair follicles. This enhancer contains a common SNP (rs12821256) that alters a binding site for the lymphoid enhancer-binding factor 1 (LEF1) transcription factor, reducing LEF1 responsiveness and enhancer activity in cultured human keratinocytes. Mice carrying ancestral or derived variants of the human KITLG enhancer exhibit significant differences in hair pigmentation, confirming that altered regulation of an essential growth factor contributes to the classic blond hair phenotype found in northern Europeans.

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Figure 1: A distant regulatory region upstream of the KITLG gene controls hair pigmentation in humans and mice.
Figure 2: The human blond-associated region contains a functional hair follicle enhancer.
Figure 3: Variant hair follicle enhancers produce altered levels of gene expression.
Figure 4: The blond-associated allele at rs12821256 alters a TCF/LEF binding site and reduces LEF responsiveness in keratinocytes.
Figure 5: Mouse lines differing at a single base-pair position in the KITLG hair enhancer (HE) show obvious differences in hair color.

References

  1. 1

    Sturm, R.A. Molecular genetics of human pigmentation diversity. Hum. Mol. Genet. 18, R9–R17 (2009).

    CAS  PubMed  Google Scholar 

  2. 2

    Sulem, P. et al. Genetic determinants of hair, eye and skin pigmentation in Europeans. Nat. Genet. 39, 1443–1452 (2007).

    CAS  Google Scholar 

  3. 3

    Cruz-Inigo, A.E., Ladizinski, B. & Sethi, A. Albinism in Africa: stigma, slaughter and awareness campaigns. Dermatol. Clin. 29, 79–87 (2011).

    CAS  PubMed  Google Scholar 

  4. 4

    Pitman, J. On Blondes (Bloomsbury Publishing, New York, 2003).

  5. 5

    Homer. The Iliad of Homer (University of Chicago Press, Chicago, 2011).

  6. 6

    Han, J. et al. A genome-wide association study identifies novel alleles associated with hair color and skin pigmentation. PLoS Genet. 4, e1000074 (2008).

    PubMed  PubMed Central  Google Scholar 

  7. 7

    Sulem, P. et al. Two newly identified genetic determinants of pigmentation in Europeans. Nat. Genet. 40, 835–837 (2008).

    CAS  PubMed  Google Scholar 

  8. 8

    Zhang, M. et al. Genome-wide association studies identify several new loci associated with pigmentation traits and skin cancer risk in European Americans. Hum. Mol. Genet. 22, 2948–2959 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  9. 9

    Kenny, E.E. et al. Melanesian blond hair is caused by an amino acid change in TYRP1. Science 336, 554 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  10. 10

    Hindorff, L.A. et al. Potential etiologic and functional implications of genome-wide association loci for human diseases and traits. Proc. Natl. Acad. Sci. USA 106, 9362–9367 (2009).

    CAS  PubMed  Google Scholar 

  11. 11

    ENCODE Project Consortium. An integrated encyclopedia of DNA elements in the human genome. Nature 489, 57–74 (2012).

  12. 12

    Praetorius, C. et al. A polymorphism in IRF4 affects human pigmentation through a tyrosinase-dependent MITF/TFAP2A pathway. Cell 155, 1022–1033 (2013).

    CAS  PubMed  Google Scholar 

  13. 13

    Yang, J. et al. Common SNPs explain a large proportion of the heritability for human height. Nat. Genet. 42, 565–569 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  14. 14

    Olalde, I. et al. Derived immune and ancestral pigmentation alleles in a 7,000-year-old Mesolithic European. Nature 507, 225–228 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  15. 15

    Morrison-Graham, K. & Takahashi, Y. Steel factor and c-kit receptor: from mutants to a growth factor system. Bioessays 15, 77–83 (1993).

    CAS  PubMed  Google Scholar 

  16. 16

    Russell, E.S. Hereditary anemias of the mouse: a review for geneticists. Adv. Genet. 20, 357–459 (1979).

    CAS  PubMed  Google Scholar 

  17. 17

    Nocka, K. et al. Molecular bases of dominant negative and loss of function mutations at the murine c-kit/white spotting locus: W37, Wv, W41 and W. EMBO J. 9, 1805–1813 (1990).

    CAS  PubMed  PubMed Central  Google Scholar 

  18. 18

    Bedell, M.A., Copeland, N.G. & Jenkins, N.A. Multiple pathways for Steel regulation suggested by genomic and sequence analysis of the murine Steel gene. Genetics 142, 927–934 (1996).

    CAS  PubMed  PubMed Central  Google Scholar 

  19. 19

    Walsh, S. et al. The HIrisPlex system for simultaneous prediction of hair and eye colour from DNA. Forensic Sci. Int. Genet. 7, 98–115 (2013).

    CAS  Google Scholar 

  20. 20

    Rajeevan, H. et al. ALFRED: the ALelle FREquency Database. Update. Nucleic Acids Res. 31, 270–271 (2003).

    CAS  PubMed  PubMed Central  Google Scholar 

  21. 21

    Abecasis, G.R. et al. An integrated map of genetic variation from 1,092 human genomes. Nature 491, 56–65 (2012).

    PubMed  Google Scholar 

  22. 22

    Beechey, C.V., Loutit, J.F. & Searle, A.G. Panda, a new Steel allele. Mouse News Lett. 74, 92 (1986).

    Google Scholar 

  23. 23

    Bedell, M.A. et al. DNA rearrangements located over 100 kb 5′ of the Steel (Sl)-coding region in Steel-panda and Steel-contrasted mice deregulate Sl expression and cause female sterility by disrupting ovarian follicle development. Genes Dev. 9, 455–470 (1995).

    CAS  PubMed  Google Scholar 

  24. 24

    Peters, E.M.J., Tobin, D.J., Botchkareva, N., Maurer, M. & Paus, R. Migration of melanoblasts into the developing murine hair follicle is accompanied by transient c-Kit expression. J. Histochem. Cytochem. 50, 751–766 (2002).

    CAS  PubMed  Google Scholar 

  25. 25

    Jordan, S.A. & Jackson, I.J. MGF (KIT ligand) is a chemokinetic factor for melanoblast migration into hair follicles. Dev. Biol. 225, 424–436 (2000).

    CAS  PubMed  Google Scholar 

  26. 26

    Boukamp, P. et al. Normal keratinization in a spontaneously immortalized aneuploid human keratinocyte cell line. J. Cell Biol. 106, 761–771 (1988).

    CAS  PubMed  Google Scholar 

  27. 27

    Giese, K., Amsterdam, A. & Grosschedl, R. DNA-binding properties of the HMG domain of the lymphoid-specific transcriptional regulator LEF-1. Genes Dev. 5, 2567–2578 (1991).

    CAS  PubMed  Google Scholar 

  28. 28

    Travis, A., Amsterdam, A., Belanger, C. & Grosschedl, R. LEF-1, a gene encoding a lymphoid-specific protein with an HMG domain, regulates T-cell receptor α enhancer function. Genes Dev. 5, 880–894 (1991).

    CAS  PubMed  Google Scholar 

  29. 29

    Waterman, M.L., Fischer, W.H. & Jones, K.A. A thymus-specific member of the HMG protein family regulates the human T cell receptor Cα enhancer. Genes Dev. 5, 656–669 (1991).

    CAS  PubMed  Google Scholar 

  30. 30

    Zhou, P., Byrne, C., Jacobs, J. & Fuchs, E. Lymphoid enhancer factor 1 directs hair follicle patterning and epithelial cell fate. Genes Dev. 9, 700–713 (1995).

    CAS  PubMed  Google Scholar 

  31. 31

    DasGupta, R. & Fuchs, E. Multiple roles for activated LEF/TCF transcription complexes during hair follicle development and differentiation. Development 126, 4557–4568 (1999).

    CAS  PubMed  Google Scholar 

  32. 32

    van Genderen, C. et al. Development of several organs that require inductive epithelial-mesenchymal interactions is impaired in LEF-1–deficient mice. Genes Dev. 8, 2691–2703 (1994).

    CAS  PubMed  Google Scholar 

  33. 33

    Zhang, Y. et al. Activation of β-catenin signaling programs embryonic epidermis to hair follicle fate. Development 135, 2161–2172 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  34. 34

    Love, J.J. et al. Structural basis for DNA bending by the architectural transcription factor LEF-1. Nature 376, 791–795 (1995).

    CAS  PubMed  Google Scholar 

  35. 35

    Yun, K., So, J.S., Jash, A. & Im, S.H. Lymphoid enhancer binding factor 1 regulates transcription through gene looping. J. Immunol. 183, 5129–5137 (2009).

    CAS  PubMed  Google Scholar 

  36. 36

    Jash, A., Yun, K., Sahoo, A., So, J.S. & Im, S.H. Looping mediated interaction between the promoter and 3′ UTR regulates type II collagen expression in chondrocytes. PLoS ONE 7, e40828 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  37. 37

    Visser, M., Kayser, M. & Palstra, R.J. HERC2 rs12913832 modulates human pigmentation by attenuating chromatin-loop formation between a long-range enhancer and the OCA2 promoter. Genome Res. 22, 446–455 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  38. 38

    Berger, M.F. et al. Compact, universal DNA microarrays to comprehensively determine transcription-factor binding site specificities. Nat. Biotechnol. 24, 1429–1435 (2006).

    CAS  PubMed  PubMed Central  Google Scholar 

  39. 39

    Newburger, D.E. & Bulyk, M.L. UniPROBE: an online database of protein binding microarray data on protein-DNA interactions. Nucleic Acids Res. 37, D77–D82 (2009).

    CAS  PubMed  Google Scholar 

  40. 40

    Tasic, B. et al. Site-specific integrase-mediated transgenesis in mice via pronuclear injection. Proc. Natl. Acad. Sci. USA 108, 7902–7907 (2011).

    CAS  PubMed  Google Scholar 

  41. 41

    Grichnik, J.M., Burch, J.A., Burchette, J. & Shea, C.R. The SCF/KIT pathway plays a critical role in the control of normal human melanocyte homeostasis. J. Invest. Dermatol. 111, 233–238 (1998).

    CAS  PubMed  Google Scholar 

  42. 42

    Kamberov, Y.G. et al. Modeling recent human evolution in mice by expression of a selected EDAR variant. Cell 152, 691–702 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  43. 43

    Veeman, M.T., Slusarski, D.C., Kaykas, A., Louie, S.H. & Moon, R.T. Zebrafish prickle, a modulator of noncanonical Wnt/Fz signaling, regulates gastrulation movements. Curr. Biol. 13, 680–685 (2003).

    CAS  PubMed  Google Scholar 

  44. 44

    DiLeone, R.J., Russell, L.B. & Kingsley, D.M. An extensive 3′ regulatory region controls expression of Bmp5 in specific anatomical structures of the mouse embryo. Genetics 148, 401–408 (1998).

    CAS  PubMed  PubMed Central  Google Scholar 

  45. 45

    Mortlock, D.P., Guenther, C. & Kingsley, D.M. A general approach for identifying distant regulatory elements applied to the Gdf6 gene. Genome Res. 13, 2069–2081 (2003).

    CAS  PubMed  PubMed Central  Google Scholar 

  46. 46

    Whitlock, M.C. Combining probability from independent tests: the weighted Z-method is superior to Fisher's approach. J. Evol. Biol. 18, 1368–1373 (2005).

    CAS  PubMed  Google Scholar 

  47. 47

    Pfaffl, M.W. A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res. 29, e45 (2001).

    CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

We thank R. Moon (University of Washington) for the XE237 LEF1 expression plasmid, R. Nusse (Stanford University) for the SuperTOPFlash plasmid, C. Lowe (Stanford University) for help with statistical and 1000 Genomes Project analysis and members of the Kingsley laboratory for useful comments on the manuscript. This work was supported in part by the University of Georgia Research Foundation (M.A.B.) and by US National Institutes of Health grants GM65393 (M.A.B.), R01-NS050835 (L.L.) and a Center of Excellence in Genomic Science award 5P50HG2568 (D.M.K.). L.L. and D.M.K. are investigators of the Howard Hughes Medical Institute.

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C.A.G. and D.M.K. conceived and oversaw the project. M.A.B. isolated and sequenced the Slpan breakpoint. B.T. and L.L. provided advice, reagents and mice for generating site-specific integrants. C.A.G. performed the gene expression analysis in Slpan mutants, carried out the transgenic analysis of the blond-associated GWAS interval, identified the hair follicle enhancer and performed in vitro and in vivo tests of the effects of the rs12821256 polymorphism. C.A.G. and D.M.K. wrote the manuscript with input from all authors.

Corresponding author

Correspondence to David M Kingsley.

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The authors declare no competing financial interests.

Integrated supplementary information

Supplementary Figure 1 H2 transgenic embryos.

Fifteen transgenic embryos produced by pronuclear injection with the 6.7-kb H2 plasmid are shown. Each embryo represents an independent genomic integration event. Embryos were collected at E16.5, stained for lacZ activity and bisected before imaging to show both external lateral (l) and internal medial (m) expression patterns. The skin (n = 13) and kidney (n = 14) were consistent sites of expression. The asterisk denotes the position of the kidney in the internal images. Scale bar, 1 mm.

Supplementary Figure 2 H2b transgenic embryos.

Thirteen transgenic embryos produced by pronuclear injection with the 1.5-kb H2b plasmid are shown. Each embryo represents an independent genomic integration event. Embryos were collected at E16.5, stained for lacZ activity and bisected before imaging to show both external lateral (l) and internal medial (m) expression patterns. The kidney (n = 12) was the only consistent site of expression. The asterisk denotes the position of the kidney in the internal images. Scale bar, 1 mm.

Supplementary Figure 3 HFE transgenic embryos.

Eleven transgenic embryos produced by pronuclear injection with the 1.9-kb HFE clone are pictured. Each embryo represents an independent genomic integration event. Embryos were collected at E16.5, stained for lacZ activity and bisected before imaging to show both external lateral (l) and internal medial (m) expression patterns. Hair/skin expression was visible in 8 of the 11 embryos. The asterisk denotes the position of the kidney in the internal images. Scale bar, 1 mm.

Supplementary Figure 4 H2-BLD transgenic embryos.

Nine transgenic embryos produced by pronuclear injection with the 6.7-kb H2-BLD plasmid are shown. Each embryo represents an independent genomic integration event. Embryos were collected at E16.5, stained for lacZ activity and bisected before imaging to show both external lateral (l) and internal medial (m) expression patterns. Hair/skin (n = 7) and kidney (n = 7) were consistent sites of expression. No clear difference in expression compared to the complete set of H2 (H2-ANC) transgenic embryos was evident (see Supplementary Fig. 1). The asterisk denotes the position of the kidney in the internal images. Scale bar, 1 mm.

Supplementary Figure 5 H2-DEL transgenic embryos.

Eight transgenic embryos produced by pronuclear injection with the 6.7-kb H2-DEL plasmid are shown. Each embryo represents an independent genomic integration event. Embryos were collected at E16.5, stained for lacZ activity and bisected before imaging to show both external lateral (l) and internal medial (m) expression patterns. Seven of the embryos show hair/skin expression. However the strength of this staining appeared reduced compared to H2-ANC and H2-BLD embryos, particularly in embryos that showed comparably strong kidney expression (such as embryos 2 and 5). The asterisk denotes the position of the kidney in the internal images. Scale bar, 1 mm.

Supplementary Figure 6 Additional pigmentation phenotypes seen in hair enhancer-Kitl mice.

BLD-Kitl/+ and ANC-Kitl/+ heterozygotes exhibit several altered pigmentation patterns compared to wild-type (FVB/C57BL/6J F1 hybrid) littermates. At 2 months, ectopic pigmentation is seen on the muzzles (arrowheads in a) and the epithelium of the antitragus and ear canal (arrows in b–d). In contrast, BLD-Kitl/+ and ANC-Kitl/+ heterozygotes show reduced pigmentation in the whiskers (arrows in a) and the hair on the digits (arrows in e), perhaps because of competitive interactions between body sites for melanocyte colonization and development48 or premature differentiation of migrating melanocytes49. (f–h) Cross-sections (6 μm) through dorsal skin from 2-month-old (f) wild-type, (g) BLD-Kitl/+ and (h) ANC-Kitl/+ heterozygotes counterstained with nuclear fast red. Elevated Kitl expression controlled by both the BLD and ANC hair enhancers leads to ectopic pigmentation of the bulge region of hair follicles (arrows) and the basal epidermis (asterisks). DP, dermal papilla. Scale bars, 30 μm.

Supplementary Figure 7 Analysis of pigment levels in zigzag hairs from site-specific transgenic mice.

(a) Photographs of zigzag hairs from wild-type (+/+; FVB/C57BL/6J F1 hybrid), BLD (BLD-Kitl/+) line 2 and ANC (ANC-Kitl/+) line 2 heterozygotes at P21. Fifteen hairs per mouse were analyzed to determine the fraction of pigmented pixels per hair shaft. (b) Mean pigmentation density in different genotypes. Both BLD-Kitl/+ and ANC-Kitl/+ heterozygotes exhibit significantly higher levels of pigmentation than wild-type controls. Notably, the amount of pigment in BLD-Kitl/+ heterozygotes is also significantly less than is found in ANC-Kitl/+ heterozygotes (P = 0.0278). Error bars indicate s.e.m. Unpaired t-test values; *P < 0.05, **P < 5 ×10–3.

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Guenther, C., Tasic, B., Luo, L. et al. A molecular basis for classic blond hair color in Europeans. Nat Genet 46, 748–752 (2014). https://doi.org/10.1038/ng.2991

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