Discovery of copy number variants by multiplex amplifiable probe hybridization (MAPH) in candidate pigmentation genes

<div><p></p><p><i>Background</i>: Copy Number Variants (CNVs) contribute to a large fraction of genetic diversity and some of them have been reported to offer an evolutionary advantage.</p><p><i>Aim</i>: To identify CNVs in pigmentary loci that could contribute to human skin pigmentation diversity.</p><p><i>Subjects and methods</i>: This study assessed the existence of CNVs in every exon of candidate genes: <i>TYR, TYRP1, DCT, MC1R</i> and <i>SLC24A5</i>, using the Multiplex Amplifiable Probe Hybridization technique (MAPH). This study analysed a total of 99 DNA samples of unrelated individuals from different populations. Validation and further analysis in a larger Spanish sample were performed by RT-qPCR.</p><p><i>Results</i>: Five CNVs were identified by MAPH: <i>DCT</i> exons 4 and 8, <i>TYR</i> exon 1 and <i>SLC24A5</i> exons 1 and 4. Real-time quantitative PCR (RT-qPCR) confirmed the CNV in exon 1 of <i>SLC24A5</i>. This study further analysed the 5′ promoter region of <i>SLC24A5</i> and found another CNV in this region. However, no association was found between the CNV and the degree of pigmentation.</p><p><i>Conclusion</i>: Although the functional role of these structural variants in pigmentation should be the subject of future work, the results emphasize the need to consider all classes of variation (both SNPs and CNVs) when exploring the genetics of skin pigmentation.</p></div

Frequency of 374F in Spain according to the intensity of annual UV irradiation.

<p>A colored map showing the frequency of 374F per UV intensity range and the correlation equations between UV intensity and frequency of 374F. UV ranges (J/m²) include: 23500–25500; 25501–27500; 27501–29500; 29501–31500; 31501–34500; and 43300. The green line corresponds to the correlation equation obtained when the Canary Islands were included. The red line corresponds to the correlation equation after excluding the Canary Islands.</p

Estimation of selection coefficient (s) of allele 314F using the haplotipic data of 82 unrelated CEU individuals from the 1KGP.

<p>The highest likelihood was obtained for s = 0.0127 (95% CI, 0.0106–0.0148).</p

EHH analysis of the 1KGP South European samples.

<p>Simulated EHH values for the 374F haplotype using the demographic model of Gutenkunst et al. (2009). Shades of blue correspond to a smoothed color density representation of the scatter plot of all EHH simulations for each region. Blue darkness is proportional to the density of points in that vicinity. The continuous blue line corresponds to the 95th percentile of the distribution. Purple dots correspond to the observed values from the 1KGP data for South Europeans.</p

Estimation of selection coefficient (s) of allele 314F using the haplotipic data of 60 unrelated CEU individuals from the HapMap.

<p>The highest likelihood was obtained for s = 0.0243 (95% CI, 0.0111–0.0375).</p

SNPs found in the resequencing of <i>SLC45A2</i>.

<p>We show the location of the 3 SNPs found in the coding region of the gene: rs16891982, rs2287949 and rs26722, plus the most frequent SNP in intron 5: rs35397.</p

Frequency of 374F (rs16891982) and the A allele (intronic rs35397) in a) the most pigmented (Dark) and least pigmented (Light) individuals of the skin reflectance distribution from our study and in b) the populations from 1KGP.

<p>n: number of chromosomes.</p>1<p>North Europe: CEU (Utah residents with Northern and Western European Ancestry, n = 174), GBR (British in England and Scotland, n = 176) and FIN (Finnish in Finland, n = 186).</p>2<p>South Europe: TSI (Toscani in Italy, n = 196) and IBS (Iberian in Spain, n = 28).</p

Admixture map for ancestral populations (k) = 6.

<p>Each vertical line represents an individual from the corresponding population. Different colors indicate the ancestry proportions. The samples inside the black square correspond to the samples analyzed in this work. D: most pigmented individuals from our samples, L: least pigmented individuals from our sample.</p

Diversity parameters and selection test for the coding region and intron 5 of <i>SLC45A2</i> gene.

<p><b>*significant after Bonferroni correction.</b></p><p><b>n.s: not significant.</b></p><p><b>n</b>: number of chromosomes; <b>SS</b>: segregating sites; pi: nucleotide diversity; <b>Hd</b>: haplotype diversity; <b>TD (p)</b>: Tajima's D (p value); <b>FLD (p)</b>: Fu & Li's D (p value); <b>FWHn (p)</b>: normalized Fay & Wu's H (p value); <b>DHEW(p)</b>: combined test DHEW (p value).</p><p>(p values in dnaSP obtained from 5000 standard coalescent simulations).</p