An intronic copy number variation inSyntaxin 17determines speed of greying and melanoma incidence in Grey horses

Abstract

The Greying with age phenotype involves loss of hair pigmentation whereas skin pigmentation is not reduced and a predisposition to melanoma. The causal mutation was initially reported as a duplication of a 4.6 kb intronic sequence inSyntaxin 17. The speed of greying varies considerably among Grey horses. Here we demonstrate the presence of two differentGreyalleles,G2carrying two tandem copies of the duplicated sequence andG3carrying three. The latter is by far the most common allele, probably due to strong selection for the striking white phenotype. Our results reveal a remarkable dosage effect where theG3allele is associated with fast greying and high incidence of melanoma whereasG2is associated with slow greying and low incidence of melanoma. Epigenetic analysis, based on nanopore sequencing of genomic DNA, reveals a drastic reduction in DNA methylation in part of the duplicated sequence harboring MITF binding sites. The copy number expansion transforms a weak enhancer to a strong melanocyte-specific enhancer that underlies hair greying (G2andG3) and a drastically elevated risk of melanoma (G3only).

White horses – non-coding sequences drive premature hair greying and predisposition to melanoma

“…Based on this result, a new nomenclature for the Grey locus has been established as follows: G1 -one copy, wild type; G2 -two copies, slow greying; G3 -three copies, fast greying (Figure 2b). In contrast to G1/G3 horses that show a high incidence of melanomas, no elevated risk compared with wild-type horses was noted for G1/G2 horses (14). Thus, there is a remarkable dosage effect of the Grey mutation, increasing the copy number of the 4.6 kb STX17 sequence shows a strong positive correlation with both speed of greying and incidence of melanoma (Table 1).…”

“…We therefore decided to explore if copy number variation of the 4.6 kb STX17 sequence may explain the dramatic difference in the speed of greying that is present in some breeds, for instance, in Connemara ponies. Copy number variation was determined using digital PCR, whole genome-sequencing and sequence capture followed by Nanopore sequencing ( 14 ). The analysis of a half-sib pedigree segregating for slow and fast greying demonstrated that slow-greying horses carry chromosomes with two tandem copies of the 4.6 kb STX17 sequence, whereas fast greying horses carry a tandem triplication ( 14 ).…”

“…Copy number variation was determined using digital PCR, whole genome-sequencing and sequence capture followed by Nanopore sequencing ( 14 ). The analysis of a half-sib pedigree segregating for slow and fast greying demonstrated that slow-greying horses carry chromosomes with two tandem copies of the 4.6 kb STX17 sequence, whereas fast greying horses carry a tandem triplication ( 14 ). Based on this result, a new nomenclature for the Grey locus has been established as follows: G1 – one copy, wild type; G2 – two copies, slow greying; G3 – three copies, fast greying ( Figure 2b ).…”

“…Thus, there is a remarkable dosage effect of the Grey mutation, increasing the copy number of the 4.6 kb STX17 sequence shows a strong positive correlation with both speed of greying and incidence of melanoma ( Table 1 ). It may be a threshold effect as regards the predisposition to melanoma, since none out of 25 G1/G2 horses 15 years of age or older showed melanoma, in contrast to a >50% incidence amongst G1/G3 heterozygotes of the same age ( 14 ).…”

“… * Genotype–phenotype relationships are uncertain since the phenotype of only one G2/G2 homozygote and one G2/G3 heterozygote has been reported so far ( 14 ). …”