Melanocortin 1 receptor (MC1R) polymorphisms’ influence on size and dermoscopic features of nevi

The melanocortin 1 receptor (MC1R) is a highly polymorphic gene. The loss‐of‐function MC1R variants (“R”) have been strongly associated with red hair color phenotype and an increased melanoma risk. We sequenced the MC1R gene in 175 healthy individuals to assess the influence of MC1R on nevus phenotype. We identified that MC1R variant carriers had larger nevi both on the back [p‐value = .016, adjusted for multiple parameters (adj. p‐value)] and on the upper limbs (adj. p‐value = .007). Specifically, we identified a positive association between the “R” MC1R variants and visible vessels in nevi [p‐value = .033, corrected using the FDR method for multiple comparisons (corrected p‐value)], dots and globules in nevi (corrected p‐value = .033), nevi with eccentric hyperpigmentation (corrected p‐value = .033), a high degree of freckling (adj. p‐value = .019), and an associative trend with presence of blue nevi (corrected p‐value = .120). In conclusion, the MC1R gene appears to influence the nevus phenotype.     

Mutations in the melanocortin 1 receptor (MC1R) gene are associated with coat colours in the domestic rabbit (Oryctolagus cuniculus)

We sequenced almost the complete coding region of the MC1R gene in several domestic rabbits (Oryctolagus cuniculus) and identified four alleles: two wild-type alleles differing by two synonymous single nucleotide polymorphisms (c.333A>G;c.555T>C), one allele with a 30-nucleotide in-frame deletion (c.304_333del30) and one allele with a 6-nucleotide in-frame deletion (c.280_285del6). A polymerase chain reaction-based protocol was used to distinguish the wild-type alleles from the other two alleles in 263 rabbits belonging to 37 breeds or strains. All red/fawn/yellow rabbits were homozygous for the c.304_333del30 allele. This allele represents the recessive e allele at the extension locus identified through pioneering genetic studies in this species. All Californian, Checkered, Giant White and New Zealand White rabbits were homozygous for allele c.280_285del6, which was also observed in the heterozygous condition in a few other breeds. Black coat colour is part of the standard colour in Californian and Checkered breeds, in contrast to the two albino breeds, Giant White and New Zealand White. Following the nomenclature established for the rabbit extension locus, the c.280_285del6 allele, which is dominant over c.304_333del30, may be allele E(D) or allele E(S).     

Differential expression of MC1R gene in Liaoning Cashmere goats with different coat colors

Melanocyte stimulating hormone receptor (MC1R) has been known as a regulator of eumelanin and phaeomelanin production in the melanocytes, and MC1R mutations causing coat color changes are known in many vertebrates; however, there are no research reports about the differentially expression of MC1R gene and its coding protein in Cashmere goats with different coat color. We examined the presence of MC1R distribution and MC1R protein and gene expression in the white Cashmere goats and black Cashmere goats, respectively; q-PCR, Western blot and immunhistochemical analysis showed that the expression of the MC1R gene in the black Cashmere goats was 3.39 fold more than the white ones (p?<?0.01), and Cashmere goats with black genotype had significantly higher (2.03, p?<?0.01) MC1R protein expression than white genotype in the all investigated samples. Moreover, all Cashmere goats with different coat color available for immunhistochemical analysis showed either lower (white Cashmere goats) or higher (black Cashmere goats) expression of the MC1R protein; these findings suggested that it had a relationship between the MC1R and the coat color of Cashmere goats. That could lay the foundation for the further research of the MC1R and coat color controllability regulation of the Cashmere goats.     

Melanocortin 1-receptor (MC1R) mutations are associated with plumage colour in chicken

The co-segregation of plumage colour and sequence polymorphism in the melanocortin 1-receptor gene (MC1R) was investigated using an intercross between the red junglefowl and White Leghorn chickens. The results provided compelling evidence that the Extended black (E) locus controlling plumage colour is equivalent to MC1R. E/MC1R was assigned to chromosome 11 with overwhelming statistical support. Sequence analysis indicated that the E92K substitution, causing a constitutively active receptor in the sombre mouse, is the most likely causative mutation for the Extended black allele carried by the White Leghorn founders in this intercross. The MC1R sequence associated with the recessive buttercup (ebc) allele indicated that this allele evolved from a dominant Extended black allele as it shared the E92K and M71T substitutions with some E alleles. It also carried a third missense mutation H215P which thus may interfere with the constitutive activation of the receptor caused by E92K (and possibly M71T).     

Unmelanized plumage patterns in Old World leaf warblers do not correspond to sequence variation at the melanocortin-1 receptor locus (MC1R)

Evolutionary changes in patterns and coloration of plumage are likely to represent a major mechanism for speciation among birds, yet the molecular basis for such changes remains poorly understood. Recently much attention has focused on the melanocortin-1 receptor (MC1R) as a candidate locus for determining the level and extent of epidermal melanin deposition. We tested the hypothesis that MC1R sequence variation is associated with interspecific variation in unmelanized plumage pattern elements in Old World leaf warblers (genus Phylloscopus). This genus is characterized by a variety of plumage patterns that nonetheless vary along similar lines. Species vary in the presence or absence of pale (unmelanized) pattern elements against a dark background, and these patterns are used in species recognition and courtship. We sequenced most of the MC1R coding region for eight Phylloscopus species, representing the full range of plumage patterns found in this genus. Although MC1R sequence varied among species, this variation was not related to melanin-based plumage variation. Rather, evolution of this locus in these birds appears to be conservative. Ratios of nonsynonymous to synonymous substitutions (dN/dS) were consistently low, suggesting that strong purifying selection has operated at this locus, and likelihood ratio testing revealed no evidence of variable selective pressures among lineages or across codons. Adaptive evolution at MC1R may be constrained by the adaptive importance of plumage pattern elements in this genus.     

棕背伯劳羽色多态与MC1R基因的相关性

黑素皮质素受体1 (melanocortin-1 receptor, MC1R)基因是控制动物黑色素合成的重要基因, 鸟类羽色的变异与MC1R基因的变异有密切关系。棕背伯劳(Lanius schach)在我国东部沿海多地存在羽色多态现象, 有棕色型、黑色型和黑色白边型的分化。为了探究MC1R基因与棕背伯劳色型分化的关系, 本研究对分布于广东省的3种色型共计11只棕背伯劳的MC1R基因编码区进行单核苷酸多态性(SNPs)分析和氨基酸多态性分析。结果表明: (1) 11个实验个体的MC1R基因序列共有4种单倍型, 其中黑色型和黑色白边型共享单倍型H3。(2) 3种色型棕背伯劳MC1R基因编码区的第34-931位的899个碱基中共有47个碱基变异位点, 相对应的氨基酸序列共有18个变异位点, 这些变异位点与黑色表型无对应关系。(3)黑色型与黑色白边型个体基因型在第268-303位编码区出现了36个碱基的缺失, 对应着12个氨基酸的缺失, 该缺失与黑色表型相对应。因此推测棕背伯劳的黑化与MC1R基因碱基片段的缺失密切相关。     

3个猪品种黑素皮质素受体1(MC1R)基因变异研究

利用测序、PCR-RFLP和PCR-SSCP等技术对杜洛克、长白、大白猪MC1R基因进行研究发现了5个多态位点。其中,668位点G→C突变发生在5′UTR,其余4个多态位点nt894insCC(894位点CC插入),1318C→T,1554G→A和1197G→A发生在编码区。nt894insCC导致编码蛋白过早终止。1318C→T,1554G→A和1197G→A突变分别导致a164Val,Ala243Thr和Asp124Asn氨基酸的改变。所有长白、大白猪个体在894位点均存在CC插入,其余多态位点基因型分别为668GG,1197AA,1318CC,1554GG。所有杜洛克个体在894位点均不存在CC插入,其余多态位点基因型分别为668CC,1197GG,1318TT,1554AA。所有突变位点无杂合子出现。由此可以推测,668G→C,1318C→T和1554G→A可能与杜洛克的红毛色存在相关,导致1197G→A突变无意义的894位点CC插入可能与长白、大白猪白毛色存在相关。     

Missense SNP of the MC1R gene is associated with plumage variation in the Gyrfalcon (Falco rusticolus)

A single nucleotide polymorphism (MC1R: c.376A>G) in the MC1R gene was found to be highly correlated with pigment phenotype in the Gyrfalcon. Homozygous genotypes c.376GG and c.376AA were found to dominate the extreme white and dark plumage types respectively, and heterozygotes occurred mainly in intermediate phenotypes. However, some heterozygotes were associated with extreme phenotypes, indicating that melanism/albinism might also involve other loci.     

A premature stop codon in the TYRP1 gene is associated with brown coat colour in the European rabbit (Oryctolagus cuniculus)

Classical genetic studies in European rabbits (Oryctolagus cuniculus) suggested the presence of two alleles at the brown coat colour locus: a wild‐type B allele that gives dense black pigment throughout the coat and a recessive b allele that in the homozygous condition (b/b genotype) produces brown rabbits that are unable to develop black pigmentation. In several other species, this locus is determined by mutations in the tyrosinase‐related protein 1 (TYRP1) gene, encoding a melanocyte enzyme needed for the production of dark eumelanin. In this study, we investigated the rabbit TYRP1 gene as a strong candidate for the rabbit brown coat colour locus. A total of 3846 bp of the TYRP1 gene were sequenced in eight rabbits of different breeds and identified 23 single nucleotide polymorphisms (SNPs; 12 in intronic regions, five in exons and six in the 3′‐untranslated region) and an insertion/deletion of 13 bp, in the 3′‐untranslated region, organised in a few haplotypes. A mutation in exon 2 (g.41360196G>A) leads to a premature stop codon at position 190 of the deduced amino acid sequence (p.Trp190ter). Therefore, translation predicts a truncated TYRP1 protein lacking almost completely the tyrosinase domain. Genotyping 203 rabbits of 32 different breeds identified this mutation only in brown Havana rabbits. Its potential functional relevance in disrupting the TYRP1 protein and its presence only in brown animals strongly argue for this non‐sense mutation being a causative mutation for the recessive b allele at the brown locus in Oryctolagus cuniculus.     

Melanism in guinea fowl (Numida meleagris) is associated with a deletion of Phenylalanine‐256 in the MC1R gene

We have characterized a deletion in the MC1R gene causing the loss of one amino acid (p.Phe256del), which is perfectly associated with melanism in guinea fowl (Numida meleagris). Co‐segregation of the p.Phe256del with melanism was confirmed in 25 offspring born from a cross of two heterozygote birds; therefore we suggest that this mutation is responsible for the black phenotype. Interestingly, this is the first case of recessive melanism linked to MC1R.     

猪黑素皮质素受体1(MC1R)基因与毛色表型的研究

猪的毛色表型虽然与经济性状没有直接相关,但它却对经济效益产生重要影响,在猪育种实践、商品猪生产等方面都有应用。结合PCR—AccⅡ—RFIP、PCR—BspH I—RFLP及PCR-SSCP技术,分析了16个全同胞家系和金华猪、嘉兴黑猪、玉山黑猪、乐平花猪、上高两头乌猪及嵊县花猪等6个地方猪种随机采样个体的黑素皮质素受体1(MCIR)基因型。结果显示,地方猪种在MCIR位点携带高频率的显性黑等位基因E^DI,表明我国地方猪种的黑毛色可能主要由显性黑等位基因E^DI调控。通过对嵊县花猪MCIR位点的分析,首次发现PCR-SSCP证据的新序列,与已知的其他5个等位基因带型不同。家系个体的分析结果进一步验证了E^DI对E^p、e为完全显性,E^p对e为不完全显性。     

High diversity in functional properties of melanocortin 1 receptor (MC1R) in divergent primate species is more strongly associated with phylogeny than coat color

We have characterized the biochemical function of the melanocortin 1 receptor (MC1R), a critical regulator of melanin synthesis, from 9 phylogenetically diverse primate species with varying coat colors. There is substantial diversity in melanocyte-stimulating hormone (MSH) binding affinity and basal levels of activity in the cloned MC1Rs. MSH binding was lost independently in lemur and New World monkey lineages, whereas high basal levels of MC1R activity occur in lemurs and some New World monkeys and Old World monkeys. Highest levels of basal activity were found in the MC1R of ruffed lemurs, which have the E94K mutation that leads to constitutive activation in other species. In 3 species (2 lemurs and the howler monkey), we report the novel finding that binding and inhibition of MC1R by agouti signaling protein (ASIP) can occur when MSH binding has been lost, thus enabling continuing regulation of the melanin type via ASIP expression. Together, these findings can explain the previous paradox of a predominantly pheomelanic coat in the red ruffed lemur (Varecia rubra). The presence of a functional, MSH-responsive MC1R in orangutan demonstrates that the mechanism of red hair generation in this ape is different from the prevalent mechanism in European human populations. Overall, we have found unexpected diversity in MC1R function among primates and show that the evolution of the regulatory control of MC1R activity occurs by independent variation of 3 distinct mechanisms: basal MC1R activity, MSH binding and activation, and ASIP binding and inhibition. This diversity of function is broadly associated with primate phylogeny and does not have a simple relation to coat color phenotype within primate clades.     

Melanocortin‐1 receptor (MC1R) genotypes do not correlate with size in two cohorts of medium‐to‐giant congenital melanocytic nevi

Congenital melanocytic nevi (CMN) are cutaneous malformations whose prevalence is inversely correlated with projected adult size. CMN are caused by somatic mutations, but epidemiological studies suggest that germline genetic factors may influence CMN development. In CMN patients from the U.K., genetic variants in MC1R, such as p.V92M and loss‐of‐function variants, have been previously associated with larger CMN. We analyzed the association of MC1R variants with CMN characteristics in two distinct cohorts of medium‐to‐giant CMN patients from Spain (N = 113) and from France, Norway, Canada, and the United States (N = 53), similar at the clinical and phenotypical level except for the number of nevi per patient. We found that the p.V92M or loss‐of‐function MC1R variants either alone or in combination did not correlate with CMN size, in contrast to the U.K. CMN patients. An additional case–control analysis with 259 unaffected Spanish individuals showed a higher frequency of MC1R compound heterozygous or homozygous variant genotypes in Spanish CMN patients compared to the control population (15.9% vs. 9.3%; p = .075). Altogether, this study suggests that MC1R variants are not associated with CMN size in these non‐UK cohorts. Additional studies are required to define the potential role of MC1R as a risk factor in CMN development.     

Association of melanocortin 4 receptor (MC4R) and high mobility group AT-hook 1 (HMGA1) polymorphisms with pig growth and fat deposition traits

The aim of this study was to analyse the combined effect of melanocortin 4 receptor (MC4R) and high mobility group AT-hook 1 (HMGA1) polymorphisms on growth and fatness traits in Duroc pigs. No significant interaction was observed between MC4R and HMGA1 for back-fat traits. An additive mode of inheritance of both gene effects was found for average daily gain and lean meat content. Maximum mean differences from combined genotypic effects were over 2 mm for back fat, 70 g/day for average daily gain and 2% for lean meat content. Therefore, utilization of polymorphisms in both MC4R and HMGA1 for marker-assisted selection could result in an economic benefit to the pig industry.     

Association of a Glu92Lys substitution in MC1R with extended brown in Japanese quail (Coturnix japonica)

We investigated melanocortin 1 receptor (MC1R) as a candidate locus for the Extended brown phenotype in quail, in which there is a general darkening throughout the plumage. An initial screen of variation in MC1R in Extended brown and in wild-type quails revealed two polymorphic non-synonymous sites. One of these sites, a G-to-A substitution leading to a Glu92Lys mutation, was perfectly associated with plumage phenotype; all Extended brown birds were homozygous for Lys92. Co-segregation of the Glu92Lys mutation with the Extended brown phenotype was confirmed in 24 progeny of an E/e(+) x E/e(+) cross. Glu92Lys is likely to be the causative mutation for the increased melanism in Extended brown, given that the same mutation is associated with melanic plumage in many breeds of domestic chicken, as well as in a wild passerine bird (the bananaquit, Coereba flaveola) and laboratory mice. Interestingly, the increase in melanization with the Glu92Lys mutation is less marked in quails than in most other birds and mammals. Phylogenetic results indicate that the Glu92Lys mutation has independently occurred in quail and chicken lineages.     

Molecular cloning and functional characterization of the canine parathyroid hormone/parathyroid hormone related peptide receptor (PTH1)

Parathyroid hormone (PTH) is a major mediator of calcium and phosphate metabolism through its interactions with receptors in kidney and bone. PTH binds with high affinity to PTH1 and PTH2, members of the superfamily of G protein-coupled receptors. In order to clone the canine PTH1 receptor, a canine kidney cDNA library was screened using the human PTH1 receptor cDNA and two clones were further characterized. The longest clone was 2177 bp and contained a single open reading frame of 1785 bp, potentially encoding a protein of 595 amino acids with a predicted molecular weight of 66.4 kD. This open reading frame exhibits >91% identity to the human PTH1 receptor cDNA and >95% identity when the putative canine and human protein sequences are compared. Competition binding following transfection of the canine PTH1 receptor into CHO cells demonstrated specific displacement of ¹²⁵I-human PTH 1-34 by canine PTH 1-34, human PTH 1-34, and canine/human parathyroid hormone related peptide (PTHrP) 1-34. Treatment of canine PTH1 receptor transfected cells, but not mock transfected cells, with these ligands also resulted in increased levels of intracellular cAMP. In contrast, the non-related aldosterone secretion inhibiting factor 1-35 neither bound nor activated the canine PTH1 receptor. Northern blot analysis revealed high levels of PTH1 receptor mRNA in the kidney, with much lower, but detectable, levels in aorta, heart, lung, prostate, testis, and skeletal muscle. Together, these data indicate that we have cloned the canine PTH1 receptor and that it is very similar, both in sequence and in functional characteristics, to the other known PTH1 receptors.     

Discovery of a new nucleotide substitution in the MC1R gene and haplotype distribution in native and non-Japanese chicken breeds

Melanocortin 1-receptor (MC1R) is one of the major genes that controls chicken plumage colour. In this study, we investigated the sequence and haplotype distribution of the MC1R gene in native Japanese chickens, along with non-Japanese chicken breeds. In total, 732 and 155 chickens from 30 Japanese and eight non-Japanese breeds respectively were used. Three synonymous and 11 non-synonymous nucleotide substitutions were detected, resulting in 15 haplotypes (H0–H14). Of these, three were newly found haplotypes (H9, H13 and H14), of which one (H9) was composed of known substitutions C69T, T212C, G274A and G636A. The second one (H13) possessed newly found non-synonymous substitution C919G, apart from the known substitutions C69T, G178A, G274A, G636A and T637C. The third one (H14) comprised a newly discovered substitution C919G in addition to the known C69T, G274A and G409A substitutions. The homozygote for this new haplotype exhibited wt like plumage despite the presence of G274A. In addition to discovering a new nucleotide substitution (C919G) and three new haplotypes, we defined the plumage colour of the bird that was homozygous for the A644C substitution (H5 haplotype) as wheaten-like for the first time; although the substitution has been already reported, its effect was not revealed. Besides detecting the new plumage colour, we also confirmed that the A427G and G274A substitutions contribute in expressing brownish and black plumage colour respectively, as reported by the previous studies. Moreover, we confirmed that the buttercup allele does not express black plumage despite possessing a G274A substitution, under the suppression effect of A644C. In contrast, the birds homozygous for the birchen allele presented solid black plumage, which was contradictory to the previous reports. In conclusion, we revealed a large diversity in the MC1R gene of native Japanese chicken breeds, along with the discovery of a new non-synonymous nucleotide substitution (C919G) and three novel haplotypes (H9, H13 and H14).