Equine coat color genetics

Equine coat color genetics determine a horse's coat color. There are many different coat colors possible, but all colors are produced by the action of only a few genes. The simplest genetic default color of all domesticated horses can be described as either "red" or "non-red", depending on whether a gene known as the "Extension" gene is present. When no other genes are active, a "red" horse is the color popularly known as a chestnut. Black coat color occurs when the Extension gene is present, but no other genes are acting on coat color.The Agouti gene can be recognized only in "non-red" horses; it determines whether black color is uniform, creating a black horse, or limited to the extremities of the body, creating a bay horse.

Chestnut, black, and bay are considered the three "base" colors that all remaining coat color genes act upon. There are a number of dilution genes that lighten these three colors in a variety of ways, sometimes affecting skin and eyes as well as hair coat, including cream, dun, pearl, champagne and silver dapple. Genes that affect the distribution of white and pigmented coat, skin and eye color create patterns such as roan, pinto, leopard, white, and even white markings. Some of these patterns may be the result of a single gene, others may be influenced by multiple alleles. Finally the gray gene, which acts differently from other coat color genes, slowly lightens any other hair coat color to white over a period of years, without changing skin or eye color.

Fundamental concepts and terminology
Much of the modern understanding of equine coat color genetics is owed to the work of Dr. Ann T. Bowling of the University of California, Davis and of Dr. Phillip Sponenberg of Virginia Polytechnic Institute. Modern discussions of horse coat color genetics are based on the distinction between "red" and "non-red" coats, a factor determined by a single gene. More detailed discussions of coat color all refer to the differing effects of separate genes on these "base" coat colors.

Coat color alleles affect melanin, the pigment or coloring of the coat. There are two chemically distinct types of melanin: phaeomelanin, which is perceived as red to yellow color, and eumelanin, is perceived as brown to black. All coloration genes in mammals affect either the production or distribution of these two chemicals. Alleles affecting melanocytes (pigment cells) do not alter the pigment chemicals themselves but rather by acting on the placement of pigment cells produce distinct patterns of unpigmented pink skin and corresponding white hair.

Heritable characteristics are transmitted, encoded, and used through a substance called DNA, which is stored in almost every cell in an organism. DNA is organized into storage structures called chromosomes. For the most part, chromosomes come in matched sets, one chromosome from each parent. The location of a gene on a chromosome is called its locus. Alternate forms of a gene are called alleles. The terms Alleles and Modifiers are used interchangeably and describe the same concept. An allele identified with a capital letter is a dominant trait, one identified with a lower-case letter is a recessive trait. Because sex cells (sperm and ova) contain only half the usual number of chromosomes, each parent contributes one allele in each gene set to the ensuing offspring. When an individual's gene set contains two copies of the same allele, it is called homozygous for that gene. When it has two different alleles, it is heterozygous. For a recessive trait to be expressed, it must be homozygous, but a dominant trait will be expressed whether it is heterozygous or homozygous. A horse homozygous for a certain allele will always pass it on to its offspring, while a horse that is heterozygous carries two different alleles and can pass on either one.

Extension
Extension controls whether or not true black pigment (eumelanin) can be formed in the hair. True black pigment may be restricted to the points, as in a bay, or uniformly distributed in a black coat. Horses capable of producing eumelanin in the hair may have a genotype of either E/E or E/e. Horses without the ability to produce eumelanin in the hair always have the genotype e/e, and are most often chestnut or "red". The e allele is also sometimes called "red factor" and can be identified through DNA testing. Horses homozygous E/E are sometimes called "homozygous black", however depending on the color of the mate, E/E status confers no guarantee of black-coated offspring; only that no offspring will be "red".



The Extension locus is occupied by the melanocortin 1 receptor (Mc1r) gene, which encodes the eponymous protein. The MC1R protein straddles the membrane of pigment cells (melanocytes). MC1R picks up a chemical called alpha-melanocyte-stimulating hormone (α-MSH), which is produced by the body, from outside the cell. When MC1R comes into contact with α-MSH, a complex reaction is triggered inside the cell, and the melanocyte begins to produce black-brown pigment (eumelanin). Without the stimulation of α-MSH, the melanocyte produces red-yellow pigment (pheomelanin) by default.

Various mutations in the human Mc1r gene result in red hair, blond hair, fair skin, and susceptibility to sundamaged skin and melanoma. Polymorphisms of Mc1r also lead to light or red coats in mice, cattle, and dogs, among others. The Extension locus was first suggested to have a role in horse coat color determination in 1974 by Stefan Adalsteinsson. Researchers at Uppsala University, Sweden, identified a missense mutation in the Mc1r gene that resulted in a loss-of-function of the MC1R protein. Without the ability to produce a functional MC1R protein, eumelanin production could not be initiated in the melanocyte, resulting in coats devoid of true black pigment. Since horses with only one copy of the defective gene were normal, the mutation was labeled e or sometimes Ee. A single copy of the wildtype allele, which encodes a fully functional MC1R protein, is protective against the loss-of-function. The normal or wildtype allele is labeled E, or sometimes E+ or EE.

Extension Phenotypes

 * E/E (+/+, E+/E+, EE/EE) wildtype, homozygous dominant. Visually, such horses are black, seal brown, bay, buckskin, perlino or smoky cream, bay dun or grullo, silver bay or silver black. Some horses with genes for gray or white spotting patterns may also have the modifier, but the color may be hidden or overlain by the loss of pigmentation. Horses that are E/E will always pass on a functional copy of the Mc1r gene to its offspring, and will never produce offspring with the e/e genotype.
 * E/e (+/e, E+/Ee, EE/Ee) wildtype, heterozygous. Visually, the horse may also be any of the colors seen with the E/E genotype. However, they statistically will only pass on the Mc1Lr gene 50% of the time.  In addition, a recent study that compared horse genotypes to their coat color phenotypes did find a statistically significant connection that suggested that lighter bay shades were heterozygous for the Extension mutation (E/e) and darker bay shades were homozygous.
 * e/e (Ee/Ee) homozygous recessive. Visually, the horse may be any color in the "red" family: chestnut, palomino, cremello, red dun, gold champagne, gray, and so on. Paired with an e/e mate, such horses will only ever produce red-family coat colors. At birth, the skin may be pink and the eyes blue, but these traits disappear after a few days and the eyes and skin of adult red coated horses are unaffected by this allele. No health defects are associated with the e allele.



Agouti
Agouti controls the restriction of true black pigment (eumelanin) in the coat. Horses with the normal agouti gene have the genotype A/A or A/a. Horses without a normal agouti gene have the genotype a/a, and if they are capable of producing black pigment, it is uniformly distributed throughout the coat. A third option, At, restricts black pigment to a black-and-tan pattern called seal brown. This allele is recessive to A and dominant to a, such that horses with the genotype A/At appear bay, while At/At and At/a horses are seal brown in the presence of a dominant Extension allele E.

The Agouti locus is occupied by the Agouti signalling peptide (Asip) gene, which encodes the eponymous protein (ASIP). Agouti signalling peptide is a paracrine signaling molecule that competes with alpha-melanocyte stimulating hormone (α-MSH) for melanocortin 1 receptor proteins (MC1R). MC1R relies on α-MSH to halt production of red-yellow pheomelanin, and initiate production of black-brown eumelanin in its place.

In many species, successive pulses of ASIP block contact between α-MSH and MC1R, resulting in alternating production of eumelanin and pheomelanin; hairs are banded light and dark as a result. In other species, Asip is regulated such that it only occurs in certain parts of the body. The light undersides of most mammals are due to the carefully controlled action of ASIP. In mice, two mutations on Agouti are responsible for yellow coats and marked obesity, with other health defects. Additionally, the Agouti locus is the site of mutations in several species that result in black-and-tan pigmentations. In normal horses, ASIP restricts the production of eumelanin to the "points": the legs, mane, tail, ear edges, etc. In 2001, researchers discovered a recessive mutation on Asip that, when homozygous, left the horse without any ASIP. As a result, horses capable of producing true black pigment had uniformly black coats. More recently, one coat color testing lab has begun offering a test for At. Further research remains to be seen.



Agouti Phenotypes

 * A/A wildtype, homozygous. Visually, the horse may be bay, buckskin, bay dun, amber champagne, and so on, or gray, or any member of the red family. However, such a horse will never be black, grullo, and so on, nor will a homozygous A horse ever produce uniform-black offspring or seal brown offspring.
 * A/At wildtype, heterozygous. Visually indistinguishable from the homozygous A horse, such horses will also never produce a uniform-black foal.
 * A/a wildtype, heterozygous. Visually indistinguishable from the homozygous A horse. With the right partner, such horses can produce uniform-black foals.
 * At/At seal brown or black-and-tan. Visually, the horse may be chestnut or gray, but in the presence of a dominant E allele, the coat will be seal brown. Variants of seal brown include dark buckskin, perlino, seal brown dun, and sable champagne.
 * At/a indistinguishable from the homozygous seal brown. Such horses may produce uniform-black offspring.
 * a/a homozygous recessive. Visually, in the presence of a dominant E allele, the horse's coat will be a uniform black, or the related smoky black, smoky cream, grullo, classic champagne, silver black, and so on.



Dun
Dun is one of several genes that control the saturation or intensity of pigment in the coat. Dun is unique in that it is simple dominant, affects eumelanin and pheomelanin equally, and does not affect the eyes or skin. Horses with the dominant D allele (D/D or D/d genotype) exhibit hypomelanism of the body coat, while d/d horses have otherwise intense, saturated coat colors. The mane, tail, head, legs, and primitive markings are not diluted. In some breeds, zygosity for Dun can be determined with an indirect DNA test.

While the Dun locus is known to be on equine chromosome 8, its precise location, the gene and protein involved, and exact mutation are not yet known. The molecular cause behind the dun coat colors is similarly not yet understood. The associated coat colors were assigned to the Dun locus in 1974 by Stefan Adalsteinsson, separate from Cream, with the presence of dun dilution indicated by the dominant D allele. The dominant D allele is relatively rare compared to the alternative d allele, and for this reason, the dominant allele is often treated as a mutation. However, the pervasive coat color among wild equids is in fact dun, and researchers from Darwin to modern day consider dun to be the wildtype state.

Dun Phenotypes

 * D/D (+/+, D+/D+) wildtype, homozygous dominant. Visually, the horse may be bay dun, grullo, red dun, palomino dun, amber dun, gray, and so on. Such a horse will always pass on the D allele and will therefore always have dun offspring.
 * D/d (+/d, D+/Dd) wildtype, heterozygous. Visually indistinguishable from the homozygous D horse.
 * d/d (Dd/Dd) non-dun, homozygous recessive. The entire coat, barring the influence of other alleles, is a rich, saturated color. The primitive markings are no longer visible. The horse may be chestnut, bay, black, gray, palomino, and so on.

Cream
Cream is another one of the genes that control the saturation or dilution of pigment in the coat. Cream differs from Dun in that it affects the coat, skin, and eyes, and unlike Dun, is dosage dependent rather than simple dominant. Furthermore, the effects on eumelanin and pheomelanin are not equal. Horses with the homozygous recessive genotype (C/C) are not affected by cream. Heterozygotes (CCr/C) have one cream allele and one wildtype non-cream allele. Such horses, sometimes called "single-dilutes", exhibit dilution red pigment in the coat, eyes, and skin to yellow or gold, while eumelanin is largely unaffected. Homozygotes (CCr/CCr) have two cream alleles, and are sometimes called "double-dilutes." Homozygous creams exhibit strong dilution of both red and black pigment in the coat, eyes, and skin to ivory or cream. The skin is rosy-pink and the eyes are pale blue. Cream is now identifiable by DNA test.

The Cream locus is occupied by the Solute carrier family 45, member 2 (SLC45A2) gene, also called the Membrane associated transport protein or Matp gene. The Matp gene encodes a protein illustrated to have roles in melanogenesis in humans, mice, and medaka, though the specific action is not known.

Mutations in the human Matp gene result in several distinct forms of Oculocutaneous albinism, Type IV as well as normal variations in skin and hair color. Mice affected by a condition homologous to cream, called underwhite, exhibit irregularly shaped melanosomes, which are the organelles within melanocytes that directly produce pigment. The first descriptions of the dosage-dependent genetic control of the palomino coat color occurred early on in equine coat color inheritance research. However, the distinction between Dun and Cream remained poorly understood until Stefan Adalsteinsson wrote Inheritance of the palomino color in Icelandic horses in 1974. The mutation responsible, a single nucleotide polymorphism in Exon 2 resulting in an aspartic acid-to-asparagine substitution (N153D), was located and described in 2003 by a research team in France.

Cream Phenotypes

 * C/C homozygous wildtype. Visually, the horse may be any color other than the cream dilute shades of palomino, buckskin, smoky black, cremello, perlino, smoky cream, and so on.
 * CCr/C heterozygous. The colors most commonly associated with this genotype are palomino, buckskin, and smoky black, though the phenotype may vary depending on other factors. Any pheomelanin in the coat is diluted to yellow or gold, and the eyes and skin are often slightly lighter than unaffected horses.
 * CCr/CCr homozygous. The colors most commonly associated with this genotype are cremello, perlino, and smoky cream. Regardless, the coat will be cream- or ivory-colored, and the skin a rosy-pink. The eyes are pale blue.

Champagne
Champagne is a gene that controls the saturation or dilution of pigment in the coat. Unlike Cream, Champagne is not strongly dosage-dependent, and affects both types of pigment equally. Champagne differs from Dun in that it affects the color of the coat, skin, and eyes, and in that the unaffected condition is the wildtype. Horses with the dominant CH allele (CH/CH or CH/ch genotype) exhibit hypomelanism of the body coat, such that phaeomelanin is diluted to gold and eumelanin is diluted to tan. Affected horses are born with blue eyes which darken to amber, green, or light brown, and bright pink skin which acquires darker freckling with maturity. The difference in phenotype between the homozygous (CH/CH) and heterozygous (CH/ch) horse may be subtle, in that the coat of the homozygote may be a shade lighter, with less mottling. Horses with the homozygous recessive genotype (ch/ch) are not affected by champagne. Champagne is now identifiable by DNA test.

The Champagne locus is occupied by the Solute carrier family 36, member 1 (SLC36A1) gene, which encodes the Proton-coupled amino acid transporter 1 (PAT1) protein. This protein is one of many which is involved in active transport. The gene associated with the Cream coat colors is also a solute carrier, and orthologous genes in humans, mice, and other species are also linked to coat color phenotypes. The single nucleotide polymorphism responsible for the champagne phenotype is a missense mutation in exon 2, in which a C is replaced with a G, such that a threonine is replaced with arginine. This mutation was identified and described by an American research team in 2008.

Champagne Phenotypes

 * ch/ch (N/N) wildtype, homozygous recessive. Visually, the horse may be any color other than the champagne shades.
 * CH/ch (CH/N) heterozygous. The colors most commonly associated with this genotype are gold champagne, amber champagne, and classic champagne, though the exact phenotype depends on a variety of factors. At birth, the skin is bright pink and the eyes bright blue, darkening to freckled and light brown or green, respectively, with age. Both red and black pigment in the hair are also diluted.
 * CH/CH homozygous champagne. Homozygotes, which will never produce non-champagne offspring, are indistinguishable from heterozygotes except that their freckling may be sparser, and their coats a shade lighter.

Sources and external links

 * "Horse coat color tests" from the UC Davis Veterinary Genetics Lab
 * "Introduction to Coat Color Genetics" from Veterinary Genetics Laboratory, School of Veterinary Medicine, University of California, Davis. Web Site accessed January 12, 2008
 * "In the Genes." Quarter Horse News, Dec 15, 2004
 * "Horse Color Calculator" From Animal Genetics Incorporated. This creates the possible coat coloring of the offspring from the imputed color of sire and dam.