Download Molecular tests for coat colours in horses

J. Anim. Breed. Genet. ISSN 0931-2668
REVIEW ARTICLE
Molecular tests for coat colours in horses
Stefan Rieder
Swiss College of Agriculture, Zollikofen BE, Switzerland
Keywords
Behavior; breed diversity; color genetics;
domestication; horse; molecular tests.
Correspondence
Stefan Rieder, Swiss College of Agriculture,
Länggasse 85, 3052 Zollikofen BE, Switzerland.
Tel: +41 31 910 22 65;
Fax: +41 31 910 22 88;
E-mail: [email protected]
Received: 9 January 2009;
accepted: 29 June 2009
Summary
Colour phenotypes may have played a major role during early domestication events and initial selection among domestic animal species. As
coat colours mostly follow a relatively simple mode of Mendelian inheritance, they have been among the first traits to be systematically analysed at the molecular level. As a result of the number of genetic tools
developed during the past decade, horse coat colour tests have been
designed and are now commercially available for some of the basic phenotypes. These tests enable breeders to verify segregation within particular pedigrees, to select specific colour phenotypes according to market
demand or studbook policies and to avoid complex inherited diseases
associated with some of the colour patterns. This paper reviews the relevance of the topic, describes all currently available tests for coat colours
in horses and addresses also ongoing research in this field.
General introduction and relevance of the topic
Colour phenotypes may have played a major role
during early domestication events and initial selection among domestic animal species (Henner et al.
2002a; Bruford et al. 2003; Andersson & Georges
2004). Domestication is a process resulting in
numerous morphological, physiological and behavioural changes. Among these, coat colour seems to be
a particularly obvious one (Grandin 1998). While
many wild animal species are coloured relatively
uniformly, domestic stock like, e.g. horses (Figure 1)
show a broad variety of coat colour patterns. This
seems to be at least partly due to differences in selection criteria for animals in the wild as opposed to
captive breeding populations. Very recently, Ludwig
et al. (2009) published a study on the occurrence
and frequency of coat colour alleles found in ancient
horse DNA samples from the Pleistocene Period to
Roman times. They found less colour variation in
ancient wild horse populations than in early and
later domesticated horse populations. Although a
particular coat colour might be essential for survival
in nature (e.g. camouflage, mating behaviour,
pathogen tolerance and environmental adaptation)
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(Gilbert 2006), human preferences and demand
might have favoured rare alleles or led to previously
unknown phenotypes due to selective breeding
(Klungland & Vage 2000). Recently, Fang et al.
(2009) presented convincing results on how humanmediated selection acted on colour phenotypes in
the pig.
As coat colours appear to follow relatively simple
modes of Mendelian inheritance most of the time
(Table 1), they were among the first traits to be systematically analysed at the molecular level. Furthermore, they provide unique models for studying gene
function and regulation, i.e. models for investigating
the relations between phenotypic variation, particular genotypes and physiological processes.
Genes affecting mammalian coat and skin colour
can be classified into two main groups: those acting on
pigment synthesis and those acting on the pigmentproducing cells, the melanocytes (Nordlund et al.
1998). Variation in coat and skin colours is therefore
likely to be understood as the effect of modified genes
causing changes to either pigment synthesis or to the
melanocyte or its combinations (constitutive pigmentation – intrinsic), supplemented by environmental
factors and hormones (facultative pigmentation –
doi:10.1111/j.1439-0388.2009.00832.x
Molecular tests for coat colours in horses
S. Rieder
Figure 1 Coat colour variation in the horse. Phenotypes and known genotypes (some of the pictures were kindly provided by the Swiss National
Stud Avenches). A hyphen (-) denominates that any of the known alleles may be present.
inducible) (Graphodatskaya 2002). Generally, mutations in the external or surface environment of the
melanocyte result in changes in the amount of pheomelanin (red ⁄ yellow pigment) versus eumelanin
(black ⁄ brown pigment), thus addressing the question
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of what kind of pigment are produced. Moreover,
changes in the amount of pigment produced are
caused by mutations affecting the internal machinery of the melanocyte, an effect which may lead to
coat colour dilution. Last but not the least, other
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Molecular tests for coat colours in horses
S. Rieder
Table 1 An overview of the genetics of horse coat colours
Phenotype
Basic coat colours
Chestnut ⁄ Sorrel
Locus name,
symbol and
gene abbreviation
Extension (E)
MC1R
Alleles and mutation types
e⁄e
C901T missense mutation;
loss-of-function mutation
E ⁄ e and EE
A ⁄ a and AA
Mode of inheritance, interaction,
association
Autosomal recessive
Epistatic over black
Bay
Extension (E)
Agouti (A)
Black
Agouti (A)
ASIP
a⁄a
11bp deletion at position
2174–2184; frameshift
loss-of-function mutation
Cream (CR)
MATP
CR ⁄ cr
G457A missense mutation
Cream (CR)
MATP
CR ⁄ CR
Dun (D)
Not known
Silver (Z)
PMEL17
D ⁄ d and D ⁄ D
QTL on ECA8 close to microsatellite
UCDEQ46
Z ⁄ z and ZZ
C1457T missense mutation
Champagne (CH)
SLC36A1
CH ⁄ ch and CH ⁄ CH
C188G missense mutation
Grey (G)
STX17
G ⁄ g and G ⁄ G
Intron 6 duplication with regulatory
consequences
Autosomal dominant
Epistatic to all colours; higher
incidence of melanoma in carriers
of loss-of-function mutation in ASIP
Progression of greying accelerated
in homozygous G ⁄ G horses
Roan (RN)
Associated to KIT
RN ⁄ rn and RN ⁄ RN
C2613T silent mutation
Overo-spotting – irregular white spotting,
often horizontal distribution
Overo (O)
EDNRB
O ⁄ o and O ⁄ O
TC353-354AG dinucleotide missense
mutation
Tobiano-spotting – regular white spotting,
often vertical distribution
Tobiano (TO)
KIT
TO ⁄ to and TO ⁄ TO
Inversion on ECA3q near KIT
Autosomal dominant
Epistatic to all colours
The homozygous RN ⁄ RN genotype is
thought to be lethal
Autosomal dominant
Epistatic to all colours
Homozygous O ⁄ O carriers are socalled lethal whites (OLWS)
OLWS follows a recessive mode of
inheritance
Autosomal dominant
Epistatic to all colours
Dilution coat colours
Palomino ⁄ Buckskin
Cremello ⁄ Perlino – creamy white coat
colour
with blue eyes
Dun – diluted basic colours and primitive
markings (e.g. dorsal stripe, zebra stripes
on legs)
Silver dapple – chocolate-to-reddish body
with white ⁄ grey mane and tail
Champagne – metallic sheen (eye and
skin colour may change with age)
Greying
Progressive greying with age; associated
with melanoma
Depigmentation
Roan – interspersed white hairs in basic
colour
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Autosomal dominant
Result of allele combinations other
than e ⁄ e at MC1R and a ⁄ a at ASIP
Autosomal recessive
Autosomal codominant
Epistatic to chestnut ⁄ sorrel and bay;
no or poor epistatic effect on black
Autosomal codominant
Epistatic effect on black in homozy
gous horses
Autosomal dominant
Epistatic to all basic colours
Autosomal dominant
Epistatic over bay and black
(eumelanin); no or poor epistatic
effect on chestnut ⁄ sorrel
(pheomelanin)
In some breeds Silver is part of a
linkage group on ECA6q harbouring
a QTL for multiple congenital ocular
anomalies (MCOA)
Autosomal dominant
Epistatic to all basic colours
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Molecular tests for coat colours in horses
S. Rieder
Table 1 (Continued)
Phenotype
Locus name,
symbol and
gene abbreviation
Alleles and mutation types
Appaloosa-spotting or Leopard-spotting;
associated with congenital stationary night
blindness (CSNB)
Leopard spotting
(LP)
Associated to
TRPM1 and yet
unknown
modifiers
LP ⁄ lp and LP ⁄ LP
Downregulation of TRPM1
White – heterogenous amount of
depigmentation
White (W)
KIT
W ⁄ w and W ⁄ W
Various independent, breed-specific
mutations in KIT
Sabino-spotting
Sabino (SB)
KIT
SB ⁄ sb and SB ⁄ SB
KI16 + 1037A base substitution T
with A – skipping of exon 17
(Sabino 1 – SB1)
White facial and leg markings
White markings
(WM)
Associated to
MC1R, KIT, MITF
WM ⁄ wm and wm ⁄ wm
1. QTL on ECA3q close to microsatel
lite AHT101 and SNP MSKITI7;
2. QTL on ECA16q
Mode of inheritance, interaction,
association
Autosomal – incompletely –
dominant
Epistatic to all colours
Homozygous LP ⁄ LP carriers tend to
show fewer pigmentation than
heterozygous horses; homozygous
horses show CSNB
Autosomal dominant
Epistatic to all colours
The homozygous W ⁄ W genotype is
thought to be lethal
Autosomal dominant
Epistatic to all colours
Homozygosity for SB1 results in a
complete or nearly completely white
phenotype
Autosomal recessive
Epistatic to chestnut ⁄ sorrel
Depending on basic colours, either
KIT or MITF primarily act on white
markings
For all listed alleles commercial genotyping is possible, except, so far, for Leopard-spotting and White markings. For Dun and Roan zygosity genotyping is possible for some breeds. Definition of mutation types: 1 = missense mutation (non-synonymous mutation) – a nucleotide change (transition, transversion ⁄ reversion) resulting in a codon that codes for a different amino acid. A silent mutation (synonymous mutation) – a nucleotide
change not affecting the amino acid code of a protein. 2 = frameshift mutation – result of insertions or deletions (indels) of nucleotides that are
not evenly divisible by three. Due to the triplet nature of gene expression by codons, the insertion or deletion disrupts the reading frame.
3 = inversion – an inversion is a chromosome rearrangement in which a segment of a chromosome is reversed. 4 = alternative splicing – alternative splicing is the RNA splicing variation mechanism in which the exons of the primary gene transcript, the pre-mRNA, are separated and reconnected to produce alternative ribonucleotide arrangements. Different modes of alternative splicing are distinguished, inter alia exon skipping.
loci, controlling differentiation, proliferation and
migration of melanocytes determine the amount and
form of white spotting (Sponenberg 2009).
Melanocytes belong to the group of neural crestderived cells. It is known that genes acting on melanocyte development and distribution also act on cells
such as primordial germ cells, haematopoietic cells
or neurones. Furthermore, enzymes and hormones
acting on pigment synthesis and regulation (e.g.
tyrosinase and melanocortins) are involved in a
number of physiological processes (Graphodatskaya
2002). These fascinating coherences explain some of
the pleiotropic effects between coat colour variation
and more complex traits such as developmental disorders (e.g. lethal dominant white), diseases (e.g.
lethal white foal disease and grey horse melanoma)
or even temperament (Trut 1999; Dobney & Larson
2006; Gilbert 2006). They might also be a key to the
question why breeders in different parts of the world
have often associated specific coat colours with particular performance characteristics in horses (fast
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and hot chestnuts, hardy bays, gentle greys, etc.).
However, this latter facet is discussed controversially
(Stachurska et al. 2007).
Today, coat colour phenotypes (especially colour
markings) are of practical relevance concerning the
identification of individual horses. Coat colours give
a first hint whether alleles between parents and offspring segregated correctly or not (e.g. grey rule – a
grey horse has at least one grey parent; chestnut
rule – breeding chestnuts always results in chestnuts – Bowling 2000; Sponenberg 2009), and they
may enable us to distinguish among horses of
particular breeds (Camargue all grey, Friesian all
black, Haflinger all chestnut with flaxen mane and
tail, etc.).
Genes controlling horse coat colours have been
known for a long time. However, it was only
recently that functional alleles or marker alleles have
been detected at the DNA level. Among them, the
chestnut allele Ee, a single nucleotide mutation
(C901T; AF288357) within the melanocortin receptor
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S. Rieder
1 (MC1R), was the first to be published in the horse
(Marklund et al. 1996).
The following paragraphs describe, step by step,
the major phenotypic coat colour groups of the
horse and their molecular genetic background, as far
as it is known and has been published to date.
Table 1 summarizes the content of the paragraphs.
Some of the phenotypes are presented in Figure 1.
First, the genetics of the so-called basic coat colours
– chestnut, bay and black – are presented. Thereafter, dilution phenotypes are discussed, followed by a
paragraph on greying and a final paragraph on
depigmentation phenotypes.
The Extension and Agouti loci – basic coat colours,
chestnut, bay and black
Apart from white markings, chestnut horses are
characterized by eumelanin pigment (black ⁄ brown
pigment) in the skin and pheomelanin pigment
(red ⁄ yellow pigment) in the hair (including mane
and tail). By contrast, black horses have uniformly
distributed black pigmented skin and hair, and bay
horses show pheomelanin (body) and eumelanin
(mane, tale and lower legs) patterns. The Melanocortin-1-receptor (MC1R), encoded by the Extension
(E) locus (ECA3p12), and its peptide antagonist, the
agouti signalling protein (ASIP), encoded by the
Agouti (A) locus (ECA22q15–16), control the relative amounts of melanin pigments in mammals.
ASIP acts as an indirect antagonist of MC1R by nullifying the action of the a-melanocyte-stimulating
hormone (a-MSH). Loss-of-function mutation of
MC1R results in red ⁄ yellow pigment (pheomelanin),
whereas gain-of-function mutation of MC1R or lossof-function mutation of ASIP results in the production of black pigment, i.e. eumelanin (Rieder et al.
2001). In the horse, the basic colours, chestnut, bay
and black, are so far determined by four alleles, two
of them encoded by the Extension (E) locus (EE and
Ee) and two encoded by the Agouti (A) locus (AA
and Aa). Chestnut and black follow a recessive mode
of inheritance (Ee ⁄ Ee and Aa ⁄ Aa), chestnut being epistatic over black. Black is therefore only expressed
when the genotype at the Extension locus differs
from Ee ⁄ Ee. Bay is the result of allele combinations
other than Ee ⁄ Ee at the Extension locus and Aa ⁄ Aa at
the Agouti locus. So far, in the horse, no gain-offunction mutation (i.e. dominant black) at the
Extension locus is known at the DNA level. Therefore, black coat colour results from a recessive allele
at the Agouti locus only (11-bp deletion in exon2;
AF288358) (Rieder et al. 2001). Shades of these basic
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Molecular tests for coat colours in horses
colour patterns might be due to epistatic effects of
the presented alleles as well as to so far unknown
genetic variation (Henner et al. 2002a). A third allele
segregating at the Extension locus, also completely
associated with the chestnut phenotype, was published by Wagner & Reissmann (2000) – AF252541.
The Cream, Dun, Silver Dapple and Champagne
loci – coat colour dilution
There are four confirmed loci in the horse which are
responsible for dilution of the basic coat colours:
Cream (C), Dun (D), Silver Dapple (Z) and Champagne (CH). They all act basically in dominant or
codominant manner. However, the effects on the
phenotypes are somehow different, depending on
the characteristics of either the dominant or codominant alleles among heterozygote and homozygote
carriers.
To start with the CCR and DD alleles, in a heterozygote state the incomplete dominant cream allele
dilutes a chestnut background to palomino and a
bay background to buckskin. However, one CCR allele
usually has no phenotypic effect on black. In homozygote state CCR dilutes all basic colours to what is
called a cremello ⁄ perlino phenotype, also featuring
blue eyes. The cream locus was mapped to ECA21q
(Locke et al. 2001). Mariat et al. (2003) showed that
a single base substitution in exon 2 of the
membrane-associated transporter protein (MATP) –
G457A and AY187093 – appears to be fully associated with the phenotypes segregating with CCR. The
mutation seems to disrupt the function of this transporter protein, leading to the known colour dilutions.
Compared with CCR, the dun allele DD shows complete dominance on all basic coat colours; heterozygotes cannot be distinguished from homozygotes.
Furthermore, the dun coat colour (red dun, yellow
dun, grullo or mouse dun, depending on the basic
colour background) appears along with a dorsal
stripe, zebra stripes and other primitive markings.
This is, in particular not the case for the phenotypes
resulting from all other horse coat colour dilution
genes mentioned. The dun coat colour has been
linkage mapped to a group of microsatellite markers
on ECA 8 (Bricker et al. 2003). However, the protein
encoded by the Dun locus is not yet known. Based
on comparative mapping data, candidate genes
would most probably be located on HSA 18q, MMU
18q and MMU 1q.
A flaxen mane and tail as well as a diluted chocolate-to-reddish body on a eumelanin background is
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Molecular tests for coat colours in horses
known as Silver Dapple (Z). Brunberg et al. (2006)
showed that the silver phenotype segregates in an
autosomal, completely dominant manner, and that it
is fully associated with a C > T transition
(DQ855465) at position five within the exon 11 of
premelanosomal protein 17 (Pmel17), also known as
the Silver locus. The mutation changes the second
amino acid in the cytoplasmic region from arginine
to cysteine (Arg618Cys). Another research group
(Reissmann et al. 2007) detected a haplotype
(DQ665301:g.697A > T ⁄ DQ665301:g.1457C > T) in
Pmel17 which again was completely associated with
the Silver Dapple phenotype. This haplotype also
included the C > T mutation mentioned above.
Pmel17 was originally mapped to ECA6q23 by
Rieder (1999). In some breeds, the Silver locus was
recently found to be part of a linkage group harbouring a quantitative trait locus (QTL) associated
with multiple congenital ocular anomalies (Andersson et al. 2008; Grahn et al. 2008).
To complete the series of dilution coat colour loci
in horses, a fourth locus, champagne (CH), is
described. Champagne is a phenotype known for its
metallic sheen, and may be confused with cream
and silver. It is described as following an autosomal
dominant mode of inheritance. Cook et al. (2008)
mapped the locus to a region on ECA 14 between
microsatellites UM010 and TKY329. The same
authors found a missense mutation at position 76 in
exon 2 of SLC36A1 (also known as PAT1 or
LYAAT1) among several candidate genes. The cysteine to guanine base change (c188C > G) is predicted
to cause a transition from a threonine to arginine, at
amino acid 63 of the protein (T63R). The mutation
is thought to affect a putative transmembrane
domain. According to Cook et al. (2008), the protein
coded by the SLC36A1 gene might play a role in
melanosome development, resulting in a reduction
in pigmentation induced by the mutation mentioned
above.
The grey locus – progressive greying with age
Progressive greying with age (G) is a coat colour
phenotype in the horse with a dominant mode of
inheritance (GG versus Gg). A grey horse is always
born with pigmented skin and coloured hair. As it
gets older, its hair greys, while its skin remains pigmented (except for white markings and vitiligo).
Grey is epistatic to all other colours. Grey horses are
known to be particularly susceptible to skin melanoma (Rieder et al. 2000). The Grey locus has
been mapped to ECA 25q by three independent
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S. Rieder
groups, and has been fine mapped by a fourth
group; all four groups worked with different horse
breeds and resource families (Henner et al. 2002b;
Locke et al. 2002; Swinburne et al. 2002; Pielberg
et al. 2005). Depending on the particular resource
family, the Grey locus (G) was found very close to
the microsatellite marker COR080. Thus, the overall
confirmative mapping data indicate that Grey is a
rather old mutation. Candidate genes for the Grey
locus were not available from comparative mapping
data until recently, neither was the type of correlation with melanoma development (Sölkner et al.
2004). Rosengren Pielberg et al. (2008) demonstrated, that the grey phenotype is caused by a
4.6-kb duplication in intron 6 of STX17 (syntaxin17) that constitutes a cis-acting regulatory mutation.
Both STX17 and the neighbouring NR4A3 gene are
overexpressed in melanomas of grey horses. Grey
horses carrying a loss-of-function mutation in the
ASIP (agouti signalling protein) have a higher
incidence of melanoma, implying that increased
melanocortin-1 receptor signalling promotes melanoma development in grey horses. Another interesting finding of the study concerned the interpretation
of how a mutation causing loss of hair pigmentation
could also cause melanomas with a massive production of melanin. The authors suggested that this
could be due to differences in the life cycle of hair
follicles and dermal melanocytes. The STX17 duplication leads to proliferation of dermal melanocytes,
thus predisposing grey horses to melanoma development. By contrast, hyperproliferation of hair follicle
melanocytes may cause premature depletion of stem
cells, and thus progressive loss of pigmentation in
the coat.
The White and White Spotting loci – coat and
skin colour depigmentation
White and White Spotting loci are responsible for a
group of phenotypes encompassing a mixture of
white and coloured hairs (roan; head usually not
affected), discrete white spots of varying size, extent
and location (tobiano, overo, sabino, leopard spotting) to nearly complete depigmentation (dominant
white). True albinos (normal structure and number
of melanocytes, but complete depigmentation of the
eyes, skin and hair), however, are not known in the
horse so far. Phenotypic and genetic definition and
distinction of these somehow heterogeneous characters may present substantial problems.
First of all, Marklund et al. (1999) described several mutations in the Tyrosine kinase receptor (KIT)
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S. Rieder
located on ECA3q21–22. A KIT exon19 substitution
(C2613T – AJ224642–45) appeared to be strongly
associated with the roan phenotype (interspersed
white and coloured hair all over the body except the
head) in most studied breeds. However, the causative mutation for roan is not yet known. Roan is
assumed to be a lethal condition in homozygous
state and segregates in a dominant manner (RNRN)
(Hintz & Van Vleck 1979).
Yang et al. (1998), Santschi et al. (1998) and Metallinos et al. (1998) demonstrated independently
that a dinucleotide mutation (TC353–354AG –
AF019072; AF038900) in the Endothelin-B receptor
gene (EDNRB; ECA17q23–24) is associated with
lethal white foal syndrome (OLWS) and the overo
coat colour pattern. The mutation leads to an amino
acid change from lysine to isoleucine in the first
transmembrane domain of this G-protein-coupled
receptor, with profound impact on the normal development of enteric ganglia and melanocytes (similar
to Hirschsprung’s disease in humans). Lethal white
foals were found homozygous for the mutation, with
all their parents showing a heterozygous state, carrying mostly the frame overo coat colour pattern.
Some carriers showed tobiano and sabino markings
or were even ‘solid’ coloured (Santschi et al. 2001),
indicating epistatic effects and variable penetrance of
the mutant allele (OO). In overo colour patterns,
white appears irregularly and will rarely cross the
dorsal line of the horse. Legs are often solid coloured, and head markings extensively white. OLWS
has a recessive mode of inheritance, while the overo
coat colour pattern follows a dominant segregation
pattern.
Brooks et al. (2002) published a PCR–RFLP
detected in the KIT gene (intron13; C693G –
AY048669) which is very closely associated with a
white spotting colour known as Tobiano (TOTO). In
2007, the same group showed a chromosomal inversion, beginning approximately 100 kb downstream
of the KIT gene on ECA3q21–22, to be the causative
mutation for the (TOTO) phenotype. The inversion
does not interrupt any annotated genes but may disrupt regulatory sequences for the KIT gene and
cause the white spotting pattern (Brooks et al. 2007;
Haase et al. 2008). Tobiano is characterized by white,
depigmented areas with distinct borders that cross
the spine and usually include all four legs. Head
markings appear to be like those in common solidcoloured horses.
Another spotting pattern is referred to as Leopard
spotting (LP), also known as Appaloosa spotting.
Horses that inherit the LPLP allele will display one of
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several different patterns of white, LPLP being incompletely dominant to the solid (non-spotted) allele
LPlp. Modifiers are likely to be responsible for the
multiplicity of patterns associated with Appaloosa.
KIT, its ligand the Mast cell growth factor (MGF)
and the Microphthalmia-associated transcription factor (MITF) have been excluded as candidates for this
coat colour by linkage analysis (Terry et al. 2001,
2002). The Leopard spotting locus was, however,
assigned to ECA1q (Terry et al. 2004), a chromosomal region containing potential candidate genes
that based on comparative mapping data are most
probably responsible for LP in the horse. Among
these are the Pink eye dilution (p) and the Transient
receptor potential cation channel subfamily M,
member 1 (TRPM1), also known as Melastatin
(MLSN1). Homozygosity for LP seems to be directly
associated with congenital stationary night blindness
(CSNB) in Appaloosa horses. Bellone et al. (2008)
demonstrated recently that LP ⁄ CSNB is the result of
a downregulation of TRPM1 expression in the skin
and retina of LP ⁄ lp and LP ⁄ LP horses. TRP proteins
are thought to play a role in controlling intracellular
Ca2+ concentration. Decreased expression of TRPM1
in the eye and skin may alter bipolar cell signalling
as well as melanocyte function, thus causing both
CSNB and LP in horses.
A final series of white pattern phenotypes concern
the Dominant white (W) and Dominant Sabino loci
(SB). Both loci have recently been mapped to
ECA3q22 and association was found with mutations
in the KIT gene. A single nucleotide polymorphism
(SNP) detected in KIT intron3 (G258A – AY232413)
revealed linkage with microsatellite ASB23 and WW
in a pedigree of Swiss Franches Montagnes horses
(Mau et al. 2004). The dominant white phenotype
was traced back to one particular founder mare in
this pedigree, indicating a spontaneous mutation
event. Dominant white is thought to be lethal in its
homozygous state, maybe as a consequence of a disordered development within the haematopoietic system (Pulos & Hutt 1969). Not all dominant white
horses in the pedigree were of an entirely white
phenotype; some of them appeared spotted, as Sabino
lookalikes. So far, it is not known to what extent
these two heterogeneous phenotypes share a common genetic and phenotypic background, apart from
their association with the KIT gene. Haase et al.
(2007, 2009a) published a study on allelic heterogeneity at the equine KIT locus among dominant
white horses of different breeds. The authors demonstrated that independent mutation events within the
equine KIT gene among non-related horse breeds
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Molecular tests for coat colours in horses
resulted in similar dominant white ⁄ sabino phenotypes.
An SNP in the KIT intron16 (KI16 + 1037A;
AY910688) was found to be responsible for skipping
exon17 and to be causative for the Sabino spotting
pattern (called Sabino 1) in the Tennessee Walking
Horse and in some other breeds (Brooks & Bailey
2005). All horses carrying the Sabino1 KIT intron16
SNP exhibited Sabino phenotypes, including homozygous all-white animals. However, not all Sabino
lookalike horses carried the Sabino1 SNP. Thus, further research is needed to unravel the relations
among those coat colour patterns and their impact
on more complex traits.
Apart from the depigmentation phenotypes mentioned above, a pattern known as White markings
(WM) is well known in many species, including the
horse. Rieder et al. (2008) demonstrated that white
markings are highly heritable (h2 > 50%), and that a
first QTL on ECA3q, the chromosomal region harbouring the KIT gene, could explain up to 80% of
the total heritability of the trait. A strong positive
correlation was found between the chestnut allele at
MC1R and the extent of white markings, confirming
data presented by, e.g. Woolf (1992) in several publications during the 1990s. Furthermore, the authors
revealed that the trait-increasing allele seemed to
follow a recessive mode of inheritance. Recent SNP
data revealed an additional QTL on ECA16q acting
on white markings – MITF. The data indicate that
KIT influences in particular white markings on a
chestnut background, whereas MITF seems to act
primarily on white markings on a non-chestnut
background (Haase et al. 2009b).
It is worth noting that all depigmentation phenotypes discussed in this paragraph, except overo and
leopard spotting, share an association with the KIT
gene. Thus, it would be interesting to study whether
there is a pattern evolving about the location of the
mutation and the type of colour seen for KIT mutations.
Conclusion
In 1995, a workshop was formed to foster collaboration and to share resources among scientists interested in the horse genome. Despite this relatively
late start, an impressive number of tools have been
developed during the past decade, enabling us today
to analyse the basis of horse phenotypic variation at
the genetic level (Chowdhary & Bailey 2003;
Chowdhary & Raudsepp 2008). Coat colours were
among the first traits to be studied, and as we can
422
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see, genetic tests, now commercially available, have
been designed for some of them (e.g. Royo et al.
2008). These tests allow breeders to verify segregation within particular pedigrees, to select specific colour phenotypes according to market demand or
studbook policies, and to avoid inherited diseases
associated with some of the colour patterns. However, more research is still needed to fully differentiate and precisely define the heterogeneity of horse
colour phenotypes (e.g. shades such as darker chestnut or darker bay, flaxen mane and tail, seasonal
coat colour change, the problem of phenotypes that
resemble one another but are genetically different,
etc.), their underlying genetics, and, as supposed to
some of them, the correlation to more complex traits
as described above. Additionally, it would be interesting to compare and study the insights obtained
from horse coat colour genetics with other members
of the larger equid family. For example, little is
known about the molecular basis of coat colour in
the donkey or the genetics behind zebra stripes. The
results of such research might give us new insights
into the genetics of various physiological processes,
i.e. related to early embryonic development, adaptation to particular environments, pathogen tolerance
and even performance. As colour phenotypes played
an important role during the development of modern livestock breeds, including the horse, the genetics of coat colour might also be helpful to better
understand the process of domestication (e.g. the
impact of white colour markings on behaviour, such
as temperament). To achieve these goals, a focus
should be given on the precise description of phenotypes and on the setting up of segregation and ⁄ or
case ⁄ control resource families.
Acknowledgements
I thank numerous colleagues for fruitful discussions
and continuous scientific exchange on horse and
mammalian coat colour genetics over the years.
Many thanks are due to two anonymous reviewers
for helpful comments and for carefully reading and
improving this manuscript.
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