Analysing the diversity of the caprine melanocortin 1 receptor (MC1R) in goats with distinct geographic origins

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j o u r n a l ho me p ag e :w w w . e l s e v i e r . c o m / l o c a t e / s m a l l r u m r e s

Short

communication

Analysing

the

diversity

of

the

caprine

melanocortin

1 receptor

(MC1R)

in

goats

with

distinct

geographic

origins

Koray

Kırıkc¸

ı

a,1,2

_,

_Antonia

_Noce

a,1

_,

_Ali

_Zidi

a,1

_,

_Juan

_Manuel

_Serradilla

b

_,

_Juan

_Carrizosa

c

_,

Baltasar

Urrutia

c

_,

_Fabio

_Pilla

d

_,

_Mariasilvia

_D’Andrea

d

_,

_Juan

_Capote

e

_,

_Iosif

_Bizelis

f

_,

Valentin

Balteanu

g

_,

_Taina

_Figueiredo

_Cardoso

a,h

_,

_Shahin

_Eghbalsaied

i

_,

_Agueda

_Pons

j

_,

Luz

Ángela

Álvarez

k

_,

_Michele

_Pazzola

l

_,

_Giuseppe

_Massimo

_Vacca

l

_,

Gabriela

Obexer-Ruff

m

_,

_Marcel

_Amills

a,∗

a_Department_of_Animal_Genetics,_Center_for_Research_in_Agricultural_Genomics_{(CSIC-IRTA-UAB-UB),}_Campus_de_la_Universitat_Autònoma_de_Barcelona, Bellaterra,08193Spain

b_Departamento_de_Producción_Animal,_Campus_de_Rabanales,_Universidad_de_Córdoba,_14071,_Córdoba,_Spain

c_Instituto_Murciano_de_{Investigación}_y_Desarrollo_Agrario_y_Alimentario_(IMIDA)._Estación_Sericícola._La_Alberca,_Murcia,_Spain d_Dipartimento_di_Agricoltura,_Ambiente_e_Alimenti,_Università_degli_Studi_del_Molise,_Campobasso,_Italy

e_Instituto_Canario_de_{Investigaciones}_Agrarias,_La_Laguna_38108,_Tenerife,_Spain f_Department_of_Animal_Science,_Agricultural_University_of_Athens,_Athens_11855,_Greece

g_Institute_of_Life_Sciences,_University_of_Agricultural_Sciences_and_Veterinary_Medicine,_Cluj-Napoca_400372,_Romania h_CAPES_Foundation,_Ministry_of_Education_of_Brazil,_Brasilia,_DF,_Zip_Code_70.040-020,_Brazil

i_Transgenesis_Center_of_Excellence,_Isfahan_(Khorasgan)_Branch,_Islamic_Azad_University,_Isfahan,_Iran j_Unitat_de_Races_Autòctones,_Servei_de_Millora_Agrària,_{(SEMILLA-SAU),}_Son_Ferriol,_07198,_Spain k_Universidad_Nacional_de_Colombia,_Sede_Palmira,_Valle_del_Cauca,_Colombia

l_Dipartimento_di_Medicina_{Veterinaria,Università}_degli_Studi_di_Sassari,_07100,_Sassari,_Italy m_{Swissuniversities,}_Department_of_Research_&_Development,_{Efﬁngerstr.}_15,_3001,_Berne,_Switzerland

a

r

t

i

c

l

e

i

n

f

o

Articlehistory: Received27May2016 Receivedinrevisedform 23September2016 Accepted6October2016 Availableonline7October2016 Keywords: Goat Pigmentation Positiveselection Coatcolor

a

b

s

t

r

a

c

t

Inhumans,thevariabilityofthemelanocortin1receptor(MC1R)genehasbeenassociatedwith geog-raphy,beingmainlydeterminedbytheamountofexposuretosunlight.Studiesperformedinpigshave alsoevidencedtheexistenceofageographiccomponentinthedistributionofMC1Rhaplotypes, prob-ablyasaconsequenceofanancientsplitbetweenAsianandEuropeanwildboars.Herewith,wehave partiallyresequencedthecaprineMC1Rcodingregionin58goatsfromdistinctgeographiclocations i.e.Colombia,Italy,Spain,France,Greece,Romania,IranandAfrica.Theresultingdatasetwasmerged with39previouslypublishedcaprineMC1Rsequencesandamedianjoiningnetworkwasbuilt.This phylogeneticanalysisdidnotyieldanyevidenceofarelationshipbetweengeographyandtheclustering ofcaprineMC1Rsequences,aresultthatwasconﬁrmedbyperformingaManteltestwithapreviously publisheddatasetofninegoatbreeds(N=319)withavailableMC1Rgenotypes.Themajorityofcaprine MC1Rvariationwasnon-synonymous(c.676A>G,c.748G>T,c.764G>Aandc.801C>G)andpredictedto havefunctionaleffects.AnanalysisofgoatMC1RsequenceswiththePAML4softwareprovidedevidence thattwoSNPs(c.764G>Aandc.801C>G)mightevolveunderpositiveselection.Theapparentlackofany linkbetweencaprineMC1Rvariationandgeographymightbeexplainedbyacomplexarrayoffactors includingartiﬁcialselectionforpigmentationphenotypesandrecentdivergenceamongstgoatbreeds.

∗ Correspondingauthor.

E-mailaddress:marcel.amills@uab.cat(M.Amills).

1 _Koray_Kırıkc¸_ı,_Antonia_Noce_and_Ali_Zidi_have_contributed_equally_to_this_work. 2 _Present_address:_Department_of_Animal_Science,_Faculty_of_Agriculture,_Ahi_Evran University,Krsehir,40000Turkey.

1. Introduction

Selectionforcoatcolorwasprobablyimplementedinancient

timesasaconsequenceofreligiousbeliefsandculturalpreferences

oflivestockbreeders(Zeder,1994).Forinstance,theBookof

Num-bersestablishesthatredheifersneedtobeusedinthepuriﬁcation

ritualsofpeoplethathavebeenincontactwithacorpse,andblack

sheepwereslaughteredinChinesesupplicationceremoniesforrain

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becausethiscolorsymbolizeswater(Tao,2007).Nonetheless,there

werealsopracticalreasonsforselectingcertaincolorse.g.white

woolis much easiertodyethanthecoloured one.In domestic

animals,thisprocessofselectioncontributedtogenerateahuge

repertoireofpigmentationpatternsthatcontraststronglywiththe

monochromecoatoftheirwildancestors.

Pigmentationisapolygenictraitdeterminedbyalarge

num-berofloci(Sturmetal.,2001).Themelanosomalproteincomplex

isformedbytyrosinase,theenzymethatcatalysesthesynthesis

ofmelanin,plustwootherenzymes:tyrosinase-relatedproteins

1and 2(TYRP1and TYRP2).Tyrosinase-relatedprotein2

catal-ysesthesynthesisof5,6-dihydroxyindole-2-carboxylicacidfrom

DOPAchrome,whichisconvertedtoeumelaninbyTYRP1(Kondo

andHearing,2011).Besides,bothTYRP1andTYRP2contributeto

thestabilizationoftyrosinase(Sturmet al.,2001).Anotherkey

playerinthedetermination of coatcoloris themelanocortin1

receptor(MC1R)gene.Bindingofthismolecule,onthecellsurface

ofmelanocytes,byproopiomelanocortin(POMC)raisesthelevels

ofcAMPandactivatestyrosinase,thusinducingthesynthesisof

blackeumelanin(Sturmetal.,2001).Theagouti-signalingprotein

(ASIP),thatisalsoaligandforMC1R,hastheoppositeeffecti.e.

byloweringtheactivationoftyrosinasepromotesthesynthesisof

red/yellowpheomelanin(MakovaandNorton,2005;Parra,2007).

Inhumans,MC1Rdiversityhasbeenlinkedtogeographyand,

morespeciﬁcally,totheamountofsunlightexposure(Savageetal.,

2008).Thisdifferentialdistributionisnotonlyexplainedbydrift

anddemographicfactors,butalsobynaturalselection.Inthisway,

dark,eumelanin-richphotoprotectivepigmentationisconsidered

tobeadvantageousattropicalandequatoriallatitudesbecauseit

isassociatedwitha decreasedrateofultraviolet-inducedfolate

degradation(JablonskiandChaplin,2010).Incontrast,alightskin

is favoured in geographic areas withreduced sunlight because

itenhancesthesynthesis of vitaminD3 (Jablonskiand Chaplin,

2010).

Geneticdiversityoflivestockpigmentationgeneshasbeenless

studiedatanintercontinentalscalethanthatofhumans(Switonski

etal.,2013).RemarkabledifferencesinthedistributionofMC1R

haplotypeshavebeendetectedwhencomparingChineseand

Euro-peanswine(Giuffraetal.,2000).Previousstudiesperformedin

goatscharacterizedthediversityoftheMC1Rgene(Fontanesietal.,

2009;Nicolosoetal.,2012;Badaouietal.,2014),butitwasdifﬁcult

toascertainifitisassociatedwithgeographybecauseonlyItalian

andSpanishpopulationsweresampled.Inthecurrentwork,we

aimedtoinvestigateifthere isalinkbetweengoatMC1R

poly-morphismand geographybyanalysingindividualsfromseveral

locationscoveringabroadgeographicrange.

2. Materialsandmethods

2.1. Goatsampling

Bloodsamples werecollectedby jugular venipuncturefrom

Colombian (N=9), Italian (Sarda breed, N=7), French (Saanen

breed,N=8),Iranian(Lori-BakhtyariandLori,N=3),Greek(Youra

breed,N=2),Romanian(Carpathianbreed,N=7),Sahelian(N=6)

andSpanish(Majorcanbreed,N=8;Palmerabreed,N=8)goats

(Supplementary TableS1). Sampling wasperformed by trained

veterinariansinthecontextofsanitationcampaignsand

parent-age controls not directly related with our research project.

In all instances, veterinarians followed standard procedures

and relevant internationalguidelines to ensurean appropriate

animalcare(ARRIVEguidelines,

https://www.nc3rs.org.uk/arrive-guidelines; EU Directive 2010/63/EU for animal experiments).

Genomic DNA was puriﬁed with the DNeasy Blood & Tissue

Kit (Qiagen, Barcelona, Spain) and resuspended in ultrapure

water.

2.2. Ampliﬁcationandsequencingprotocols

ByusingprimersFW1,5-CCTGCACTCCCCCATGTAC-3and

REV1,5-TGCGGAAGGCATAAATGAGG-3,weampliﬁeda

frag-mentofapproximately0.7kboftheMC1Rgenethatinprevious

studieswasshowntocontainmostofitspolymorphism(Fontanesi

etal.,2009;Badaouietal.,2014).Polymerasechainreactionswere

performedinaﬁnalvolumeof15␮Lcontaining1.5␮Lof10xPCR

buffer, 2.5mM MgCl2,0.3␮M ofeach primer, 0.25mM of each

dNTP,0.75UTaqGoldDNApolymerase(AppliedBiosystem,

Fos-terCity,CA)and50nggenomicDNA.Thisreactionmixturewas

heatedto95◦Cfor10min,followedby35cyclesof95◦Cfor1min,

62◦Cfor1minand72◦Cfor1min.Subsequently,aﬁnalextension

stepat72◦Cfor10minwascarriedout.Ampliﬁcationproducts

werepuriﬁed withtheExoSAP-IT PCR Cleanupkit (Affymetrix,

SantaClara,CA) andsequencedinbothdirections withprimers

FW2, 5-ACC TGC TGG TGA GCG TCAG-3 and REV1.

Sequenc-ingreactionswerepreparedwiththeBigDyeTerminatorCycle

SequencingKitv1.1(AppliedBiosystems)andelectrophoresedin

anABI3730DNAAnalyzer(AppliedBiosystems).Chromatograms

wereeditedwiththeSeqScapesoftwarev2.5(AppliedBiosystems),

andallsequencesweresubmittedtotheGenBankdatabase

(acces-sioncodes:KT071610-KT071667).

2.3. Phylogeneticandpositiveselectionanalyses

Wecarriedoutphylogeneticandstatisticalanalysesbyusing

ourMC1Rdatasetplus39goatMC1Rsequences(Supplementary

TableS1)retrievedfromGenBank(Badaouietal.,2014).A

median-joiningnetworkwasbuiltwiththeNetwork4.6software(Bandelt

etal.,1999)byusingdefaultparameters.Thecodemlprogramof

thePAML4package(Yang,2007)wasemployedtodetectpositive

selection.Maximumlikelihoodestimatesofthew-ratio(dN/dS),i.e.

therateofnon-synonymoussubstitutionspernon-synonymous

site (dN) divided by the rate of synonymous substitutions per

synonymoussite(dS),wereobtainedforeachcodonofthe

MC1R-encodingregionunderanalysis.Wecontrastedmodels7(neutral

model),whichassumesa␤-distributionforthew-ratio(0≤w≤1),

withmodelM8(selectionmodel),whichtakesintoaccountanextra

categoryofsiteswithw1>1(positiveselection).Theperformance

ofalikelihood ratiotest,wheretwicethedifferenceinthe

log-likelihoodvalues correspondingtomodels7and8is compared

witha ␹2 _with_two _degrees_of_freedom,_was_used_to_assess_the

statisticalsigniﬁcanceofpositiveselection.BayesEmpiricalBayes

inferencewasemployedtodeterminetheposteriorprobabilitythat

agivencodonevolvesunderpositiveselection.Wealsoperformed

a Manteltest (Mantel,1967)toinvestigateifthere isany

rela-tionshipbetweengeographyandMC1Rvariation.Inthisway,we

usedapublisheddataset(Badaouietal.,2014)of319goats

geno-typedfor the c.673C>T, c.676A>G,c.748G>T, c.764G>A MC1R

singlenucleotidepolymorphisms(SNPs).Thesegoatsbelongedto

thefollowingbreeds:CilentanaNera(N=26),Garganica(N=41),

GrigiaMolisana(N=13),Girgentana(N=19),Malague ˜na(N=43),

Murciano-Granadina(N=81),Tinerfe ˜na(N=38),Majorera(N=20)

and Palmera(N=38).The Mantel testestimates thelinear

cor-relationbetweentwomatricesofthesamerank,i.e.matricesof

geographic (measured in km) and genetic(FST coefﬁcient)

dis-tances, in order to ﬁnd out if both parameters are associated

(Mantel, 1967). We computedFST coefﬁcients amongst these9

breedswiththeGenepopontheWebsoftware(http://genepop.

curtin.edu.au,Rousset,2008).TheManteltestwascarriedoutwith

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Table1

InsilicopredictionofthefunctionaleffectsofgoatMC1Rpolymorphismswiththreesoftwares(SIFT,SNAP2andMutPred)anddetectionofpositivelyselectedsiteswith PAML4.

MC1RSNP Aminoacidchange SIFT SNAP2 _MutPred _PAML₄_w₌_d

N/dS PAML4Prob(BEB)a

c.673C>T Q225X Stopcodon Stopcodon Stopcodon – –

c.676A>G K226E Nottolerated Effect(score=41) 0.491 4.69±3.73 0.651

c.748G>T F250V Nottolerated Effect(score=70) 0.650 4.36±3.71 0.609

c.764G>A G255D Nottolerated Effect(score=72) 0.647 6.54±3.11 0.922

c.801C>G C267W Nottolerated Effect(score=81) 0.802 6.56±3.10 0.924

a_Posterior_probability_deduced_with_a_Bayes_Empirical_Bayes_approach.

Fig.1.MedianjoiningnetworkofMC1RsequencescorrespondingtoColombian,Spanish(Murciano-Granadina,Malague ˜na,Payoya,Mallorquina),Canarian(Palmera, Major-era,Tinerfe ˜na),Italian(Sarda,CilentanaNera,DerivatadeSiria,GrigiaMolisana,Maltese,Ionica,Girgentana,Garganica),French(Saanen),Greek(Youra),Iranian(Lori-Bakhtyari andLori),SahelianandRomanian(Carpathian)goats.

3. Resultsanddiscussion

Byexaminingindividualswithverydistinctgeographicorigins,

weexpectedtoincreasesigniﬁcantly thecatalogof MC1R

poly-morphismsdetectedingoats.Nevertheless,wejustfoundtheset

ofﬁvemissensepolymorphismsthatwerepreviouslyreportedby

Fontanesietal.(2009),Nicolosoetal.(2012)andBadaouietal. (2014),plusasilentmutationc.825C>T(Table1,Supplementary

Fig.S1).Fouroftheﬁvepolymorphic aminoacidsites detected

ingoatshappenedtobevariablewhencomparingthegoatMC1R

sequencewiththoseofﬁveadditionalmammalianspecies

(Supple-mentaryFig.S2)ThemedianjoiningnetworkshowninFig.1made

evidentthatgoatsdidnotclusteraccordingtotheirgeographic

ori-gin.Therewasonemainhaplotypewithabroadintercontinental

distributionandseveralotherMC1Rhaplotypessharedby

individ-ualsfromdistinctcountries(Fig.1,SupplementaryTableS2).This

patterncontrastswiththeoneobservedinhumans,wheremarked

differencesinMC1RallelefrequenciesbetweenAsian,Africanand

Caucasian populationsexist, probablyas a consequenceof

nat-uralselection(eumelanin-richpigmentationisphotoprotective),

geneticdriftandotherfactors(Savageetal.,2008;Jablonskiand

Chaplin2010).Wealsocarried outofa Manteltest basedona

datasetof319goatsdistributedin9populationsandgenotyped

for4MC1Rpolymorphisms.AsshowninSupplementaryFig.S3,

the correlation between geographic and genetic distances was

low (r=0.244) and non-signiﬁcant (P-value=0.152). Thisresult

supportstheabsenceofadetectablelinkbetweengoatMC1R

poly-morphismandthegeographicdistributionofcaprinebreeds.

Thetypeofvariationthatweandothers(Fontanesietal.,2009;

Badaouietal.,2014)havefoundinthecaprineMC1Rgeneisquite

particularofthislocus.Inthisway,thevastmajorityof

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SIFT(Kumaretal.,2009),SNAP2₍_Hecht_et_al.,₂₀₁₅₎_and_MutPred

(Lietal.,2009)indicatethattheymighthavefunctional

conse-quences(Table1).Inastudyperformedinpigs,Fangetal.(2009)

alsodetectedanexcessofnon-synonymousvariationattheMC1R

gene.Oneplausibleexplanationforthispatternofvariationcould

betheoccurrenceof positiveselectioni.e.thesystematic

selec-tionofmutationsassociatedwithcoatcolorwouldhaveinvolved

thelossoflinkedneutralvariants.Wetestedthishypothesisby

analysingourdatasetofMC1Rsequenceswiththecodemlprogram

ofthePAML4package(Yang,2007).Theperformanceofa

like-lihoodratiotestprovidedstatisticalsupportfortheexistenceof

positiveselection(P<0.005).Moreover,twocodonswithposterior

probabilitiesabove0.90wereidentiﬁedwiththeBayesEmpirical

Bayesmethod(Table1).Thisisconsistentwithpreviousreports,

whereevidencesof positive selectionacting ontheMC1R gene

ofpigs(Fangetal.,2009)andcattle(Zhaoetal.,2015;Xuetal.,

2015)havebeenprovided.Interestingly,inapreviousstudywe

determinedthatthegenotypicfrequencies ofthecaprineMC1R

c.801C>GSNPareremarkablydifferentinblackand mahogany

Murciano-Granadinagoats(Zidietal.,2012),aresultthatis

con-sistentwithdataprovidedbyFontanesietal.(2009).Moreover,we

foundthatthegenotypicfrequenciesofthec.764G>A

polymor-phismaresigniﬁcantlydifferentinredandblackPalmeragoats,

andthatblondeandwhiteMalague ˜nadisplaydifferentgenotypic

frequenciesofthec.676A>Gandc.748G>TSNPswhencompared

withtheirdarkchestnutcounterparts(Zidietal.,2012).

ThelackofdetectionofatightlinkbetweencaprineMC1R

varia-tionandgeographyisnotunexpectedbecausetheeffectsofneutral

forcesonthevariationofpigmentationgenescanbecounteracted,

tosomeextent,byartiﬁcialselectionforcoatcolor.Indeed,breeds

thataregeographicallyclosemayhavebeenselectedforvery

differ-entcoatcolors,whilstpopulationsfromdistantlocationsmayhave

beenconvergentlyselectedforsimilarpigmentationphenotypes.

Notably,thelackofcorrespondencebetweengoatMC1R

polymor-phismandgeographycontrastsstronglywithdataevidencingthat

ChineseandEuropeanpigscarrydifferentMC1Rhaplotypes(Fang

etal.,2009).Onepossibleexplanationforthisdiscrepancywould

bethatthequantitativeeffects ofMC1Rvariation oncoatcolor

arenotthesameingoatsandpigs,sotheintensityofselection

exertedonthislocuswouldbealsodifferentinthesetwo

live-stockspecies. Indeed, Fontanesi et al. (2009)presented several

casesof incomplete associations betweenMC1R genotypes and

pigmentationingoats.Forinstance,theprotein-truncating

muta-tionc.673C>T(Q225X)washighlyassociated withred colorin

theGirgentana breed,butin theDerivatadeSiriabreed,which

isalsored,only15%oftheindividualswerehomozygousforthis

polymorphism (Fontanesi et al., 2009).Similarly, thec.801C>G

mutationwashighlyassociatedwiththeblackvsmahoganycolor

inMurciano-Granadinagoats,whilethisrelationshipwasless

evi-dentintheMaltesebreed.AccordingtoFontanesi etal.(2009),

theseobservationsmaybeexplainedbytheexistenceofundetected

mutationswithregulatoryeffectsonMC1Rexpression,genetic

het-erogeneityinthedeterminationofcoatcolorandtheexistenceof

epistaticinteractionswithothergenes,suchasPOMCandASIP,with

strongeffectsonpigmentationpatterns.

AnotherfactorthatmayexplainthedifferentialMC1Rgenetic

patternsobserved in goats and pigs is the time of divergence.

ModerngoatsdescendfromindividualsdomesticatedatEastern

Anatolia10,000YBP(Naderietal.,2008).Incontrast,FarEastern

and Western wild boars diverged 1 Mya and they were

inde-pendentlydomesticatedinChinaandtheNearEast,respectively

(Larsonetal.,2005).Inconsequence,goatbreedsdivergedmuch

more recently than Asian and European pigs, a feature that is

expectedtoweakentheamountofgeneticdifferentiationamongst

populationsdistributedindifferentgeographiclocations.

4. Conclusions

The topologyof themedian-joining network based ongoat

MC1RsequencesandtheManteltestdidnotprovideevidenceofa

relationshipbetweenthevariationofthecaprineMC1Rgeneand

geography,aresultthatcanbeexplainedbymultipledemographic,

biologicalandselectionfactors.Besides,wehaveobtainedevidence

thattwocodonsinthegoatMC1Rgenemightevolveunderpositive

selection.

Conﬂictofinterest

Theauthorsdeclarethattheyhavenoconﬂictofinterest.

Acknowledgments

Thanks toJosé Gutiérrez Plasencia for helping in goat

sam-pling.WealsoacknowledgethesupportoftheSpanishMinistry

ofEconomyandCompetitivityfortheCenterofExcellenceSevero

Ochoa 2016–2019 (SEV-2015-0533) grant awarded to the

Cen-terfor Research inAgricultural Genomics.TainaF Cardoso was

fundedwithafellowshipfromtheCAPESFoundation-Coordination

ofImprovementofHigherEducation,MinistryofEducation(MEC)

oftheFederalGovernmentofBrazil.

AppendixA. Supplementarydata

Supplementarydataassociatedwiththisarticlecanbefound,

intheonlineversion,athttp://dx.doi.org/10.1016/j.smallrumres.

2016.10.010.

References

Badaoui,B.,Manunza,A.,Castelló,A.,D’Andrea,M.,Pilla,F.,Capote,J.,Jordana,J., Ferrando,A.,Martínez,A.,Cabrera,B.,Delgado,J.V.,Landi,V.,Gómez,M.,Pons, A.,ElOuni,M.,Vidal,O.,Amills,M.,2014.Technicalnote:advantagesand limitationsofauthenticatingPalmeragoatdairyproductsbypyrosequencing themelanocortin1receptor(MC1R)gene.J.DairySci.97,7293–7297. Bandelt,H.J.,Forster,P.,Röhl,A.,1999.Median-joiningnetworksforinferring

intraspeciﬁcphylogenies.Mol.Biol.Evol.16,37–48.

Fang,M.,Larson,G.,Ribeiro,H.S.,Li,N.,Andersson,L.,2009.Contrastingmodeof evolutionatacoatcolorlocusinwildanddomesticpigs.PLoSGenet.5, e1000341.

Fontanesi,L.,Beretti,F.,Riggio,V.,Dall’Olio,S.,González,E.G.,Finocchiaro,R., Davoli,R.,Russo,V.,Portolano,B.,2009.Missenseandnonsensemutationsin melanocortin1receptor(MC1R)geneofdifferentgoatbreeds:associationwith redandblackcoatcolourphenotypesbutwithunexpectedevidences.BMC Genet.10,47.

Giuffra,E.,Kijas,J.M.,Amarger,V.,Carlborg,O.,Jeon,J.T.,Andersson,L.,2000.The originofthedomesticpig:independentdomesticationandsubsequent introgression.Genetics154,1785–1791.

Hecht,M.,Bromberg,Y.,Rost,B.,2015.Betterpredictionoffunctionaleffectsfor sequencevariants.BMCGenomics16(8),S1.

Jablonski,N.G.,Chaplin,G.,2010.Colloquiumpaper:humanskinpigmentationas anadaptationtoUVradiation.Proc.Natl.Acad.Sci.U.S.A.107(Suppl(2)), 8962–8968.

Kondo,T.,Hearing,V.J.,2011.Updateontheregulationofmammalianmelanocyte functionandskinpigmentation.Exp.Rev.Dermatol.6,97–108.

Kumar,P.,Henikoff,S.,Ng,P.C.,2009.Predictingtheeffectsofcoding

non-synonymousvariantsonproteinfunctionusingtheSIFTalgorithm.Nat. Protoc.4,73–81.

Larson,G.,Dobney,K.,Albarella,U.,Fang,M.,Matisoo-Smith,E.,Robins,J.,Lowden, S.,Finlayson,H.,Brand,T.,Willerslev,E.,Rowley-Conwy,P.,Andersson,L., Cooper,A.,2005.Worldwidephylogeographyofwildboarrevealsmultiple centersofpigdomestication.Science307,1618–1621.

Li,B.,Krishnan,V.G.,Mort,M.E.,Xin,F.,Kamati,K.K.,Cooper,D.N.,Mooney,S.D., Radivojac,P.,2009.Automatedinferenceofmolecularmechanismsofdisease fromaminoacidsubstitutions.Bioinformatics25,2744–2750.

Makova,K.,Norton,H.,2005.WorldwidepolymorphismattheMC1Rlocusand normalpigmentationvariationinhumans.Peptides26,1901–198. Mantel,N.,1967.Thedetectionofdiseaseclusteringandageneralizedregression

approach.CancerRes.27,209–220.

Naderi,S.,Rezaei,H.R.,Pompanon,F.,Blum,M.G.,Negrini,R.,Naghash,H.R.,Balkiz, O.,Mashkour,M.,Gaggiotti,O.E.,Ajmone-Marsan,P.,Kence,A.,Vigne,J.D., Taberlet,P.,2008.Thegoatdomesticationprocessinferredfromlarge-scale

(5)

mitochondrialDNAanalysisofwildanddomesticindividuals.Proc.Natl.Acad. Sci.U.S.A.105,17659–17664.

Nicoloso,L.,Negrini,R.,Ajmone-Marsan,P.,Crepaldi,P.,2012.Onthewayto functionalagrobiodiversity:coatcolourgenevariabilityingoats.Animal6, 41–49.

Parra,E.J.,2007.Humanpigmentationvariation:evolution,geneticbasis,and implicationsforpublichealth.Am.J.Phys.Anthropol.,85–105.

Rousset,F.,2008.Genepop’007:acompletereimplementationoftheGenepop softwareforWindowsandLinux.Mol.Ecol.Res.8,103–106.

Savage,S.A.,Gerstenblith,M.R.,Goldstein,A.M.,Mirabello,L.,Fargnoli,M.C.,Peris, K.,Landi,M.T.,2008.Nucleotidediversityandpopulationdifferentiationofthe melanocortin1receptorgene,MC1R.BMCGenet.9,31.

Sturm,R.A.,Teasdale,R.D.,Box,N.F.,2001.Humanpigmentationgenes:

identiﬁcation,structureandconsequencesofpolymorphicvariation.Gene277, 49–62.

Switonski,M.,Mankowska,M.,Salamon,S.,2013.Familyofmelanocortinreceptor (MCR)genesinmammals-mutations,polymorphismsandphenotypiceffects.J. Appl.Genet.54,461–472.

Tao,W.,2007.Shangritualanimals:colourandmeaning(part1).B.Sch.Orient. Afr.St.70,305–372.

Xu,L.,Bickhart,D.M.,Cole,J.B.,Schroeder,S.G.,Song,J.,Tassell,C.P.,Sonstegard, T.S.,Liu,G.E.,2015.Genomicsignaturesrevealnewevidencesforselectionof importanttraitsindomesticcattle.Mol.Biol.Evol.32,711–725.

Yang,Z.,2007.PAML4:phylogeneticanalysisbymaximumlikelihood.Mol.Biol. Evol.24,1586–1591.

Zeder,M.A.,1994.Ofkingsandshepherds:specializedanimaleconomyinURIII Mesopotamiae.In:Stein,G.,Rothman,M.(Eds.),ChiefdomsandEarlyStatesin theNearEast:theOrganizationalDynamicsofComplexity.PrehistoryPress, Madison,pp.175–191.

Zhao,F.,McParland,S.,Kearney,F.,Du,L.,Berry,D.P.,2015.Detectionofselection signaturesindairyandbeefcattleusinghigh-densitygenomicinformation. Genet.Sel.Evol.47,49.

Zidi,A.,Badaoui,B.,Manunza,A.,Serradilla,J.M.,Capote,J.,Urrutia,B.,Carrizosa,J., Pilla,F.,D’Andrea,M.S.,Jordana,J.,Ferrando,A.,Pons,A.,Sánchez,A.,Landi,V., Gómez,M.,Martínez,A.,Vidal,O.,Amills,M.,2012.Mutationsatthecaprine MC1RgeneareassociatedwithcoatcolorinSpanishgoats.In:11th InternationalConferenceonGoats,GranCanaria,Spain.