Derived immune and ancestral pigmentation alleles in a 7000-year-old Mesolithic European

13 Pages·2014·6.36 MB·English

by  Iñigo Olalde

LETTER
doi:10.1038/nature12960
Derived immune and ancestral pigmentation alleles
in a 7,000-year-old Mesolithic European
In˜igoOlalde1*,MortenE.Allentoft2*,FedericoSa´nchez-Quinto1,GabrielSantpere1,CharlestonW.K.Chiang3,
MichaelDeGiorgio4,5,JavierPrado-Martinez1,JuanAntonioRodr´ıguez1,SimonRasmussen6,JavierQuilez1,OscarRam´ırez1,
UrkoM.Marigorta1,MarcosFerna´ndez-Callejo1,Mar´ıaEncinaPrada7,JulioManuelVidalEncinas8,RasmusNielsen9,
MihaiG.Netea10,JohnNovembre11,RichardA.Sturm12,PardisSabeti13,14,Toma`sMarque`s-Bonet1,15,ArcadiNavarro1,15,16,17,
EskeWillerslev2&CarlesLalueza-Fox1
Ancientgenomicsequenceshavestartedtorevealtheoriginandthe
demographicimpactoffarmersfromtheNeolithicperiodspread- a b
ing into Europe1–3. The adoption of farming, stock breeding and
sedentarysocietiesduringtheNeolithicmayhaveresultedinadapt-
ivechangesingenesassociatedwithimmunityanddiet4.However,
the limited data available from earlier hunter-gatherers preclude
LLLaaa BBBrrraaaññaaa---AAArrriiinnnttteeerrrooo
anunderstandingoftheselectiveprocessesassociatedwiththiscru- sssiiittteee
cialtransitiontoagricultureinrecenthumanevolution.Herewe
sequenceanapproximately7,000-year-oldMesolithicskeletondis-
coveredattheLaBran˜a-ArinterositeinLeo´n,Spain,toretrievea
complete pre-agricultural European human genome. Analysis of
this genome in the context of otherancient samples suggests the c
existenceofacommonancientgenomicsignatureacrosswestern 0.08 LFILFI
aTmwevwvnahaeeirdrsdenieaenLtcnanaeoattlcntrisBeoteyarrnateasahdtsgln˜ayoeuEatcnpbuiiaeranirseqtsad,esuissiedgivuinantiwgotdifgfuiuirietnscoahsamltitnpinhcntaaitgMstrhnhrtheueihoeusmaUgsontelbpattnienhtephrrrcee-eierglocssiafgiPttstrhidathamtaleelensrreaokiceslvliel.rineet.Midlhone,isfocmpmrintueooootsdadvetteeehvirrrveen,nerMwalEEyleuuesasrrkpdoooiranlppoipteevpthaaiiidinnvgceess-. 00..0046 La Braña 1 FI FFII FIFIFFFIIFIFFIFIFIFIIIFFFFFFIIIIFIIIFFFFFIFIFIFIFFFFIIFIFIIFFIIFIFIIIFIIIFFIFFFFIIFFIIIIIIFFFFILFIIIFFIFFFIILIFIILFFIFFFLIFIFFIILIFIIILFLLILFLIFFLFIFLIILFLLIIFFIFLFFIIFIIILILFFLFFLLLIFILLILIFFFLIFLFFLFLIIILFLLFIILIFIFLFIFLFILFFLILIFLIIFILFLLIIFFFILIFIFFIILILFLIIILFFFIFFLIIIILFILLLLFLIFLFFLLFIFIFIIFFILILLFLLIILILLFFFLFIFLFFLFIIIFIIFIIFIIII LLLFFFIIILLLFFFIIILFI
FI
Next-generation sequencing (NGS) technologies are revolution- C1 Ire8 AFIjv52 Ajv70 FI
wdgMhiczieiuhinanenTmtgesegenhotaloitelyncitfhtwIphecbcboroioofeceimmdremcpileypadpeadnlrfoleiooeinstosfueigsidtantkiaenotd(enhncclcceeiiiarenitoeNlconlfnatentehdets1Doeag0l(LrNieAn,lt0anihAael0opiBmr0cms(r–h1aepa.5eudDne˜Hs,nr0a5NLi,to0-6oae,AA0wdrsB-r))eyug,i.rvencaaaeahttnn˜rreh,dasraevosbrh1eeetawrrahfvyonaapersldtoieetodpnt2pfliu)aersOblce¨waisolstteeezvidknorie,enntrt,aehosfBdoweNfPursn;ieonneiommqdol2ufimit0entth0hhnei6acee-- P –000...000202 IEIIIIIIEEEEEEIIEEUIEIKUEKIUIUUUEEUIIIUIUEEKEDLIKUEKKUUUUEKUKIVSKUIEKIEKKKEUKIEKIcEPNKUDEUUUKIIUtIIUNLEUELEUEKDEIKUKKUIUUKUEIIUEKOUKIUEUKEUEUIIUNEIUKUUUKEEUIKKKUIUUENDSUUDUUKIKEUDEUKKUUKIKEUUKUILUDKUUKKEKIKUUEKESUKKELUEKKUKKNEIUEKUKNUKKDKKKSEUKEUUKKDIKIEKFUNUNUKKDDUKLEUEKUUPIUKOUKEEUUSPUKSUUKUKRUEESUIUIUUDUSUKSLKLKEIEUKEUUEDPKLUUFDKIKKUKIEDcLSUKPUNEKUKUEUIKKKCUEUKUKUUFEEKKFDEKKcUUUKUEUNRKULKKCUPEtUDKKEDEKKLUKDKLUKRUKUINtKUHKKKKKUEBUKKIUNSUKKUKLUUUIULKHRKCBBEIKEUUUEEUNUKEBKEDKKLKDDDIBUKEBDDcKUIUKOKBUKKKCKKNFEUIEEBUHUCEUUKKCKUIUBUPKELEKIUFEUtEEBBEFEEUEKCPEECDEBKRKSEBUKDUBHCLUFFCEKKHKDKHELKFRKUKDPDUCRKUNUUKUBEECLKHEcUEHCERRIUUUKEEBKHNKHDUNRNUUEEUSUCUUDLCKUEUEDKHDHBCKtDELKKUUKUHPHKHFFBKUFKKECPFLDUEKLUCLEBKKFKKHKDEKUHUCKEKCEEUDHFESBCLKKKRRRUPSPBREULKHCURCFFBFEKDBKEUHPFUCEURKUKHCPCEFNHEEFHCULELCBECCPDCRIRHCFRUFKUBUREHBKBHLEEBDUKKHCBRDKRLKBHCCHLCUDBHDRKRBLERHFUCHFEBHHHCRHCKEKDKEEPCBCHUEFFEKPEHFCCCECEDCCDUZHCHEEKRCFREDRNEEPDUCERCHHDFZCLIREDCECIHCHFURBUKLCFCCHHCHCHFCEDCERECFHHKHCBSCCCCLHADUEDDRLECHCHCERHCECHRDFZECHBDCKRCCZCHRFHCHHFCECHHHFCEHCCHTDFEHCCHHDEHEECCHCCCHCBCHRHAFZHEUCFCCPERHEHCIHFRHCFHHCFCCCRHRCCDHCSHCFHCUHHETCFCCDHCHCFECCDHCHUFCCHFSHHERFCTCHCZCRBCCRLBCRCRFBKCRDCBHHHCFFZFCCCBCUHACUHHCHHRDCCREHHRHPCHCRRHABRHEHHHHIEHHAHCHHCEHCFHHCRHCEFOCCZARERCHRCEEHHEHHCFFHCTFACHHHHHKCCHAEFCHHHCLCHFCFDTHHEEHFCCRTHSEERYHCHCCHHHCTCCRRAFHHCHRCBHADECRHEHRHFHRTCHRCCHHPDHIAURCHCFSHRHEECEASDUSHHFRGDFHCTACREFHCCHHCCTZHHSCCTSARFHFCHCUECHUFCRRTHZCHTCESEESHHERTRHFERPSTECEAUHFCEHHHRRHECHCPCHHRHECEHHFHSFSHBCHSHPCCRUCPCIRSTFCCITCCSHEHCEHFSFHCTCSRTHHSHEUTRPCHPPIACTCPHTFREHHKEHHCSERPRSTHHHUHUUYHCEISHHTCDFTCYHTTRLSHYRHSEEPHFSYETHEEGYSEAHPFRFFGEHEPEEIHHIEGRSSCSSERTFGYHRPPSUTSCEEFSEGTRCCYCSYPTRRRAPSSSTIPEIIESUISEPESGTHTTEESOCCRSRETTSIHEEEPGPPIEGTYPEPHIHBHTUTCIIYTISBTSCPTEPSCSETTTISSPPTPPEPSPIESSUHSFHETSTETESEGETTRETFEIATEIFPGPBHYEERASPDTTESTEIPCHCTPHSPTETTTTSPRPTESICFFPSSTRSSKPSFESFEPPEEEORYSETTTSSGIEAEEBTESSIISEEPTEESTPTHPZPTTSRTREPETTRTRIPCESESHTESPSSETTSGPEIHSSTSPPSIPPASSIITSPTTITTEPICSITSTPESSTTTSEIPIPSREPTHEETSKTIRITTYPPPTTETTRETPHTICPPIBTPETYETEPEPSPPHBPITPPSETIITHTESPSETTEETSSOPITEPSYPPOTTGSETETTPSPIPPTTTTSUIMITSGEHSGPSPTTTHTTTTASTKPUSPECSTESITSTIITPSETGTETEPRPSSETETRTTTTTEMSEPTTPEPSUTSTSKHSTSIIRSSMTYTSOSPOPPSCEPETTTIISPTEPTEIPKITEPYTPPYOTGPTTPTISTPSTIKBEEIHSTPYSTTTIGSTPPTGTITTPYGTTRITTITYSSPAYTGTITTTPPRPTTGPOPYRIGCBIETGIYTYTTTYTTEIPTTGMPIFOSPYRGHTPGTIGPPGSEIITTEIITAIYTGTTORAYHKTERTEITIIAITTSPPSTTTTILGIYPLGSOITUSOYIITLITTTRETTITIYGTIIYTPTGITTPISIEOTIIGITTGTTIITITYTITITSTGIGITTIITTIGITTHGTTRYRIGIITYYYIRIRTTIGIIITITITTTRTTGYTGGIIIYITTITIDTIGTITITIITGIITITTEITTITTTIITITIIIIITITTTTTIIIIITIITTTTTTITIIIIIIITIITTTTIITTTTTSTGTIIIIITITTIUIITKTGITIIRTCITTTTTTIKIITHITTIITGTIToIIITITITITÖRITTTITIIITCIITTTIIkTTTTIIHIITTtITTI4ITTITzITCIITTGTTIIIRYiTIITTRTITITTIITTICTTYRITITITITTCRY TCRYUK
deepcavesystem,1,500mabovesealevelintheCantabrianmountain
–0.06 –0.04 –0.02 0.00 0.02 0.04 0.06
range(Leo´n,NorthwesternSpain)(Fig.1a).Theskeletonsweredated
PC2
toapproximately7,000yearsBP(7,940–7,690calibratedBP)7.Because
ofthecoldenvironmentandstablethermalconditionsinthecave,the Figure1|GeographiclocationandgeneticaffinitiesoftheLaBran˜a1
preservationofthesespecimensprovedtobeexceptional(Fig.1b).We individual.a,LocationoftheLaBran˜a-Arinterosite(Spain).b,TheLaBran˜a1
identifiedatoothfromLaBran˜a1withhighhumanDNAcontent(48.4%) skeletonasdiscoveredin2006.c,PCAbasedontheaverageoftheProcrustes
transformationsofindividualPCAswithLaBran˜a1andeachofthefive
and sequenced this specimen to a final effective genomicdepth-of-
Neolithicsamples1,3.ThereferencepopulationsaretheFinnishHapMap,
coverageof3.403(ExtendedDataFig.1).
FINHMandPOPRES.Populationlabelswithlabellingofref.12withthe
Weusedseveralteststoassesstheauthenticityofthegenomesequence
additionofFI(Finns)orLFI(late-settlementFinns).Ajv70,Ajv52,Ire8and
andtodeterminetheamountofpotentialmodernhumancontamina- Gok4areScandinavianNeolithichunter-gatherersandafarmer,respectively3.
tion.First,weobservedthatsequencereadsfromboththemitochondrial O¨tziistheTyroleanIceMan1.
1InstitutdeBiologiaEvolutiva,CSIC-UPF,Barcelona08003,Spain.2CentreforGeoGenetics,NaturalHistoryMuseumofDenmark,UniversityofCopenhagen,DK-1350CopenhagenK,Denmark.
3DepartmentofEcologyandEvolutionaryBiology,UniversityofCalifornia,LosAngeles,California90095,USA.4DepartmentofIntegrativeBiology,UniversityofCalifornia,Berkeley,California94720,USA.
5DepartmentofBiology,PennsylvaniaStateUniversity,502WartikLaboratory,UniversityPark,Pennsylvania16802,USA.6CenterforBiologicalSequenceAnalysis,TechnicalUniversityofDenmark,DK-
2800KongensLyngby,Denmark.7I.E.S.O.’LosSalados’,JuntadeCastillayLeo´n,E-49600Benavente,Spain.8JuntadeCastillayLeo´n,ServiciodeCulturadeLeo´n,E-24071Leo´n,Spain.9Centerfor
TheoreticalEvolutionaryGenomics,UniversityofCalifornia,Berkeley,California94720,USA.10DepartmentofMedicineandNijmegenInstituteforInfection,InflammationandImmunity,Radboud
UniversityNijmegenMedicalCentre,6500Nijmegen,TheNetherlands.11DepartmentofHumanGenetics,UniversityofChicago,Illinois60637,USA.12InstituteforMolecularBioscience,Melanogenix
Group,TheUniversityofQueensland,Brisbane,Queensland4072,Australia.13CenterforSystemsBiology,DepartmentofOrganismicandEvolutionaryBiology,HarvardUniversity,Cambridge,
Massachusetts02138,USA.14BroadInstituteoftheMassachusettsInstituteofTechnologyandHarvard,Cambridge,Massachusetts02142,USA.15Institucio´CatalanadeRecercaiEstudisAvançats
(ICREA),08010Barcelona,Catalonia,Spain.16CentredeRegulacio´Geno`mica(CRG),Barcelona08003,Catalonia,Spain.17NationalInstituteforBioinformatics(INB),Barcelona08003,Catalonia,Spain.
*Theseauthorscontributedequallytothiswork.
13 MARCH 2014 | VOL 507 | NATURE | 225
©2014Macmillan Publishers Limited. All rights reserved
RESEARCH LETTER
DNA(mtDNA)andthenuclearDNAofLaBran˜a1showedthetypical theLaBran˜a1genometoadatasetofmodernhumanswithsimilar
ancientDNAmisincorporationpatternsthatarisefromdegradationof coverage (3–43). The overall genomic heterozygosity was 0.042%,
DNAovertime8(ExtendedDataFig.2a,b).Second,weshowedthatthe lowerthanthevaluesobservedinpresentdayAsians(0.046–0.047%),
observednumberofhumanDNAfragmentswasnegativelycorrelated Europeans (0.051–0.054%) and Africans (0.066–0.069%) (Extended
withthefragmentlength(R2.0.92),asexpectedforancientdegraded DataFig.6a).Theeffectivepopulationsize,estimatedfromheterozyg-
DNA, and that the estimated rate of DNA decay was low and in ositypatterns,suggestsaglobalreductioninpopulationsizeofapproxi-
agreementwithpredictedvalues9(ExtendedDataFig.2c,d).Wethen mately20%relativetoextantEuropeans(SupplementaryInformation).
estimatedthecontaminationrateinthemtDNAgenome,assembledto Moreover,noevidenceoftractsofautozygositysuggestiveofinbreed-
a high depth-of-coverage (913), by checking forpositions differing ingwasobserved(ExtendedDataFig.6b).
fromthemtDNAgenome(haplogroupU5b2c1)thatwaspreviously To investigatesystematicallythe timingofselectioneventsinthe
retrievedwithacapturemethod2.Weobtainedanuppercontamina- recenthistoryofmodernEuropeans,wecomparedtheLaBran˜agen-
tionlimitof1.69%(0.75–2.6%,95%confidenceinterval,CI)(Supplemen- ometomodernpopulationsatlocithathavebeencategorizedasof
taryInformation).Finally,togenerateadirectestimateofnuclearDNA interestfortheirroleinrecentadaptiveevolution.Withrespecttotwo
contamination,wescreenedforheterozygouspositions(amongreads recentwell-studiedadaptationstochangesindiet,wefoundtheancient
with.43coverage)inknownpolymorphicsites(SingleNucleotide genometocarrytheancestralalleleforlactoseintolerance4andapproxi-
PolymorphismDatabase(dbSNP)build137)atuniquelymappedsec- matelyfivecopiesofthesalivaryamylase(AMY1)gene(ExtendedData
tionsontheXchromosome6(SupplementaryInformation).Wefound Fig. 7 and Supplementary Information), a copy numbercompatible
that the proportion of false heterozygous sites was 0.31%. Together with a low-starch diet18. These results suggest the LaBran˜a hunter-
these results suggest low levels of contamination in the La Bran˜a 1 gathererwaspooratdigestingmilkandstarch,supportingthehypo-
sequencedata. thesesthattheseabilitieswereselectedforduringthelatertransitionto
ToinvestigatetherelationshiptoextantEuropeansamples,wecon- agriculture.
ducted a principal component analysis (PCA)10 and found that the Toexpandthesurvey,weanalysedacatalogueofcandidatesignals
approximately7,000-year-oldMesolithicsamplewasdivergentfrom for recent positive selection based on whole-genome sequence vari-
extantEuropeanpopulations(ExtendedDataFig.3a,b),butwasplaced ationfromthe1000GenomesProject13,whichincluded35candidate
inproximitytonorthernEuropeans(forexample,samplesfromSweden non-synonymousvariants,tenofwhichweredetecteduniquelyinthe
and Finland)11–14. Additional PCAs and allele-sharing analyses with CEU(UtahresidentswithnorthernandwesternEuropeanancestry)
ancientScandinavianspecimens3supportedthegeneticsimilarityof sample19.ForeachvariantweassessedwhethertheMesolithicgenome
theLaBran˜a1genometoNeolithichunter-gatherers(Ajv70,Ajv52, carriedtheancestralorderived(putativelyadaptive)allele.
Ire8)relativetoNeolithicfarmers(Gok4,O¨tzi)(Fig.1c,ExtendedData Ofthetenvariants,theMesolithicgenomecarriedtheancestraland
Figs 3c and 4). Thus, this Mesolithic individual from southwestern non-selectedalleleasahomozygoteinthreeregions:C12orf29(agenewith
Europerepresentsaformerlywidespreadgenepoolthatseemstobe unknownfunction),SLC45A2(rs16891982)andSLC24A5(rs1426654)
partiallypreservedinsomemodern-daynorthernEuropeanpopula- (Table 1). The latter two variants are the two strongest known loci
tions,assuggestedpreviouslywithlimitedgeneticdata2,3.Wesubse- affectinglightskinpigmentationinEuropeans20–22andtheirancestral
quentlyexploredtheLaBran˜aaffinitiestoanancientUpperPalaeolithic allelesandassociatedhaplotypesareeitherabsentorsegregateatvery
genomefromtheMal’tasitenearLakeBaikalinSiberia15.Outgroupf lowfrequencies in extant Europeans (3% and 0% for SLC45A2 and
3
andDstatistics16,17,usingdifferentmodernreferencepopulations,sup- SLC24A5,respectively)(Fig.2).Wesubsequentlyexaminedallgenes
portthatMal’taissignificantlyclosertoLaBran˜a1thantoAsiansor knowntobeassociatedwithpigmentationinEuropeans22,andfound
modernEuropeans(ExtendedDataFig.5andSupplementaryInfor- ancestral alleles in MC1R, TYR and KITLG, and derived alleles in
mation).Theseresultssuggestthatdespitethevastgeographicaldis- TYRP1,ASIPandIRF4(SupplementaryInformation).Althoughthe
tanceandtemporalspan,LaBran˜a1andMal’tasharecommongenetic precisephenotypiceffectscannotcurrentlybeascertainedinaEuropean
ancestry,indicatingageneticcontinuityinancientwesternandcentral geneticbackground,resultsfromfunctionalexperiments20indicatethat
Eurasia.Thisobservationmatchesfindingsofsimilarculturalartefacts thealleliccombinationinthisMesolithicindividualislikelytohave
acrosstimeandspaceinUpperPaleolithicwesternEurasiaandSiberia, resultedindarkskinpigmentationanddarkorbrownhair.Further
particularlythepresenceofanthropomorphic ‘Venus’figurinesthat examinationrevealedthatthisindividualcarriedtheHERC2rs12913832*C
havebeenrecoveredfromseveralsitesinEuropeandRussia,including singlenucleotidepolymorphism(SNP)andtheassociatedhomozygous
theMal’tasite15.Wealsocomparedthegenome-wideheterozygosityof haplotypespanningtheHERC2–OCA2locusthatisstronglyassociated
Table1|Mesolithicgenomeallelicstateat10nonsynonymousvariantsrecentlyselectedinEuropeans
Allelicstate Gene Name SNP Amino-acidchange Function
LaBran˜a1carriesthe PTX4 Pentraxin4 rs2745098 Arg281Lys Maybeinvolvedininnate
derivedallele immunity
UHRF1BP1 UHRF1bindingprotein1 rs11755393 Gln454Arg Risklocusforsystemic
lupuserythematosus
GPATCH1 Gpatchdomaincontaining1 rs10421769 Leu520Ser ReceptorforOmpAexpressed
byE.coli
WWOX WWdomain-containingoxidoreductase rs12918952 Ala179Thr Actsasatumoursuppressorand
hasaroleinapoptosis
CCDC14 Coiled-coildomain-containingprotein rs17310144 Thr365Pro Unknown
14
LaBran˜a1carriesboth SETX Senataxin rs1056899 Val2587Ile Involvedinspinocerebellarataxia
theancestralandthe andamyotrophiclateralsclerosis
derivedallele
TDRD12 Tudordomaincontaining12 rs11881633 Glu413Lys Unknown
LaBran˜a1retainsthe C12orf29 Chromosome12openreadingframe29 rs9262 Val238Leu Unknown
ancestralallele
SLC45A2 Solutecarrierfamily45,member2 rs16891982 Leu374Phe Associatedwithskinpigmentation
SLC24A5 Solutecarrierfamily24,member5 rs1426654 Ala111Thr Associatedwithskinpigmentation
226 | NATURE | VOL 507 | 13 MARCH 2014
©2014Macmillan Publishers Limited. All rights reserved
LETTER RESEARCH
rs16891982
1.0
ycneuqerf elellA 0000....2468 CEU
0.0
Genotype La Braña 1
1.0
ycneuqerf elellA 0000....2468 YRI
0.0
rs75567877rs192097188rs35402rs79371483rs187050917rs73074891rs35403rs876363rs80064515rs10461928rs111912886rs111845075rs35404rs77278088rs140309535rs35405rs113214512rs35406rs180707453rs78855057rs35407rs57959046rs55746663rs114794536rs6874998rs6894408rs142167897rs10067723rs115630314rs146144555rs143546871rs16891977rs181922817rs188216331rs35396rs138891424rs40132rs141135868rs191934001rs35397rs78505258rs16891982rs113891096rs114237189rs138005661rs1364038rs185145rs149480117rs185146rs73077106rs75344149rs250417rs116828782rs113928135rs2287949rs114068287rs35389rs7729962rs2113097rs115793513rs114573901rs35391rs76081820rs186125987rs35393rs114452043rs1364037rs76152294rs1010872rs28777rs77366462rs114048710rs75092413rs189002334rs73077127rs149427409rs190934666rs141821415rs80292549rs111618495rs78704525rs7444256rs147821561rs145058219
rs1426654
1.0
ycneuq 00..68
erf elellA 00..24 CEU
0.0
Genotype La Braña 1
1.0
ycneuqerf elellA 0000....2468 YRI
0.0
rs75626786rs75804106rs143538859rs185587149rs116688269rs2244657rs150271785rs10162789rs79875456rs55728404rs2675346rs142889253rs78065868rs111337581rs186348543rs191207217rs183165868rs188836177rs2433354rs2459391rs77575793rs191740934rs190436254rs77294837rs142736532rs139737156rs2675347rs184200025rs144562056rs2555364rs6493306rs142039743rs4775737rs113182462rs145907211rs17426596rs79122997rs2459393rs8041528rs16960624rs80228496rs1426654rs150379789rs140666229rs2433357rs4143849rs75179742rs76613407rs115878304rs192753244rs141123757rs9302141rs76547866rs201749426rs2470102rs138354861rs141380477rs79443233rs186162343rs2469597rs16960631rs16960633rs2433359rs190676599rs77920254rs149290668rs36075490rs12914834rs74011910rs184550982rs117103804rs139802263rs189335331rs74011911rs59318801rs56044246rs77618450rs114906353rs183566272rs114321279rs188030968rs141680414rs150885139rs76332840
Figure2|AncestralvariantsaroundtheSLC45A2(rs16891982,above)and alsoabsentinmodernEuropeans(CEU)(n5112)butpresentinYorubans
SLC24A5(rs1426654,below)pigmentationgenesintheMesolithicgenome. (YRI)(n5113).ThispatternconfirmsthattheLaBran˜a1sampleisolderthan
TheSNPsaroundthetwodiagnosticvariants(redarrows)inthesetwogenes thepositive-selectioneventintheseregions.Blue,ancestral;red,derived.
wereanalysed.TheresultinghaplotypecomprisesneighbouringSNPsthatare
withblueeyecolour23.Moreover,apredictionofeyecolourbasedon describedeQTLs),includingthreeToll-likereceptorgenes(TLR1,TLR2
genotypesatadditionallociusingHIrisPlex24produceda0.823maximal andTLR4)involvedinpathogenrecognition26.
and0.672minimalprobabilityforbeingnon-brown-eyed(Supplemen- TheseobservationssuggestthattheNeolithictransitiondidnotdrive
taryInformation).Thegenotypiccombinationleadingtoapredicted allcasesofadaptiveinnovationonimmunitygenesfoundinmodern
phenotypeofdarkskinandnon-browneyesisuniqueandnolonger Europeans.Severalofthederivedhaplotypesseenathighfrequency
presentincontemporaryEuropeanpopulations.Ourresultsindicate todayinextantEuropeanswerealreadypresentduringtheMesolithic,
that the adaptive spread of light skin pigmentation alleles was not asneutralstandingvariationorduetoselectionpredatingtheNeolithic.
completeinsomeEuropeanpopulationsbytheMesolithic,andthat Denovomutationsthatincreasedinfrequencyrapidlyinresponseto
the spread of alleles associated with light/blue eye colour may have zoonoticinfectionsduringthetransitiontofarmingshouldbeiden-
precededchangesinskinpigmentation. tifiedamongthosegeneswhereLaBran˜a1carriesancestralalleles.
Fortheremainingloci,LaBran˜a1displayedthederived,putatively ToconfirmwhetherthegenomictraitsseenatLaBran˜a1canbe
adaptivevariantsinfivecases,includingthreegenes,PTX4,UHRF1BP1 generalizedtootherMesolithicpopulations,analysesofadditionalancient
andGPATCH1(ref.19),involvedintheimmunesystem(Table1and genomesfromcentralandnorthernEuropewillbeneeded.Nevertheless,
ExtendedDataFig.8).GPATCH1isassociatedwiththeriskofbacterial thisgenomesequenceprovidesthefirstinsightastohowthesehunter-
infection.Wesubsequently determined the allelic statesin 63SNPs gatherers are related to contemporary Europeans and other ancient
from40immunitygeneswithpreviousevidenceforpositiveselection peoplesinbothEuropeandAsia,andshowshowancientDNAcanshed
andforcarryingpolymorphismsshowntoinfluencesusceptibilityto lightonthetimingandnatureofrecentpositiveselection.
infections in modern Europeans (Supplementary Information). La
Bran˜a1carriesderivedallelesin24genes(60%)thathaveawiderange METHODSSUMMARY
of functions in the immune system: pattern recognition receptors, DNAwasextractedfromtheLaBran˜a1toothspecimenwithapreviouslypub-
intracellular adaptor molecules, intracellular modulators, cytokines lishedprotocol2.Indexedlibrarieswerebuiltfromtheancientextractandsequenced
and cytokine receptors, chemokines and chemokine receptors and ontheIlluminaHiSeqplatform.ReadsgeneratedweremappedwithBWA27tothe
effectormolecules. Interestingly,four outofsixSNPs fromthe first humanreferencegenome(NCBI37,hg19)afterprimertrimming.Ametagenomic
categoryareintracellularreceptorsofviralnucleicacids(TLR3,TLR8, analysisandtaxonomicidentificationwasgeneratedwiththeremainingreads
usingBLAST2.2.271andMEGAN4(ref.28)(ExtendedDataFig.9).SNPcalling
IFIH1(alsoknownasMDA5)andLGP2)25.
wasundertakenusingaspecificbioinformaticpipelinedesignedtoaccountfor
Finally,toexplorethefunctionalregulationofthegenome,wealso
ancientDNAerrors.Specifically,thequalityofmisincorporationslikelycausedby
assessedtheLaBran˜a1genotypeatallexpressionquantitativetraitloci
ancientDNAdamagewasrescaledusingthemapDamage2.0software29,andaset
(eQTL) regions associated to positive selection in Europeans (Sup- ofvariantswithaminimumreaddepthof4wasproducedwithGATK30.Analyses
plementaryInformation).Themostinterestingfindingisarguablythe includingPCA10,Outgroupf16andDstatistics17wereperformedtodeterminethe
3
predictedoverexpressionofeightimmunitygenes(36%ofthosewith populationaffinitiesofthisMesolithicindividual(SupplementaryInformation).
13 MARCH 2014 | VOL 507 | NATURE | 227
©2014Macmillan Publishers Limited. All rights reserved
RESEARCH LETTER
OnlineContentAnyadditionalMethods,ExtendedDatadisplayitemsandSource 23. Sturm,R.A.etal.AsingleSNPinanevolutionaryconservedregionwithinintron86
Dataareavailableintheonlineversionofthepaper;referencesuniquetothese oftheHERC2genedetermineshumanblue-browneyecolor.Am.J.Hum.Genet.
sectionsappearonlyintheonlinepaper. 82,424–431(2008).
24. Walsh,S.etal.TheHIrisPlexsystemforsimultaneouspredictionofhairandeye
Received22October;accepted17December2013. colourfromDNA.ForensicSci.Int.Genet.7,98–115(2013).
25. Aoshi,T.,Koyama,S.,Kobiyama,K.,Akira,S.&Ishii,K.J.Innateandadaptive
Publishedonline26January2014.
immuneresponsestoviralinfectionandvaccination.Curr.Opin.Virol.1,226–232
(2011).
1. Keller,A.etal.NewinsightsintotheTyroleanIceman’soriginandphenotypeas
26. Moresco,E.M.Y.,LaVine,D.&Beutler,B.Toll-likereceptors.Curr.Biol.21,
inferredbywhole-genomesequencing.NatureCommun.3,698(2012).
R488–R493(2011).
2. Sa´nchez-Quinto,F.etal.Genomicaffinitiesoftwo7,000-year-oldIberianhunter-
27. Li,H.&Durbin,R.FastandaccurateshortreadalignmentwithBurrows–Wheeler
gatherers.Curr.Biol.22,1494–1499(2012).
transform.Bioinformatics25,1754–1760(2009).
3. Skoglund,P.etal.OriginsandgeneticlegacyofNeolithicfarmersandhunter- 28. Huson,D.H.,Mitra,S.,Ruscheweyh,H.-J.,Weber,N.&Schuster,S.C.Integrative
gatherersinEurope.Science336,466–469(2012). analysisofenvironmentalsequencesusingMEGAN4.GenomeRes.21,
4. Laland,K.N.,Odling-Smee,J.&Myles,S.Howcultureshapedthehumangenome: 1552–1560(2011).
bringinggeneticsandthehumansciencestogether.NatureRev.Genet.11, 29. Jo´nsson,H.,Ginolhac,A.,Schubert,M.,Johnson,P.L.F.&Orlando,L.
137–148(2010). mapDamage2.0:fastapproximateBayesianestimatesofancientDNAdamage
5. Rasmussen,M.etal.AncienthumangenomesequenceofanextinctPalaeo- parameters.Bioinformatics29,1682–1684(2013).
Eskimo.Nature463,757–762(2010). 30. McKenna,A.etal.TheGenomeAnalysisToolkit:aMapReduceframeworkfor
6. Rasmussen,M.etal.AnAboriginalAustraliangenomerevealsseparatehuman analyzingnext-generationDNAsequencingdata.GenomeRes.20,1297–1303
dispersalsintoAsia.Science334,94–98(2011). (2010).
7. VidalEncinas,J.M.&PradaMarcos,M.E.Loshombresmesol´ıticosdeLaBran˜a-
Arintero(Valdelugueros,Leo´n)(Leo´n:JuntadeCastillayLeo´n,2010). SupplementaryInformationisavailableintheonlineversionofthepaper.
8. Overballe-Petersen,S.,Orlando,L.&Willerslev,E.Next-generationsequencing
AcknowledgementsTheauthorsthankL.A.GrauLobo(MuseodeLeo´n)foraccessto
offersnewinsightsintoDNAdegradation.TrendsBiotechnol.30,364–368(2012).
theLaBran˜aspecimen,M.RasmussenandH.Schroederforvalidinputintothe
9. Allentoft,M.E.etal.Thehalf-lifeofDNAinbone:measuringdecaykineticsin158
experimentalwork,andM.RaghavanforearlyaccesstoMal’tagenomedata.
datedfossils.Proc.R.Soc.BBiol.Sci.279,4824–4733(2012). SequencingwasperformedattheDanishNationalHigh-ThroughputDNA-Sequencing
10. Patterson,N.,Price,A.L.&Reich,D.Populationstructureandeigenanalysis.PLoS Centre,UniversityofCopenhagen.ThePOPRESdatawereobtainedfromdbGaP
Genet.2,e190(2006). (accessionnumber2038).Theauthorsaregratefulforfinancialsupportfromthe
11. Nelson,M.R.etal.Thepopulationreferencesample,POPRES:aresourcefor DanishNationalResearchFoundation,ERCStartingGrant(260372)toTM-B,and
population,disease,andpharmacologicalgeneticsresearch.Am.J.Hum.Genet. (310372)toM.G.N.,FEDERandSpanishGovernmentGrantsBFU2012-38236,the
83,347–358(2008). SpanishMultipleSclerosisNetowrk(REEM)oftheInstitutodeSaludCarlosIII(RD12/
12. Novembre,J.etal.GenesmirrorgeographywithinEurope.Nature456,98–101 0032/0011)toA.N.,BFU2011-28549toT.M.-B.,BFU2012-34157toC.L.-F.,ERC
(2008). (MarieCurieActions300554)toM.E.A.,NIHNRSApostdoctoralfellowship
13. Anintegratedmapofgeneticvariationfrom1,092humangenomes.Nature491, (F32GM106656)toC.W.K.C.,NIH(R01-HG007089)toJ.N.,NSFpostdoctoral
56–65(2012). fellowship(DBI-1103639)toM.D.,theAustralianNHMRCtoR.A.S.andapredoctoral
14. Surakka,I.etal.Founderpopulation-specificHapMappanelincreasespowerin fellowshipfromtheBasqueGovernment(DEUI)toI.O.
GWAstudiesthroughimprovedimputationaccuracyandCNVtagging.Genome
Res.20,1344–1351(2010). AuthorContributionsC.L.-F.andE.W.conceivedandleadtheproject.M.E.P.and
15. Raghavan,M.etal.UpperPalaeolithicSiberiangenomerevealsdualancestryof J.M.V.E.providedanthropologicalandarchaeologicalinformation.O.R.andM.E.A.
NativeAmericans.Nature505,87–91(2014). performedtheancientextractionsandlibraryconstruction,respectively.I.O.,M.E.A.,
16. Reich,D.,Thangaraj,K.,Patterson,N.,Price,A.L.&Singh,L.ReconstructingIndian F.S.-Q.,J.P.-M.,S.R.,O.R.,M.F.-C.andT.M.-B.performedmapping,SNPcalling,mtDNA
populationhistory.Nature461,489–494(2009). assembly,contaminationestimatesanddifferentgenomicanalysesontheancient
genome.I.O.,F.S.-Q.,G.S.,C.W.K.C.,M.D.,J.A.R.,J.Q.,O.R.,U.M.M.andA.N.performed
17. Green,R.E.etal.AdraftsequenceoftheNeandertalgenome.Science328,
functional,ancestryandpopulationgeneticanalyses.R.N.andJ.N.coordinatedthe
710–722(2010).
ancestryanalyses.M.G.N.,R.A.S.andP.S.coordinatedtheimmunological,
18. Perry,G.H.etal.Dietandtheevolutionofhumanamylasegenecopynumber
pigmentationandselectionanalyses,respectively.I.O.,M.E.A.,T.M.-B.,E.W.andC.L.-F.
variation.NatureGenet.39,1256–1260(2007).
wrotethemajorityofthemanuscriptwithcriticalinputfromR.N.,M.G.N.,J.N.,R.A.S.,P.S.
19. Grossman,S.R.etal.Identifyingrecentadaptationsinlarge-scalegenomicdata.
andA.N.
Cell152,703–713(2013).
20. Lamason,R.L.etal.SLC24A5,aputativecationexchanger,affectspigmentationin AuthorInformationAlignmentdataareavailablethroughtheSequenceReadArchive
zebrafishandhumans.Science310,1782–1786(2005). (SRA)underaccessionnumbersPRJNA230689andSRP033596.Reprintsand
21. Norton,H.L.etal.Geneticevidencefortheconvergentevolutionoflightskinin permissionsinformationisavailableatwww.nature.com/reprints.Theauthorsdeclare
EuropeansandEastAsians.Mol.Biol.Evol.24,710–722(2007). nocompetingfinancialinterests.Readersarewelcometocommentontheonline
22. Sturm,R.A.&Duffy,D.L.Humanpigmentationgenesunderenvironmental versionofthepaper.Correspondenceandrequestsformaterialsshouldbeaddressed
selection.GenomeBiol.13,248(2012). toE.W.([email protected])orC.L.-F.([email protected]).
228 | NATURE | VOL 507 | 13 MARCH 2014
©2014Macmillan Publishers Limited. All rights reserved
LETTER RESEARCH
ExtendedDataFigure1|AlignmentandcoveragestatisticsoftheLaBran˜a chromosomecoveredbyatleastonereadisshown,aswellasthemeanread
1genome. a,AlignmentsummaryoftheLaBran˜a1sequencedatatohg19 depthofallpositionsandpositionscoveredbyatleastoneread.c,Percentageof
assembly.b,Coveragestatisticsperchromosome.Thepercentageofthe thegenomecoveredatdifferentminimumreaddepths.
©2014Macmillan Publishers Limited. All rights reserved
RESEARCH LETTER
ExtendedDataFigure2|DamagepatternofLaBran˜a1sequencedreads. nucleargenome(c)andmtDNAgenome(d).Coefficientsofdetermination
a,b,FrequenciesofCtoT(red)andGtoA(blue)misincorporationsatthe59 (R2)foranexponentialdeclineareprovidedforthefourdifferentdatasets.
end(left)and39end(right)areshownforthenuclearDNA(nuDNA)(a) Theexponentialcoefficientsforthefourdatasetscorrespondtothedamage
andmtDNA(b).c,d,Fragmentlengthdistributionofreadsmappingtothe fraction(l);eisthebaseofthenaturallogarithm.
©2014Macmillan Publishers Limited. All rights reserved
LETTER RESEARCH
ExtendedDataFigure3|GeneticaffinitiesoftheLaBran˜a1genome. Tuscans,British,FinnsandCEU;andIndianGujaratifromTexas.c,Eachpanel
a,PCAoftheLaBran˜a1SNPdataandthe1000GenomesProjectEuropean showsPC1andPC2basedonthePCAofoneoftheancientsampleswiththe
individuals.b,PCAofLaBran˜a1versusworld-widedatagenotypedwiththe mergedPOPRES1FINHMsample,beforeProcrustestransformation.The
IlluminaOmni2.5Marray.ContinentaltermsmakereferencetoeachOmni ancientsamplesincludetheLaBran˜a1sampleandfourNeolithicsamplesfrom
populationgroupingasfollows:Africans,YorubaandLuyha;Asians,Chinese refs1and3.
(Beijing,Denver,South,Dai),JapaneseandVietnamese;Europeans,Iberians,
©2014Macmillan Publishers Limited. All rights reserved
RESEARCH LETTER
ExtendedDataFigure4|Allele-sharinganalysis. Eachpanelshowsthe Ajv70,Ire8,Gok4andO¨tzi).Intheupperrightofeachpanel,thePearson’s
allele-sharingofaparticularNeolithicsamplefromrefs1and3withLaBran˜a1 correlationcoefficientisgivenwiththeassociatedPvalue.
sample.ThesampleIDsarepresentedintheupperleftofeachpanel(Ajv52,
©2014Macmillan Publishers Limited. All rights reserved
LETTER RESEARCH
ExtendedDataFigure5|Pairwiseoutgroupf statistics. a,SardinianversusKaritiana.b,SardinianversusHan.c,LaBran˜a1versusMal’ta.d,Sardinianversus
3
Mal’ta.e,LaBran˜a1versusKaritiana.Thesolidlinerepresentsy5x.
©2014Macmillan Publishers Limited. All rights reserved
RESEARCH LETTER
ExtendedDataFigure6|Analysisofheterozygosity. a,Heterozygosity GBR,GreatBritain;IBS,Iberians;JPT,Japanese;LWK,Luhya;TSI,Tuscans;
distributionsofLaBran˜a1andmodernindividualswithsimilarcoveragefrom YRI,Yorubans.b,Heterozygosityvaluesin1-Mbwindows(with200kb
the1000GenomesProject(using1-Mbwindowswith200kboverlap).CEU, overlap)acrosseachchromosome.
northern-andwestern-Europeanancestry.CHB,HanChinese;FIN,Finns;
©2014Macmillan Publishers Limited. All rights reserved