Runcie, D. E., Wiedmann, R. T., Archie, E. A., Altmann, J., Wray, G. A., Alberts, S. C. & Tung, J

12 Pages·2013·0.49 MB·English

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Social environment influences the
relationship between genotype and
gene expression in wild baboons
Daniel E. Runcie1,2, Ralph T. Wiedmann5, Elizabeth A. Archie6,7,
rstb.royalsocietypublishing.org Jeanne Altmann7,8, Gregory A. Wray1,2, Susan C. Alberts1,7
and Jenny Tung2,7,3,4
1DepartmentofBiology,2InstituteforGenomeSciences&Policy,3DepartmentofEvolutionaryAnthropology,
and 4DukePopulation Research Institute, DukeUniversity, Durham, NC 27708,USA
Research 5USDA, ARS,USMeat AnimalResearch Center,ClayCenter,NE68933,USA
6DepartmentofBiologicalSciences, UniversityofNotreDame, NotreDame, IN 46556,USA
7Institute ofPrimate Research, National Museums ofKenya, Nairobi, Kenya
Cite this article: Runcie DE, Wiedmann RT,
8DepartmentofEcologyand EvolutionaryBiology,Princeton University, Princeton,NJ 08544,USA
Archie EA, Altmann J, Wray GA, Alberts SC,
TungJ.2013Socialenvironmentinfluencesthe
Variationinthesocialenvironmentcanhaveprofoundeffectsonsurvivaland
relationship between genotype and reproduction in wild social mammals. However, we know little about the
gene expression in wild baboons. Phil degreetowhichtheseeffectsareinfluencedbygeneticdifferencesamongindi-
Trans R Soc B 368: 20120345. viduals,andconversely,thedegreetowhichsocialenvironmentalvariation
mediates genetic reaction norms. To better understand these relationships,
http://dx.doi.org/10.1098/rstb.2012.0345
weinvestigatedthepotential fordominancerank,social connectedness and
group size to modify the effects of genetic variation on gene expression in
One contribution of 15 to a Theme Issue thewildbaboonsoftheAmboselibasin.Wefoundevidenceforanumberof
‘Flexibility and constraint in the evolution of gene–environmentinteractions(GEIs)associatedwithvariationinthesocial
mammalian social behaviour’. environment, encompassing social environments experienced in adulthood
aswellaspersistenteffectsofearlylifesocialenvironment.Socialconnected-
ness,maternaldominancerankandgroupsizeallinteractedwithgenotypeto
Subject Areas:
influencegeneexpressioninatleastonesex,andeitherinearlylifeorinadult-
behaviour, genetics hood.Theseresultssuggestthatsocialandbehaviouralvariation,akintoother
factorssuchasageandsex,canimpactthegenotype–phenotyperelationship.
WeconcludethatGEIsmediatedbythesocialenvironmentareimportantin
Keywords:
the evolution and maintenance of individual differences in wild social
gene–environment interaction, dominance
mammals,includingindividualdifferencesinresponsestosocialstressors.
rank, Amboseli baboons, social connectedness,
allele-specific gene expression
1. Introduction
Author for correspondence:
Jenny Tung Thesocialinteractionsandsocialstructuresthatcharacterizegroup-livingmam-
e-mail: [email protected] malsarenotonlyproductsofadaptivechange,butcanthemselvesinfluencethe
evolutionaryprocess.Forexample,behaviouralpatternsthatgovernmatingand
dispersalaredirectlyreflectedinpatternsofpopulationgeneticstructure[1–5].
Social behaviour thus affects the distribution of genetic variation upon which
selection can act. Social behaviour also shapes the environment experienced
by individual animals within a social group. Low-status versus high-status
individuals,orsociallyintegratedversussociallyisolatedanimalsmaydifferin
adaptively important aspects of steroid hormone physiology ([6,7]; reviewed
in Cavigelli & Chaudhry [8]), immune function [9–14] and access to mates or
otherresources[15–20].Suchdifferencesprovidescopeforsocialbehaviourto
createnewsourcesofenvironmentalselectivepressure.Indeed,thisobservation
ledWest-Eberhard[21]toarguethatevolutionarytransitionstoobligatesociality
oftenalterthesetoftraitsshapedbystrongselection.
Muchofwhatweknowabouttheevolutionaryimpactofsocialbehaviour
Electronic supplementary material is available
insocialmammalshasarisenfromfieldstudies,whichhaveproduceddetailed
at http://dx.doi.org/10.1098/rstb.2012.0345 or
illustrations of the relationship between social structure and genetic structure
via http://rstb.royalsocietypublishing.org. [22,23] and the impact of social interactions on fitness [24,25]. By contrast, we
knowfarless aboutathirdpotential effectofsocialbehaviouronthegenetics
&
2013TheAuthor(s)PublishedbytheRoyalSociety.Allrightsreserved.
Downloaded from http://rstb.royalsocietypublishing.org/ on December 31, 2014
of these species: the role of the social environment in shaping Genetic effects on gene expression that behave in this 2
genetic reaction norms. Specifically, we know little about manner (often referred to as cis-regulatory variants) can
whether and to what extent the social environment, similarly thereforebedetectedbymeasuringallele-specificgeneexpres- rstb
to otherenvironmentaleffects,canproducenormsofreaction sion (ASGE), which measures differences in gene expression .ro
y
that differ for individuals of different genotypes (i.e. gene– between the two alleles of a gene, within each individual als
o
environmentinteractions,GEIs[26]).Viewedfromacomplemen- [40,41]. ASGE assays therefore capture the ratio of gene c
ie
tary perspective, we also do not know the degree to which expression between two alleles in the same environmental ty
p
u
physiological changes in response to the social environment andgeneticbackground(becausethetwoallelesarecontained b
lis
arecontingentongenotype.GEIsinvolvingthesocialenviron- withinthesameindividual).Importantlyforthisstudy,ASGE h
in
mentarethusimportantfortworeasons.First,GEIsmayhelp levelscanalsoindicatethepresenceofGEIswhendifferences g.o
us understand how genetic differences among individuals inASGEacrossindividualsareassociatedwithenvironmental rg
affectsusceptibilitytoselectivelyrelevantsocialenvironmental variation. Specifically, GEIs are implicated in cases in which
P
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conditions.Second,byalteringhowgeneticvariationistrans- ASGEvaluesforagivengenotypevaryacrossenvironments, il
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lated into trait variation, such GEIs may alter the strength of implying that environmental exposure changes the relative ra
n
s
selectiononthegeneticvariantsthemselves. amountsofgeneexpressiondrivenbythetwoallelespresent R
AtleasttwolinesofevidencesuggestthatGEIsinvolving inastudysubject[42–44]. So
c
social environmental effects are likely to arise in natural Here, we took advantage of ASGE measurements B
3
animal populations. First, datafrom a range of species indi- to identify genes for which gene expression is affected by 6
8
:
cate that GEIs often involve environmental variation that cis-regulatory variation and to detect GEIs. We asked two 2
0
1
has large direct effects on fitness. Low-quality or quantity sets of questions. First, we tested for evidence that several 2
0
3
offoodresources,forexample,altersgeneticeffectsonsexu- aspects of the social environment are involved in naturally 4
5
ally selected traits in collared flycatchers [27], body size in occurringGEIs,focusingondominancerank,socialconnect-
blue tits [28] and lifespan in Drosophila [29–31]. Similarly, edness and group size as the key social environments of
in plants, classical ecological stressors such as drought and interest. To place these analyses in context, we also tested
leaf damage influence genetic effects on flowering time [32]. for interactions between genotype and age, and between
Because the social environment impacts fitness for many genotype and sex, two interactions that are distinct from
social mammals, it might also participate in GEIs. Second, (although potentially related to) social environmental GEIs.
studiesofcaptiverhesusmacaqueshaveindicatedthepoten- Second, we investigated whether GEIs were more likely to
tialforsociallymediatedGEIstotakeplace[33].Forinstance, be associated with early life social environmental variables,
in male rhesus macaques, early social environment (rearing in aggregate, rather than with adult social environments,
of male infants with their mothers versus rearing of male and whetherone sex was more likely to experience rank- or
infantswithsame-agepeers)appearstomediategeneticeffects social connectedness-related GEIsthantheother.
onadultaggression[34],alcoholconsumption[35]andstress
hormone physiology [36]. Whether similar effects occur in
2. Methods
the context of natural variation in the social environment,
however,remainsunknown.
(a) Study subjects
Wesetouttotestthispossibilitybytakingadvantageofa
long-term study of a well-characterized population of social Studysubjects were 96 members (50 females and 46 males) of a
natural population of baboons monitored by the Amboseli
mammals: the baboons of the Amboseli basin of Kenya.
BaboonResearchProjectintheAmboselibasin,Kenya.Thispopu-
The Amboseli baboons have been under continuous study
lationconsistsprimarilyofyellowbaboons(Papiocynocephalus)with
forover 41 years [37], providing an opportunity to focus on
somehybridadmixturefromimmigrationofanubisbaboons(Papio
aspects of the social environment of known importance to
anubis)fromoutsidethebasin[45,46].Ninety-oneofthe96individ-
these animals [7,9,15,19]. We combined detailed observa-
ualsincludedinthisstudyweremembersofoneoffiveintensively
tional data on dominance rank, social connectedness and studied socialgroups (i.e. ‘study groups’) sampled between 2005
group size with new data on genetic variation in the same and2009.Theotherfiveindividualswerebornintostudygroups
set of individuals. butemigratedtonon-studygroupasadultsandweresampledin
Wealsogathereddataongeneexpressionvariationasthe those non-study groups. All study subjects were recognized on
phenotype of interest for testing for GEIs. We chose gene sightbyobserversbasedonuniquephysicalcharacteristics.
expressionlevels,becausetheyrepresentaccessiblequantita- For most individuals (n¼78), social environmental infor-
mation was available for both early life and adult life (close to
tive traits that can be readily measured at multiple loci, are
thetimeofdarting),asaconsequenceofnear-dailybehavioural
responsiveto social environmental variation[11,38],and are
anddemographicmonitoring.Forasubsetofmaleswhoimmi-
influenced byGEIs. Forexample, for47percent of genes in
grated into the study population as adults (n¼15), data were
the yeast genome, the effects of genetic variation on gene
missing on early life social environment, and for three females,
expression levels depend on feeding substrate (glucose or
maternal rank and social connectedness data were missing
ethanol). That is, differences in gene expression between because of sparse datacollection during theirearly lives. Birth-
yeaststrainswereeitherpresentinonlyonefeedingcondition, dates and ages for the majority of individuals were known
orwerelargerinoneconditionthantheother[39].Wealsotook within several days’ error (n¼81); for immigrant males, birth-
advantage of the fact that genetic variants that affect gene dates were estimated based on known patterns of age-related
expression often lie close to the genes they regulate, on the changeinphysicalcharacteristics[47].
same physical chromosome. Hence, the maternally inherited
allele ‘controls’ gene expression of the maternally inheri- (b) Collection of blood samples
ted copy of the gene, and the paternally inherited allele Toobtainbloodsamplesforgeneexpressionanalysisandgeno-
‘controls’ gene expression of the paternally inherited copy. typing,studysubjectswereanaesthetizedwithaTelazol-loaded
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dart using a handheld blowpipe. Adult animals were darted (grooming, being groomed and proximity data) separately 3
opportunistically,resultinginanoverallsamplethatwasrandom- against the number of point samples per adult female per day
iwzeedawnaitlhysreedspheecrtet,oeaxgcee,pstexthaantdinthaedednivtiiornontmoeonntlaylcdhaarrtainctgeraisdtuiclst tshoeciasltucdoynnseucbtjeedcntewssasasinthaegmiveeanngvraoluupe.oFfinthalelyr,eswideucaalslcuolfattehde rstb.ro
(post-pubertal)animals,wealsoavoidedfemaleswithdependent respective models for grooming, being groomed and proximity y
a
infants andpregnant females beyond the first trimesterof preg- forthefocalindividual.NotethatwhileSCI-Fwasaconsequence lso
c
nancy. Following anaesthetization, study subjects were quickly ofinteractions betweenfocalfemales andbothadult malesand ie
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transferredtoaprocessingsitedistantfromtherestofthegroup. adult females, SCI-M reflects interactions between focal males p
u
b
Blood samples for gene expression analysis were collected by andadultfemales. lis
h
drawingwholebloodintoPaxGeneVacutainertubes(BDVacu- For measures of social connectedness during early life, we in
g
tainer),andbloodsamplesforsequencingandgenotypingwere used the focal individual’s mother’s social connectedness value .o
collected into BD Vacutainer EDTA tubes. Following sample (SCI-F), during the year the focal individual was conceived rg
collection, study subjects were allowed to regain consciousness (i.e. if the focal individual was conceived when its mother was
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inacoveredholdingcageuntilfullyrecoveredfromtheeffectsof 4.5 years old, then we used theyear interval from age 4 to age il
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the anaesthetic. They were then released within view of their 5 for the mother). In 15 cases for natal individuals, maternal ra
n
social group; all subjects promptly rejoined their respective SCImeasureswerenotavailableforthetimeintervalsurround- s
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groupsuponrelease,withoutincident. ing conception. We then used the closest available measure of S
o
Blood samples were stored for no more than 3 days in an SCI for that individual’s mother, provided it overlapped the c
B
evaporatively cooled charcoal structure at Amboseli, which calendar year before or after the year of the focal individual’s 3
6
maintains a daily maximum temperature of 20–258C. They conception (n¼6; otherwise, the SCI was considered missing 8:
were then shipped to Nairobi, where they were frozen at data:n¼9).Formeasuresofsocialconnectednessinadulthood, 20
1
2208C until transport to the USA. Our previous work has weconsideredtheSCI-MorSCI-Fvaluefortheage–yearover- 20
3
demonstratedstabilityofASGEmeasurementsunderthesecon- lapping the date each individual was darted (n¼83), or the 45
ditions [44,48]. For pyrosequencing assays, RNA was extracted closest available measure of SCI to the dart date, within 1 year
using the PaxGene RNA blood kit (Qiagen) and reverse tran- (n¼3forfemalesandn¼2formales).
scribed into cDNA (High Capacity cDNA Archive Kit; Applied
Biosystems). For genotyping and sequencing, DNA samples
(iii) Group demography
wereextractedusingtheDNeasyDNAextractionkit (Qiagen).
To capture social competition for resources and availability of
mates,wemeasuredthenumberofadultspresentinanindividu-
(c) Social environmental effects
al’s social group either at the month of birth (to measure the
(i) Dominance rank influence of early life group demography) or at the month of
darting(tomeasuretheinfluence ofgroupdemographyduring
Inbaboons,dominancerankofmatureindividualsislinearwithin
adulthood).
sexesandismeasuredbytheabilityofdominantindividualsto
consistently win agonistic encounters with their subordinates.
Adultdominance ranks inAmboseliare assigned on a monthly (d) Pyrosequencing assay development
basis,separatelyforeachsex,basedontheoutcomesofallpairwise
WemeasuredASGEusingpyrosequencingonaPYROMARKQ96
encounters during that month. Baboons do not achieve adult
MD instrument. This approach depends on the presence of at
dominance ranks until they are 2–4 years old (for females) or
least one single nucleotide polymorphism (SNP) in the tran-
6–8 years old (for males) [49,50]. Hence, an individual’s status
scribed region of a target gene (the ‘assay SNP’), which allows
during early life largely reflects the status of its mother, which
a PCR-based assay to discriminate between the twovariants of
we term its maternal dominance rank. Forour measure of rank
thetargetgene(thesevariantsneednotbefunctionallydifferent
effects during earlylife, we therefore considered maternal dom-
intheproteintheyproduce).InheterozygotesfortheassaySNP,
inance rank, measured as the rank of each focal individual’s
ASGEcanthenbemeasuredasthelog -transformedratioofthe
motherduringthemonththatindividualwasconceived[51].As 2
signal derived from one variant of the target gene versus the
ourmeasureofrankinadulthood,weusedthesex-specificdom-
signal derived from the alternative variant of the target gene
inance rank for each individual, assigned in the month that
[40].Theadvantageofthisapproachisthatcorrelationsbetween
individualwasdarted.
ASGE and environmental variation indicate GEIs, not simply
changes in total gene expression; in other words, they indicate
(ii) Social connectedness
differencesintheproportionalexpressionoftwodifferentalleles
Social connectedness measures (SCI-M for males and SCI-F for asafunctionofchangesinthesocialenvironment.Alimitationis
females) capture the degree to which individuals are socially thatASGEmeasurementscanonlybetakenforthosegenesthat
integratedwithotherindividualsintheirgroups.Wecalculated harbouracommonSNPinatranscribedregion;ifsuchaSNPis
one SCI value per individual per year of age, as a composite notavailable,thenthismethodwillnotwork.
indexofthefrequencytheindividualwasgroomedandgroomed Becauseofthislimitation,webeganourassaydevelopment
others (for males) at that age; for females, we also included efforts using a large initial set of 166 loci (figure 1). This set
whether the individual was in close proximity to others [19]. was chosen because they were likely to be expressed in our
Specifically,weidentifiedthenumberoftimesthefocalindividu- samples(i.e.inblood)andbecausetheyscoredhighlyonapre-
alwasgroomedbyanotheradult,thenumberoftimesthefocal dictivealgorithmforcommonASGE[53].Wealsoaddedseveral
individual groomed another adult and (in the case of females) locibecausetheyhadpreviouslybeenstudiedinassociationwith
thenumberoftimesthefocalanimalwasthenearestneighbour geneexpressionvariationinhumansorotherprimates.Thisset
of an adult female (within 5m) [52]. These counts were not wasthenfilteredtothosegenes(i)forwhichwecouldgenerate
directlycomparableacrossgroupsofdifferentsizesbecausethe high-qualitySangersequencefromputativetranscribedregions,
number of observations per animal was reduced in larger based on primers derived from the then-current draft baboon
groups relative to smaller groups. Hence, to control for these genome sequence (Pham1.0; 18 genes failed this filter); (ii) that
differences in observer intensity across groups, we collated the harboured one or more common SNPs in these regions, based
same data for all other same-sex adults alive in the population on Sanger sequencing runs from 10 to 12 unrelated Amboseli
during the same interval. We then regressed each measure individuals (33 genes failed this filter; note that we did not
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(a) (b) (c) 4
initial gene set
asmsaeya sSuNreP A hSetGerEo zfoyrg oaltles rstb
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166 genes y
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successful pyrosequencing ty
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assay designed ub
lis
identify cis-regulatory test for social environment– hin
89 genes sites associated with all 34 mediated GEIs across g.o
ASGE (elevated ASGE in genes all ASGE-associated genes rg
heterozygotes)
ASGE measured in a Ph
test set of individuals ilT
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54 genes 34 genes s
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resequence 8–10 kb c
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no common common regulatory DNA around 3
6
ASGE ASGE detected the gene TSS in all 8:
2
individuals 0
1
2
0
3
Figure1.Overallworkflow.(a)AninitialsetofgeneswasscreenedforthoselociforwhichwecouldperformASGEmeasurementsandthenforwhichwedetected 45
common ASGE within an initial test set of Amboseli individuals. (b) Genesthat exhibited common ASGE (n¼34) were subjected to ASGE measurements forall
ASGEassaySNPheterozygotesandapproximately7–9kboftheputativecis-regulatoryregionwasresequencedtoidentifygeneticeffectsongeneexpression(see
TableS2forexactnumbers).(c)ThesegeneswerethenanalysedjointlytotestforevidenceofGEIsinvolvingeachenvironmentofinterest.Thenumberofgenes
that survived each progressive screening set is shown in bold at each transition between steps.
sequencealltranscribedregionsforeachgene);and(iii)forwhich individuals in the study sample that were heterozygous for
bothvariantsofapotentialassaySNPweredetectableinASGE the assay SNP. For each individual–gene combination, we ran
assays,basedonsuccessfulamplificationoftheregionsurround- four replicate cDNA measurements and two replicate gDNA
ing a target assay SNP from cDNA and good pyrosequencing measurements on two separately prepared pyrosequencing
signal strength (22 genes failed this filter). An additional four plates. After log -transforming the relative intensities of the
2
geneswerefilteredduetounacceptablyhighvarianceacrosstech- two alternative alleles at the assay SNP, we performed three
nicalreplicates. Afterfiltering,we measuredASGE for89genes levels of quality filtering. First, we removed measurements in
(figure1).Thisgenesetwasenrichedforgenesinvolvedinimmun- which one of the two alleles was detected at low intensity (less
ity, as cell types found in blood play important roles in the than 20 units; assay variance is higher for low-intensity meas-
immuneresponse. urements because ASGE measurements are ratios). Second,
we removed outlier gDNA measurements (approx. 10% of all
measurements), conservatively identified as those deviating by
(e) Allele-specific gene expression measurements
more than 0.5 log units from the plate-specific median for all
2
To test for GEIs, we further restricted our analysis to genes gDNA measurements. This approach corrected for potential
that exhibited common ASGE, which signified the presence of assay failures. If all gDNA measurements for an individual–
common segregating genetic variation that affects gene expres- genecombinationwereoutliers,thentheindividualwasremoved
sion levels. This filter was necessary because our interest lay in from the analysis for that gene. Finally, we averaged the log -
2
testing whether thesocial environment modifiesthe magnitude transformedcDNAmeasurementsandgDNAmeasurementsfor
of ASGE, and we did not have power to detect such effects for each individual–gene combination, and corrected the cDNA
genesthat rarelyexhibited non-zeroASGE. To identify cases of measurementsbysubtractingthemean gDNA log -transformed
2
common ASGE, we genotyped the assay SNPs identified for ratio.ThisisstandardpracticeforassessingASGEusingpyrose-
each gene to identify heterozygous individuals. We then tested quencing [40,41,48]: the idea isthat some level of technical bias
for common ASGE in six to eight individuals, based on four maybeinherenttoanASGEassayitself,andthisbiascanbecor-
replicate cDNA PCRs (to measure gene expression) and two rectedbasedonestimatingthemagnitudeofthebiasfromgDNA
replicate genomic DNA (gDNA) PCRs (to control for technical samples(whichhaveaknownratio;1:1inmostcases).Thispro-
bias in the relative signal strength for the two alleles) for each cedurealsocorrectsforplateeffects(i.e.systematicallyhigheror
individual. We log2-transformed the ratio of the signal strength lower signal from one of the two alternative bases on a specific
fromthetwoallelesforeachreactionandtestedwhetherthedis- plate)becausetheyaffectbothcDNAandgDNAmeasurements.
tributionofASGEvaluesobtainedfromcDNAdifferedfromthe AftercorrectingcDNAmeasurementswithgDNAmeasurements
corresponding distribution obtained from gDNA (two-tailed, on the same plate, these plate effects are removed. Following
non-parametric Wilcoxon-summed ranks test, with significance these three quality-filtering steps, we obtained a single measure
assessedviapermutation,followingTungetal.[44]).Thisprocedure of corrected ASGE for each individual–gene combination (see
allowed ustoexcludecasesinwhich ASGEmeasurementsfrom theelectronicsupplementarymaterial,tableS1forasummaryof
cDNAwerelargelyindistinguishablefromthesameassayrunon numbersofindividualsassayedforeachgene).
gDNA,indicatingabsentorrareASGEintheAmboselibaboons.
Based on this procedure, we identified 35 genes that puta-
(f) Cis-regulatory genotyping
tivelyshowedcommonASGE.Uponfurtherevaluation(see§3),
we believe one of these genes (CYP17A1) was a false positive; Sequence variants that influence gene expression differences in
hence, 34 of these genes formed the core of the remainder of cis tend to be clustered near transcription start sites (TSSs). We
our study. For each of these genes, we measured ASGE in all thereforefocusedontheseregionstosearchforsequencevariants
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that might help explain ASGE in the Amboseli population. To alleles of a gene similarly. An environmental effect involved in 5
identify and genotype these putative regulatory variants, we a GEI, on the other hand, should influence the expression
ufoslelodwaedtarbgyetheingrhic-thhmroeungthappuptrosaecqhue(nAcginilgentonSurtehSeeleIclltu)m[5in4]a, liedveenltsitoyfothfetthweocaisl-lerelegsuolaftoarygenveardiaifnfte(rse)ntlliyn,kdedepetondtinhgatonalltehlee rstb.ro
HiSeq 2000 platform. For each of the 34 genes with common [42–44]. Under this model, individuals heterozygous for a y
a
ASGE, we identified an approximately 7–9kb target (see the functional cis variant will exhibit different levels of ASGE lso
c
electronicsupplementarymaterial,tableS2)coveringtheregion depending on the environment (provided that this variant is, ie
ty
upstream of the gene TSS and the region between the TSS and at least to some degree, linked to the transcribed SNP used in p
u
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its downstream translation start site. To do so, we used the theASGEassayandtheenvironmentisnotconfoundedbygen- lis
h
rhesus macaque genome (rhemac2), because the draft baboon etic background effects: the social environments we considered in
g
genome available at the time (Pham1.0) contained too many are unlikely to be confounded by genetic background in our .o
gaps and missing regions to adequately cover these regions. sample, as they are poorly correlated with measures of both rg
Importantly, rhesus macaque and baboon have highly similar admixture and kinship; see electronic supplementary material,
P
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sequence in these regions, and cross-species sequence capture figures S1 and S2). We took advantage of this property to test il
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has been validated in primates, including more distant pairs whether models of ASGE that included GEIs involving the ra
n
thanmacaque–baboon[55,56].Wethendesigned120bpbiotin- social environments of interest were favoured over modelsthat s
R
ylated RNA probes tiled to cover our target regions at a mean didnotincludeGEIs. S
o
2(cid:2) coverage. We used these probes to capture our regions of ASGEitselfsignifiesthepresenceoffunctionalcis-regulatory c
B
interest. We added a unique 6bp barcode (Agilent) to each variation,wheretheresponsiblefunctionalvariant(s)islinked,to 3
6
library,whichenabledustopoolcapturedDNAfromall96indi- somedegree,totheASGEassaySNP.Theaprioriexpectationis 8:
2
viduals and sequence a single, multiplexed sample on a single thereforethatheterozygotesfortheassaySNP(i.e.theindividu- 0
1
laneoftheHiSeq2000. als we were able to assay) are probably heterozygotes for the 20
3
Wegenerated182million,50bpreadsfromthissample(see functional site(s) as well, which is likely to be the case if the 45
the electronic supplementary material, table S3). Reads were assay SNP is closely linked to this site. Such a pattern would
generally evenly distributed across the 96 individual subjects bereflectedbynon-zeroASGElevelsforallassayedindividuals.
(median¼1.85 million reads+0.060 million reads s.d.), with Alternatively, some individuals might not be heterozygous for
the exception of two outlier individuals for whom we obtained the functional cis-regulatorysite if the assay SNPand the func-
very few reads (‘Face’ and ‘Morris’: electronic supplementary tional variant were not tightly linked (e.g. in the cases when
material, table S3). Reads were mapped to the baboon genome the assay SNP is far from the putative promoter region). This
(Panu2.0, released after the probes were designed) using the possibility, in turn, would be reflected by a pattern in which
default settings in NovoAlign (NovoCraft). Across individuals, someassayedindividuals(thoseheterozygousforthefunctional
a median of 82 per cent (median range: 60–85%) of reads SNP) exhibited non-zero ASGE, but others (homozygotes for
mapped to the genome with Phred-scaled mapping quality the functional SNP) exhibited ASGE levels close to zero. In
greater than or equal to 20. To translate between the regions thesecases,heterozygosity/homozygosityatasitemoreclosely
we targeted using rhesus macaque genome sequence and the linked to the functional SNP would better explain variance in
baboon genome assembly, we used lastz [57] and axtChain [58] ASGE levels. Because ASGE levels in homozygotes would not
to find the corresponding target regions in baboon. For 89 per correlate with environmental variation even in the presence
centofgenes,wecouldclearlyidentifyasingleregionthatcor- of GEIs, including these individuals would dilute anysignal of
responded to the size expected based on the rhesus macaque GEIs in the population. For each gene, we therefore compared
sequence fromwhich theprobesweredesigned. Forfour genes models reflecting these two alternative possibilities: either non-
(CLC, GBP1,APOBE3G and RNASE2), our probes mapped to a zeroASGEinallassayedindividuals(reflectingcloselinkagedis-
larger region than we had anticipated in the original probe equilibrium between the assay SNP and the functional site), or
design; we therefore analysed genotype data from all baboon non-zero ASGE only in heterozygotes for a putative regulatory
regionsthat were on the same chromosome asthe ASGE assay SNP (reflecting closer linkage between the functional site and
SNP and that matched the target macaque sequence, as SNPs this SNP instead of the assay SNP). We chose the best model
that were farther than expected from a gene transcription start as that which yielded the highest model r2-value. This model
site could still plausibly be functionally relevant. Note that for identified probable heterozygotes at the (unknown) functional
these four genes, we also performed SNP and genotype calls regulatory site responsible for ASGE as either heterozygotes at
based only on reads from the captured sequences that mapped a regulatory region SNP or heterozygotes for the assay SNP
uniquely to the baboon genome; we simply captured a larger itself (see the electronic supplementary material, figure S3). We
region than we had originally intended. In total, a median of used only these heterozygous individuals in subsequent GEI
72percentofhigh-quality-mappedreadsfellwithinthetargeted analyses. In all these analyses, we excluded putative regulatory
regionsinbaboon(range:24–80%). SNPs that were in apparently perfect linkage disequilibrium
We conducted variant discovery and genotyping on the with other sites for the same gene (we retained a single SNP
mapped, quality-filtered reads, using the Genome Analysis foreachcorrelatedset)andputativeregulatorySNPswithmiss-
Toolkit (GATK) ([59,60]; see electronic supplementary material ing or littledata(fewer than fiveheterozygous individuals and
for additional details). We then used the program BEAGLE [61] five homozygous individuals). Note that this process was
to impute missing genotypes in the resulting dataset. As input, completelyblindtodataonenvironmentalvariation.
weused thegenotype likelihoods produced by GATK; we then We then asked whether, foreach environmental variable of
filtered the BEAGLE results to include only genotypes with a interest, our dataset supported the potential for GEIs involving
posteriorprobabilitygreaterthan0.98. the social environment. We did so using the following nested
setofmodels(includingheterozygotesorinferredheterozygotes
ataputativefunctionalregulatorySNPonly):
(g) Identification of gene–environment interactions
M :y ¼g þe
0 ij i ij
ToidentifyGEIsinourdataset,wereasonedthatanenvironment
thathasanunconditionaleffectongeneexpression(i.e.isinde- and
pendent of cis-regulatory variation that might be associated
with the gene) should influence the expression levels of both M1:yij¼giþgi(cid:2)vjþeij;
Downloaded from http://rstb.royalsocietypublishing.org/ on December 31, 2014
whereyijisthelog2-transformed,normalizedASGEmeasurefor (a) OAS2 (b) AIM2 6
geneiinindividualj;g fitsaninterceptforeachgene;g(cid:2)v fitsa
i i j 39 13 16 11
seaenrpveaisrrioadntuemaler,enwgtarheliscvshaiorwinaebslaelsostpouemAefSodGrtEoevabcaehluinegdseefnpoere,ntdhreaeltnattgieannnged;tnahonerdmfeoaijcllaiysl atio 1 0 26 1 0 5 rstb.roya
gdeisnteri)b.uFtoerdeawchithenEv[ierioj]n¼m0enat,nedxcveaprtiadnocmeisn2ian(cdeifrfaenrekntanfdorsoecaicahl ASGE r 0 0 lsociety
connectedness (which are calculated differently for males and 2 pu
g b
femalesand havedifferent biologicalinterpretations), wetested lo−1 −1 lish
maleMsoadnedlsfemwearleesbfiotthussienpgaraatelmyaaxnidmjuomin-tlliyk.elihood criterion P– 55– P– 57– ing.o
with the function gls in R [62], after excluding genes with SN 76 SN 06 rg
fewer than five heterozygous individuals. If M wasa better fit y 17 y 96
to the data than M0, as assessed by a likeliho1od ratio test, we assa 112 assa 132 Phil
T
interpreted the data as supportive of GEI(s) arising as a con- Chr11 Chr1 ran
sequence of the social environment tested in those models. s
R
Importantly, this approach specifically tests the hypothesis that Figure2.ExamplegeneticassociationswithASGE.(a)VariationintheASGE S
o
a given social environment participates in GEIs in Amboseli, data for OAS2 is best explained by heterozygosity/homozygosity at a SNP c
B
rather than testing each gene–environment combination for contained in the captured, resequenced region upstream of the OAS2 3
6
each gene separately (which would incuran unacceptably high transcription start site (r2 for a model including the best site in the 8:
2
multiple testing burden, given our sample size). We adjusted resequencing data¼0.54 versus r2 for a model including the assay 01
for multiple testing using the false discovery rate method of SNP¼0.01). (b) Variation in the ASGE data for AIM2 is best explained by 203
Benjamini&Hochberg[63]inthefunctionp.adjustinR. heterozygosity/homozygosity at the assay SNP, and not at any SNP in the 45
Finally, we tested two hypotheses about how GEIs differ
captured, resequenced region (r2 for a model including the assay SNP¼
betweensexesandinrelationshiptothetimingofenvironmental
0.88 versus r2 for a model including the best site in the resequencing
effects.First,wetestedwhetherGEIsweremorestronglyassoci-
ated with early life social environments (maternal dominance data¼0.73). (a,b) In both panels, boxplots show the range of ASGE vari-
rank, maternal social connectedness and social group size in ation across all assayed individuals (left: ‘assay SNP’) and ASGE variation
earlylife) thanwith adultsocialenvironments (theindividual’s subdivided by a SNP in the captured, resequenced region (right: base pair
own rank, social connectedness index and group size at the coordinates for these SNPs are provided as labels). Numbers above each
timeofdarting),orviceversa.Becauserankandsocialconnect- setofboxplotsprovidethenumberofheterozygotes(red)andhomozygotes
edness were calculated separately for males and females, we (black) foreach site, and the site associated with the best ASGE partition is
tested for early versus late life GEIs separately for each sex.
highlighted in yellow. For the assay SNP, all assayed individuals are hetero-
Second, we tested whether, among sex-specific effects (domin-
zygousbecauseassaySNPheterozygosityisarequirementfortheassaytobe
ance rank and social connectedness), GEIs were more evident
performed. Boxplots represent the data distributions as follows: heavy bars
inmaleorfemalesubjects.
show the sample median; boxes cover the interquartile range of the data
Forbothtests,wecomparedthevarianceinASGEexplained
and whiskers extend to the most extreme data point (excluding outliers
foreachgeneby(i)earlyversuslateenvironmentaleffects,and
(ii) male-specific versus female-specific environmental effects, that were more than 1.5 times the interquartile range from the box;
using a two-sample paired t-test. Genes were removed from small open circles mark outliers beyond this range).
these analyses if fewer than nine individuals for the early
versus adult life comparison, or eight individuals for the male
sequencing-based estimates of genetic diversity for the same
versus female comparison (which involved fitting fewer
population(seetheelectronicsupplementarymaterial).
parameters),weretestableforeachofthetwocompetingmodels.
For 13 of the 34 genes that exhibited common ASGE, the
presenceorabsenceofASGEinanindividualbaboonwasbest
explained by heterozygosity versus homozygosity at a SNP
3. Results in the captured, resequenced regulatory region (figure 2a). In
these cases, genetic variation responsible for the observed
(a) Genetic effects on gene expression
ASGE was likely to be more closely linked to variants in the
We identified commonASGEin34blood-expressedgenes in upstreamregulatoryregionthantotheassaySNPitself(consist-
theAmboselibaboonpopulation,outof166genesweoriginally ent with the expectation that these regions are enriched for
surveyed and 89 genes for which we could reliably measure functional cis-regulatory variants). For these genes, genotype
ASGE.Wethereforeestimatethatcommonlysegregatingfunc- at theputative regulatorysite explained a large proportionof
tional cis-regulatory variants influence gene expression in overallvarianceinASGE(medianr2¼55%;range¼31–81%:
approximately 38 per cent of genes in the Amboseli baboon notethatinsubsequentGEIanalysis,wewereconcernedonly
population. This frequency agrees with a previous, smaller- withvarianceamongheterozygotes,notthevarianceexplained
scaleanalysisofthispopulation,whichyieldedanestimateof by the contrast between heterozygotes and homozygotes
36.4 per cent [44]. In addition, the putative cis-regulatory reported here). For 21 genes, heterozygosityat the assay SNP
regions near gene TSSs that we surveyed exhibited substan- best explained the patterns of ASGE in our sample, and thus
tial segregating genetic variation. Overall, we identified 3527 weincludedallassayedindividuals(allofwhomwereheterozy-
high confidence segregating sites (among 250333 total base gotesattheassaySNP)indownstreamGEIanalyses.Thatis,for
pairs for which at least 10 individuals were sequenced at a these 21 genes, the assay SNP was likely to be more closely
coverage of at least 30(cid:2)), indicating a frequency of variants linkedtothefunctionalvariantdrivingASGEthananyofthe
identified (including rare variants) of about one per 70 bp. resequenced upstream SNPs in the sample. Thus, the rese-
DiversitylevelsbasedontheseSNPswereinexcellentconcor- quenced variants explained variance in ASGE levels more
dance with estimated diversity levels from prior, Sanger poorly than a model in which all assayed individuals were
Downloaded from http://rstb.royalsocietypublishing.org/ on December 31, 2014
Table 1. Evidence for GEIs involving testedinteraction effects. 7
interaction effect p-value (females)a p-value (males) p-value (both sexes) rstb
.ro
y
social environments a
ls
o
dominance rank cie
ty
early (maternal) 0.464(31) 0.020(24) 0.304(34) pu
b
adult 0.133(31) 0.285 (30) n.a.b lis
h
in
social connectedness g
.o
early (maternal) 0.122(30) 0.237 (24) 0.108(34) rg
adult 0.014 (31) 1.1331023 (28) n.a. Ph
il
group size Tra
n
early 4.731024 (31) 0.433 (24) 0.024 (34) sR
S
adult 9.2031023 (31) 2.8831023 (30) 0.012 (34) o
c
B
non-social effects 3
6
8
age 0.0023 (31) 0.542 (30) 0.098(34) :
2
0
sex n.a. n.a. 6.5031023 (34) 12
0
3
ap-values from likelihoodratiotests comparing amodel with GEIstoamodel without GEIs are provided in each cell. Bold values are those thatsurvivemultiple 45
hypothesistestcorrection using the method of Benjamini & Hochberg [63],atafalse discovery rate of10%. Values in parentheses provide thenumberof genes
included in each test.
bSocial connectedness and dominanceranks in adulthood could not beevaluated for both sexes combined, as both statistics arecalculatedseparately for males
and females.
probably heterozygous for the (unknown) functional site (p¼1.13(cid:2)1023) and females (p¼1.41(cid:2)1022; figure3b,d),
(figure2b:thebestSNPidentifiedintheresequencedregulatory but maternal social connectedness was not associated with
regionisshownagainsttheassaySNPforcomparison).Finally, GEIs for either sex. Conversely, for dominance rank, we
foronegene,CYP17A1,weidentifiednobestSNP:neitherthe observed no signal of an individual’s own rank near the
assay SNP nor any of the resequenced, putative regulatory time of sampling. The only evidence for a rank effect was
SNPsexplainedASGE variationwell(noSNPexplainedvari- also sex-specific: maternal dominance rank was detected as
anceinASGElevelswithap-valuebelowanominalthreshold a contributor to GEIs for males (p¼1.97(cid:2)1022) but not
of0.05).Thisgene(CYP17A1)probablyrepresentsafalseposi- forfemales (p¼0.464).
tive result from our earlier, more restricted test for common
ASGEandwasexcludedfromfurtheranalysis.
(c) Sex- and timing-related differences in the
environmental components of gene–environment
(b) Environmental modification of allele-specific gene
interactions
expression levels
Finally,wetookadvantageofouranalysisofmultiplesocial
We asked whether any of the social environmental variables environmental effects to ask whether, in aggregate, social
we tested had the capacity to influence the genotype–gene environments relevant to early versus adult life stages (i.e.
expression relationship identified through our ASGE assays all three early life environments versus all three adult
(see table 1 and electronic supplementary material, tables S4 environments) tended to explain more variance in ASGE
andS5).Ofallthesocialenvironmentsweanalysed,weobserved across all measurable genes. That is, we asked whether
the most consistent GEIs in relationship to group size. The early or adult social environmental variation more consist-
numberofadultsinanindividual’ssocialgroupatthetimeof ently contributed to variance in gene expression via GEIs.
darting, which indexes competition for mates and resources, For females, early life social environmental characteristics
was associated with GEIs in males (p¼2.9(cid:2)1023), females (maternal dominance rank, maternal social connectedness
(p¼9.2(cid:2)1023)andalsojointlywhenmalesandfemaleswere and group size) and social environmental characteristics in
tested together (p¼0.012; figure 3a,c). Similarly, we found adulthood(groupsize,adultdominancerankandsocialcon-
evidence that group size at birth, the early life analogue of nectednessatdarting)wereindistinguishablewithrespectto
numberofadultsinthegroupatdarting,wasalsoinvolvedin explaining ASGE (p¼0.96; figure 4a). In males, despite our
GEIs in females (p¼4.72(cid:2)1024), but no evidence for such reduced power to detect such differences compared with
GEIsinmales(p¼0.433).Supportforthisearlylifeeffectwas females (for whom we had a larger sample size for early
weakerbutstillevidentwhenmalesandfemalesweremodelled life effects, and thus retained more genes for this analysis),
together(p¼0.024;seealsoelectronicsupplementarymaterial, we observed a trend towards a greater impact of adult
tableS4forasummaryofallmodels). social environments than early life environments on ASGE
In contrast to group size, for which both early life and (p¼0.085; figure4b).
adult exposures were associated with GEIs, adult social We performed a similar analysis to test whether female-
connectedness was associated with GEIs for both males specific social environments (SCI-F and adult dominance
Downloaded from http://rstb.royalsocietypublishing.org/ on December 31, 2014
(a) 1.0 (b) 8
males males
0.8 p = 0.030 p = 0.0371 rstb
E ratio 0.6 .royalsoc
G ie
S ty
A p
d log 2 0.4 ublishin
e g
aliz 0.2 .org
m
or P
n h
0 il
T
ra
n
s
–0.2 R
S
o
20 30 40 50 –0.5 0 0.5 1.0 c
B
3
group size at sampling male social conectedness index 6
8
:
(c) (d) 2
1.0 01
2
0
females females 3
4
0.8 p = 0.0293 p = 0.0030 5
o
ati
E r 0.6
G
S
A
g 2 0.4
o
d l
e
z
ali 0.2
m
or
n
0
–0.2
20 30 40 50 –0.6 –0.4 –0.2 0 0.2 0.4 0.6
group size at sampling female social conectedness index
Figure3.SociallymediatedGEIs.ASGElevelsthataremodifiedbyanindividual’senvironmentimplyGEIs,becausetherelativegeneexpressionlevelsofthetwo
allelesofagenechangedependingontheenvironment.Eachpaneldepicts,foroneexemplargene,achangeinASGEwithasocialenvironment(notethatourcore
questions focusedonanalysesforeach environment,acrossgenes, however).Weidentified consistentsociallymediatedGEIsrelatedtogroupsizeandsocial con-
nectedness in adulthood for both (a,b) males and (c,d) females. (a) PHF11, (b) RNASE2, (c) CD8A and (d) SLAMF7.
rank)tendedtobemoreorlessimportantthanmale-specific Our results serve as proof of principle that, like age and
social environments (SCI-M and adultdominance rank). We sex [64,65], social environmental characteristics have the
observed no evidence for sex differences in the impact of capacity to influence the relationship between genotype and
these effects (p¼0.687;figure4c). geneexpressioninwildsocialmammals.Althoughnotallof
the individual variables that we tested revealed GEIs, we
identified significant support for GEIs for each of the broad
4. Discussion categoriesofsocialenvironmentsweinvestigated:dominance
rank,socialconnectednessandgroupdemography.Ourdata
Genetic differences make important contributions to vari- thussuggestthattheimportanceofthesepredictorstosocial
ationinbehaviouralphenotypes.However,behaviouraltraits competition, survival and reproductive success extends, to
also feed back to influence population genetic structure and somedegree,togeneticreactionnormsoperatingatthemol-
evolutionary genetic change. Understanding flexibility and ecular level. Viewed from a complementary perspective,
constraint in the evolution of behaviour therefore demands generegulatoryresponsestothesocialenvironmentinsocial
thatwedevelopabetterunderstandingofthereciprocalmech- mammals [11] are therefore likely to be mediated in part by
anismsthroughwhichgenesandbehaviourarelinked.Inthis genetic differences among individuals. Given that social
study, we tested whether GEIs involving the social environ- status and the quality of social interactions are powerful
ment act as one such mechanism. To do so, we combined a predictors of survival and longevity [20], untangling these
long-termdatasetonthedemographyandbehaviourofwild interactions will be important for understanding how
baboons with novel data on ASGE variation and genotype, individualanimalsdifferinsusceptibilitytotheseeffects.
focusingonindividuallyidentifiedindividualsthathadbeen ByinvestigatingthepotentialforGEIsatmultipleloci,we
trackedoverthecourseoftheirlives. were also able to ask whether social environment-mediated
Downloaded from http://rstb.royalsocietypublishing.org/ on December 31, 2014
(a) (b) (c) 9
early biased adult biased early biased adult biased male biased female biased
–0.4 –0.2 0 0.2 0.4 –0.4 –0.2 0 0.2 0.4 –0.4 –0.2 0 0.2 0.4
GIGMZAMPA2 GIMCDAP8A2 HLAS−LFA rstb.roy
APOBEICR3FG2 IFNGABRP11 PPHRFN1P1 also
OKILAL4FSRL6 pf e=m 0a.9le6s0 CXICILLR41R40 p m= 0a.l0es85 APOBEOKCLA3FSGL6 p = 0.687 cietypu
HPLHAF−1F1 APOBECCC3RG1 CGDB8PA1 blish
PRNP RNASE2 IL4R in
IFNNAMR1I SLPAHMFF171 MCSLRC1 g.org
GBP1 NMI IL10
CXCR4 PRNP CXCR4
P
SLAMF7 OASL IFNAR1 h
CLC HLA−F NMI il
T
IL10 IRF2 ra
S100A9 GIMAP2 ns
CD8A CCR1 R
SLA SLAMF7 S
AIM2 oc
CCR1 B
3
6
Figure 4. Testing for biases in the variance explained by GEIs. In each panel, the difference in percentage variance in ASGE explained byearly life versus adult 8
:
environments(a,b)ormale-specificversusfemale-specificenvironments(c)isplottedforeachgene.(a)ThedistributionofvarianceinASGEexplainedbyearlylife 20
1
effectsversuseffectsduringadulthooddoesnotdifferinfemales(pairedt-test:p¼0.960).(b)ThedistributionofvarianceinASGEexplainedbyearlylifeeffects 20
3
versuseffectsduringadulthoodexhibitsaweaktrendsuggestingbiastowardsadultsocialenvironmentalexposuresinmales(pairedt-test:p¼0.085).(c)Forsex- 45
specificenvironmentaleffectsofdominancerankandsocialconnectedness,femalesandmalesdonotdifferinthedegreeofvarianceaccountedforbyGEIs(paired
t-test: p¼0.687).
GEIstendedtobebiasedbysexorbythetimingofenviron- will be necessary to confirm these results, they suggest that
mental exposures. We found no evidence for an overall sex earlylifesocialenvironments,whileimportant,arenotneces-
bias related to the variance in ASGE accounted for by GEIs. sarily privileged over social exposures in adulthood with
However, we did observe support for GEIs involving respecttotheireffectsongeneexpressionreactionnorms.
maternalrankformales,butnotfemales.Thisresultissome- Interestingly, male baboons arguably experience increas-
what surprising, as maternal rank is a pervasive maternal ed social environmental variability over their lifetimes
effectinfemalebaboonsthatinfluencesgrowthduringdevel- relative to females: unlike females, males often change
opment [66,67], the timing of reproductive and social social groups multiple times as adults, thus also changing
maturation in females [50,66], and the rank they achieve in theirrank,socialbondsanddemographiccontext.Maledom-
adulthood [68–70]. By contrast, while maternal rank affects inance rank is more dynamic within groups than female
male growth as it does female growth [66,67], it seems to dominance rank as well [68,69,73]. It therefore seems quite
havelessofanimpactonamale’slifehistorythaninfemales possible that the environment that a male currently experi-
[50,68], and maternal rank appearsto have little or no influ- ences, which affects immediate access to mates [15,17,74],
ence on the dominance rank that sons eventually achieve tenurelengthinagroup[47]andtestosteroneandglucocor-
[68]. However, long-term effects of maternal rank on male ticoid levels [7], may be more physiologically relevant than
physiologyhavebeenpreviouslydemonstratedintheAmbo- their experiences during early life. We conjecture that this
seli baboons: subadult sons of higher-ranking females have pattern may hold especially true for blood-expressed genes
lower glucocorticoid levels than sons of lower-ranking such as those we studied, as many of these genes are re-
mothers [51]. In the light of these previous findings, our lated to immune function that itself can vary depending on
results suggest that maternal rank-mediated GEIs in males social environmental conditions. In Amboseli, for example,
may fit into a broader pattern of persistent maternal effects higher-ranking males heal faster from wounds and injuries
on physiologythat, asyet,is poorlyunderstood. than low-ranking males [9]. Tissues that have less of a role
Possible differences in the timing of the social environ- in sensing or responding to the external environment may
mentalexposuresinvolvedinGEIsarealsosuggestedbyour be less likely to exhibit social environment-mediated GEIs.
analysis of early life versus adult life social environments. Additionally, because immune-related genes sometimes har-
Early life, especially the gestational and periparturitional bour unusually high levels of genetic variation, GEIs may
periods,hasbeenproposedasa‘sensitiveperiod’inphenotyp- alsobemoreimportantinblood.Testingthesehypotheses—
icdevelopment,withpotentiallyprofoundeffectsonlaterlife aswellaswhetherspecificclassesofgenesaresusceptibleto
traits [71,72]. This idea has garnered support from empirical GEIsandwhetherGEIsareplasticinthefaceofenvironmental
evidence tying early life exposures to disease susceptibility change—will require gathering considerably more data on a
andstressreactivity inadulthood,raising the possibilitythat widervarietyofsamples.
earlylifeenvironmentaleffectsmightbeimportantforsocial Suchworkwillbefacilitatedbynewmethodsformeasur-
environment-mediatedGEIs.Amongthefemaleswestudied, ingASGEonagenome-widescale.Genome-wideapproaches
however, the amount of variance in ASGE explained by couldextendtheapproachweusedheretofulltranscriptomes
early life social environments was indistinguishable from [75],althoughtheywouldlikelyusedifferentmethodsforboth
that explained by adult social environments. By contrast, in measuring and analysing ASGE levels. In an alternative
males,weobservedweakevidencethatadultsocialenviron- strategy, genes that are highly responsive to a given social
mentsmightexertstrongereffects.Althoughfollow-upwork environment could be tested for environment–contingent
Downloaded from http://rstb.royalsocietypublishing.org/ on December 31, 2014
geneticeffects.Indeed,recentworkhassuccessfullydiscovered of species social structure: for example, dominance rank- 10
geneticmodifiersofthegeneexpressionresponsetophysical mediated environmental variation always occurs within
stressorssuchastuberculosisinfection[76],radiationexposure baboon social groups because no two individuals can rstb
[77] and synthetic glucocorticoid treatment [78]. Intriguing- occupy the same rank. These differences raise an intriguing .ro
y
ly, these studies suggest that genetic differences contribute possibility that the evolution of complex social structures als
o
only weakly to the effects of the environment when envir- among social mammals has also had an effect on the ways c
ie
onmental effects are strong (i.e. explain a large fraction of in which genetic variation is maintained and expressed— ty
p
u
varianceingeneexpressionlevels).Incomparison,ourobser- strong motivation for investigating social environment- b
lis
vations here suggest that environmental variation only mediated GEIs in the context of long-term genetic evolution h
in
weakly modifies the effects of genotype on gene expression aswell. g.o
levelswhengenotypeeffectsarestrong.Applyingthesecomp- rg
lementary approaches together might therefore yield a more
This work was supported by the National Science Foundation P
h
completeunderstandingofthenatureandeffectsizeofsocial (IOS-0919200 to S.C.A., DEB-0846286 to S.C.A and G.A.W., and il
T
environment-mediatedGEIs. BCS-0846532toJ.A.)andtheNationalInstituteofAging(NIAR01- ra
n
Our study suggests that behaviour and social structure AG034513-01 and NIA P01-AG031719 to S.C.A.). We thank the sR
influence the function of genetic variation in wild social OfficeofthePresident,RepublicofKenya;theKenyaWildlifeService So
anditsAmboselistaffandwardens;ourlocalsponsor,theInstituteof c
mammals on a proximate (mechanistic) timescale by chan- B
PrimateResearch;theNationalMuseumsofKenya;andthemembers 3
ging its effects on gene regulation. However, like other of the Amboseli–Longido pastoralist communities. Particular 68
:
behavioural phenomena, social environment-mediated GEIs thanks go to the Amboseli fieldworkers who contribute to genetic 2
0
can also be investigated on an evolutionary timescale [79], and environmental sampling, especially R. Mututua, S. Sayialel 12
0
and K. Warutere, and to T. Wango and V. Atieno, who provided 3
and our findings raise new questions about the degree to 4
assistance with the samples and logistical support in Nairobi. 5
which socially mediated GEIs might shape evolution over
WearealsogratefulforthecontributionsofJ.Stroud,J.Gordon,and
the long term. GEIs have long been proposed, albeit with S.Morrowtothepyrosequencingandgenotypingdata,S.Mukherjee
mixed support from theoretical analyses, as a mechanism forstatisticaladvice,T.Brown-BrandlandJ.Horvathforreadingan
through which selectively relevant genetic variation might earlydraftofthemanuscript,twoanonymousreviewersfortheirhelp-
ful comments and Baylor College of Medicine (Human Genome
bemaintainedinnaturalpopulations[80–83].Suchanalyses
SequencingCenter)forthepreliminarydraftassemblyofthebaboon
have generally assumed that the relevant environmental
genome. Data included in this study were obtained in accordance
players in GEIs are structured either across space (across with the Institutional Animal Care and Use Committee protocols
populations that experience different ecological conditions) approvedbyDukeUniversityandPrincetonUniversity.Dataincluded
ortime(asecologicalconditionschangewithinapopulation). inthisstudyaredepositedinDryad(doi:10.5061/dryad.s5j81)andin
theNCBIShortReadArchive(SRP018756).Mentionoftradenames
By contrast, social environmental variation tends to be
or commercial products in this publication is solely for providing
structured among individuals, within populations or social
specificinformationanddoesnotimplyrecommendationorendorse-
groups.Unlikemanyothertypesofenvironmentalvariation, ment by the US Department of Agriculture (USDA). USDA is an
it is also frequently omnipresent, as a direct consequence equalopportunityproviderandemployer.
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