Genetic Influences on Brain Gene Expression in Rats Selected for Tameness and Aggression

Inter-individual differences in many behaviors are partly due to genetic differences, but the identification of the genes and variants that influence behavior remains challenging. Here, we studied an F2 intercross of two outbred lines of rats selected for tame and aggressive behavior towards humans for more than 64 generations. By using a mapping approach that is able to identify genetic loci segregating within the lines, we identified four times more loci influencing tameness and aggression than by an approach that assumes fixation of causative alleles, suggesting that many causative loci were not driven to fixation by the selection. We used RNA sequencing in 150 F2 animals to identify hundreds of loci that influence brain gene expression. Several of these loci colocalize with tameness loci and may reflect the same genetic variants. Through analyses of correlations between allele effects on behavior and gene expression, differential expression between the tame and aggressive rat selection lines, and correlations between gene expression and tameness in F2 animals, we identify the genes Gltscr2, Lgi4, Zfp40 and Slc17a7 as candidate contributors to the strikingly different behavior of the tame and aggressive animals

A Comparison of Brain Gene Expression Levels in Domesticated and Wild Animals

Domestication has led to similar changes in morphology and behavior in several animal species, raising the questionwhether similarities between different domestication events also exist at the molecular level. We used mRNA sequencing toanalyze genome-wide gene expression patterns in brain frontal cortex in three pairs of domesticated and wild species (dogsand wolves, pigs and wild boars, and domesticated and wild rabbits). We compared the expression differences with thosebetween domesticated guinea pigs and a distant wild relative (Cavia aperea) as well as between two lines of rats selectedfor tameness or aggression towards humans. There were few gene expression differences between domesticated and wilddogs, pigs, and rabbits (30–75 genes (less than 1%) of expressed genes were differentially expressed), while guinea pigs andC. aperea differed more strongly. Almost no overlap was found between the genes with differential expression in thedifferent domestication events. In addition, joint analyses of all domesticated and wild samples provided only suggestiveevidence for the existence of a small group of genes that changed their expression in a similar fashion in differentdomesticated species. The most extreme of these shared expression changes include up-regulation in domesticates of SOX6and PROM1, two modulators of brain development. There was almost no overlap between gene expression in domesticatedanimals and the tame and aggressive rats. However, two of the genes with the strongest expression differences betweenthe rats (DLL3 and DHDH) were located in a genomic region associated with tameness and aggression, suggesting a role ininfluencing tameness. In summary, the majority of brain gene expression changes in domesticated animals are specific tothe given domestication event, suggesting that the causative variants of behavioral domestication traits may likewise bedifferent.funding agencies|Max Planck Society||European Research Council|233297|German Science Foundation (DFG)|AL 1525/1-1|CAS young scientists fellowship|2009Y2BS12|National Science Foundation of China research grant|31010022||SFRH/BPD/65464/2009|</p

Expression levels of four genes with common expression in domesticated dogs, pigs, rabbits, and guinea pigs.

<p>Blue: domesticated animals, red: wild animals. Shown are the four genes with the lowest p-values for the domestication factor across dogs, pigs and rabbits, and with expression change in the same direction in these three species as well as guinea pigs. Expression levels are from variance stabilized data, separately normalized to the median in each species pair.</p

Pairwise differential expression.

1<p>variance explained by domestication/species, mean across expressed genes.</p>2<p>absolute fold change, mean across expressed genes.</p>3<p>Fraction of all possible permutations of the domestication factor where the respective statistic is matched or exceeded.</p

Gene expression across domesticated species.

<p>A and B) For each of three models, the figure shows the number of genes that match or exceed the p-value for the domestication factor and that are expressed in the same direction in domesticated animals. The thick black line shows the real data. Each grey line shows the result from one of the possible extreme permutations (see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002962#pgen.1002962.s013" target="_blank">Note S1</a> for details). p = 1 is included to show the effect of only requiring genes to be expressed in the same direction, irrespective of significance. vsd: variance stabilized data. A) joint analyses of dogs, pigs and rabbits. B) As in A, but including guinea pigs.</p

Expression differences between tame and aggressive rats.

<p>A. Heatmap showing expression levels of DE genes. Genes were individually normalized and sorted by DE p-value, separately for genes up- and downregulated in domestication. Red (blue): lower (higher) expression. B. A QTL for tameness is located on chromosome one <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002962#pgen.1002962-Albert2" target="_blank">[25]</a>. The x-axis shows the genetic position along chromosome one in centiMorgan (cM). The F-value is a measure of the likelihood of the presence of a QTL. The dashed horizontal line is the genome-wide significance threshold. See <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002962#pgen.1002962-Albert2" target="_blank">[25]</a> for details. C. Expression differences in the tameness QTL region. Top panel: significance and location for each gene. P-values were signed so that positive (negative) values correspond to genes with higher expression in aggressive (tame) rats. Dashed lines: genome-wide 10% FDR threshold, dotted lines: p = 0.05. Lower panel: fold changes. Red: genes with FDR<10%, orange: genes with p<0.05. D: positions of <i>Dll3</i> and <i>Dhdh</i> (green vertical lines) compared to patterns of DNA polymorphism in the founder animals of the QTL pedigree used to identify the tameness QTL <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002962#pgen.1002962-Albert3" target="_blank">[49]</a>. Blue (red) line: nucleotide diversity in the tame (aggressive) founder animals.</p

Expression differences between domesticated and wild animals.

<p>A. Pairwise expression differences. Plotted for each comparison is in black on the left the mean variance explained by domestication, along with 95% confidence intervals (box whiskers) based on 10,000 bootstraps across genes. On the right in light blue is the null distribution of mean variance explained by domestication based on permutations of the domestication factor (box whiskers comprise 95% of the distribution, central horizontal bar is the median). B. For each comparison, the mean variance explained by domestication across genes is plotted as a function of sequence divergence expressed as the median fraction of nucleotides that differ between any two domesticated and wild animals in that comparison.</p

Pairwise overlap of DE genes among domestication events.

<p>Above diagonal: number of genes that are DE in both species, with Fisher's exact test p-values.</p><p>Below diagonal: Odds ratios of FET testing if DE genes in the one species predict expression direction in the other species, with Fisher's exact test p-values. Only the more significant direction is shown (see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002962#s4" target="_blank">Materials and Methods</a> for details).</p