Background
Autism Spectrum Conditions (ASC) are a group of neurodevelopmental conditions characterized by difficulties in social interaction and communication, alongside unusually repetitive behaviors and narrow interests. Classic autism and Asperger Syndrome (AS) are two subgroups of ASC. They differ in that AS (unlike classic autism) has no history of language or cognitive delay [
1]. The prevalence of ASC in the general population is around 1% [
2], with a male:female sex ratio of 4:1 in classic autism [
3], increasing to as high as 9:1 in AS [
4]. ASC are highly heritable, as indicated in three different twin studies [
5‐
7]. Despite the high heritability, they are characterized by complex patterns of inheritance where multiple genes, epigenetic and environmental factors are involved. Many loci have been implicated in the predisposition to these conditions [
8‐
10], but only a few of these loci have been well-replicated.
The oxytocin receptor (
OXTR) gene encodes a member of the class I family of G protein-coupled receptors, which contains seven transmembrane domains. It occupies 17 Kb on chromosome 3p25 and includes four exons and three introns. It is mainly expressed in the mammalian reproductive system and brain (pre-limbic circuits, nucleus accumbens, thalamus and amygdala) [
11‐
13]. Oxytocin receptor (OXTR) regulates the activity of oxytocin (OXT), a neuropeptide implicated in labor, uterine contraction and lactation. OXT is also involved in the establishment of maternal [
14] and social [
15] behaviors. The
OXT and
OXTR genes are good candidate genes for studying the genetic basis of ASC, because of their role in social-emotional behaviors [
16‐
19]. Intranasal inhalation of OXT represents a potential treatment of ASC, improving social interaction [
20] and emotion recognition [
21].
A previous genetic study from our group identified a nominal association between the single nucleotide polymorphism (SNP) rs237880 in
OXTR and autistic traits [
22]. Several other studies reported an association between genetic variations in
OXTR and ASC or related phenotypes. An association study in a Chinese population found that two SNPs (rs2254298 and rs53576) and haplotypes, including rs53576, are involved in the predisposition to ASC [
23]. These findings were partially replicated in Japanese [
24] and Caucasian [
25] populations. Yrigollen
et al. [
26] identified an association between rs2268493 and affiliative behavior in ASC in a largely (93%) Caucasian sample, while Campbell
et al. [
27] reported three SNPs (rs2268493, rs1042778, rs7632287) were nominally associated with ASC and related phenotypes. Another study conducted in a mixed-ethnicity population (84% Caucasian, 4.9% African-Canadian, 11.1% ‘other’) found that a genetic variation (rs237885) and one haplotype (rs237885-2368493) in
OXTR were associated with callous-unemotional traits, which are related to reduced empathy [
28]. Two other studies reported an association between genetic variations in
OXTR and autism-related phenotypes: empathy [
29] and mind-reading [
30]. Finally epigenetic studies show that different methylation patterns in
OXTR are involved in the predisposition to classic autism [
31] and variability of social perception [
32].
In the current study we tested for a possible association of nine SNPs in
OXTR with AS, performing a case–control study in a Caucasian population sample. We hypothesized that the SNPs in
OXTR, alone or in combination, would be associated with AS. We did not have a specific hypothesis about any particular SNP, and hence mapped the gene using multiple SNPs. This approach is useful in testing the role of a candidate gene (rather than a specific SNP), since it allows a test of genetic association directly and by proxy [
33].
Discussion
In the present study we conducted a case–control study in a Caucasian sample to test for genetic association between nine SNPs in the
OXTR gene and AS. Our results indicate a significant association between the common genetic variant rs2268493 in
OXTR and AS. These corroborate previously reported association between SNPs in
OXTR and individuals with an ASC more broadly. The two most reported SNPs in
OXTR that are associated with ASC are rs2254298 and rs53576 [
23‐
25], while rs2268493 has been significantly [
26] and nominally [
27] associated with ASC in two different studies.
In the current study we find a genetic association of rs2268493 with AS, which remains significant after Bonferroni correction. Haplotype analysis reports significant association of five haplotypes in OXTR with AS (rs2268493-rs2254298, rs2268490-rs2268493-rs2254298, rs2268493-rs2254298-rs53576, rs237885-rs2268490-rs2268493-rs2254298, rs2268490-rs2268493-rs2254298-rs53576). rs2268493 is included in all of these haplotype combinations and it localizes, along with rs2268490, in an LD block associated with AS in our sample. In the CEU population, rs2268493 localizes in an LD block with other four SNPs (rs6790151, rs6777612, rs6790467 and rs6793234).
rs2268493 is included in intron 3 of
OXTR (Figure
1) and it localizes 5930 bp upstream of the intron 3-exon 4 splice junction. Functional annotation indicates that this genetic variation alters six different transcription factor-binding sites (F-SNP, HaploReg). Among them are SOX_3 and Pou3f3, which are involved in neural development [
47,
48]. Moreover, rs2268493 and haplotypes that include this variation have been reported to be associated with ASC in earlier reports [
26,
27].
OXTR has four splice variants and three of them are protein-coding; one transcript includes four exons and three introns, another variant is composed of the first three exons (1, 2 and 3) and introns 1 and 2, and the other two variants include exon 1, intron 1 and part of exon 2 (UCSC genome browser). Genetic variations that disrupt regulatory regions can affect gene expression in a time- and tissue-specific manner. Molecular studies are needed to understand this further. rs2268493 localizes in an LD block when calculated in the general population, where four other SNPs are included. Three SNPs (rs6777612, rs6790467 and rs6793234) alter regions that regulate the chromatin states in several neuronal cell lines (such as fetal brain and substantia nigra) (HaploReg).
This is the first study which reports a statistically significant association between one SNP (rs2268493) in
OXTR with AS in a Caucasian sample, as other studies have focused on the whole spectrum of ASC. A previous study from our laboratory analyzed the genetic association between candidate genes (including
OXTR) and AS [
22] in an independent sample, while the current study represents a more in-depth analysis of the involvement of
OXTR in AS. Campbell
et al. have previously reported a nominally significant association between this SNP and a narrow ASC diagnosis in family-based samples [
27]. However, another study by Li
et al. failed to identify any association between the same SNP and ASC in a Japanese population [
24]. Such differences can arise due to differences between populations studied, as well as differences in criteria for diagnoses between AS and other subgroups of ASC. It is currently unclear if these differences can be explained by a different genetic architecture underlying these conditions. One way to test this hypothesis is by investigating common variants in candidate genes in AS specifically.
A limitation of this study is the moderate sample size and so further studies are necessary to confirm our findings. Multiple genes are involved in the predisposition for ASC, interacting with epigenetic and environmental factors. For this reason a single polymorphism or haplotype is unlikely to be the sole cause of these conditions. Testing this SNP for association with endophenotypes related to ASC will be an interesting next step.
Conclusions
In summary, we report a significant association between a common genetic variant in the OXTR gene (rs2268493) with AS. Five haplotypes that include this variant are also associated with AS. This provides further evidence suggesting a role of the oxytocinergic system in the aetiology of ASC, and shows that rs2268493 in OXTR is associated with AS, a high-functioning form of ASC.
Acknowledgments
This study was funded by project grants to SBC from Target Autism Genome, the Nancy Lurie Marks (NLM) Family Foundation, the Autism Research Trust, the Medical Research Council United Kingdom, the Wellcome Trust Sanger Centre, and the Max Planck Institute for Psycholinguistics. BC was funded by the Medical Research Council United Kingdom. VW was funded by the Nehru Trust for Cambridge University, St John’s College Cambridge, and the Cambridge Commonwealth Trust, and this work was submitted in part fulfilment of the degree of MPhil at Cambridge University by VW. The study was undertaken in association with the National Institute of Health Research (NIHR) Collaborations for Leadership in Applied Health Research and Care (CLAHRC) East of England (EoE). We are thankful to Robert Plomin, Frank Dudbridge, Lindsey Kent and Ian Craig for discussions, and to Laura Murphy, Jon Breidbord, Allen Chan, Sylvia Lakatosova, Sally Wheelwright, Carrie Allison, Uma Mallya, Alex Politt, and the late Leena Peltonen for support at different stages of the project. Simon Fisher provided part funding for the project.
Competing interests
The authors declare that they have no competing interests.
Authors’ contributions
ADN analyzed the data, under the guidance of VW and BC. BC and SBC designed the study and SBC obtained funding for the study. All authors contributed to the drafting of the manuscript. All authors read and approved the final manuscript.