Jun Egawa

A detailed association analysis between the tryptophan hydroxylase 2 (TPH2) gene and autism spectrum disorders in a Japanese population

We conducted a detailed association analysis between the tryptophan hydroxylase 2 gene and autism spectrum disorders in a Japanese population using 19 markers, including tagging single nucleotide polymorphisms and a novel missense variation, p.R225Q, identified through exon resequencing. However, we failed to obtain supportive evidence for an association.

Interleukin 1 beta gene and risk of schizophrenia: detailed case-control and family-based studies and an updated meta-analysis.

Interleukin‐1 beta (IL‐1β) has been implicated in the pathophysiology of schizophrenia. To assess whether the IL1B gene confers increased susceptibility to schizophrenia, we conducted case–control and family‐based studies and an updated meta‐analysis.

Oxytocin receptor (OXTR) gene and risk of schizophrenia: Case–control and family-based analyses and meta-analysis in a Japanese population

OXYTOCIN (OXT), SIGNALING through its receptor (OXTR), has been implicated in the pathophysiology of schizophrenia. Thus, OXTR is a potential candidate gene for schizophrenia. The association between OXTR and schizophrenia has been tested in only one study reporting negative results using Caucasian case–control and trio samples (n = 358 and 34, respectively). To assess whether OXTR confers increased susceptibility to schizophrenia, case–control and familybased analyses and a meta-analysis were performed using Japanese case–control and trio samples (n = 1218 and 105, respectively). The present study was approved by the Ethics Committee on Genetics of the Niigata University School of Medicine, and written informed consent was obtained from all participants. The case–control subjects consisted of 544 patients with schizophrenia (290 men and 254 women; mean age, 41.8 13.6 years) and 674 mentally healthy individuals (341 men and 333 women; mean age, 38.4 10.8 years). The family-based subjects consisted of 105 trios, made up of patients (59 men and 46 women; mean age, 28.8 9.2 years) and both parents. A psychiatric assessment of every participant was conducted as previously described. We selected 17 tagging single nucleotide polymorphisms (SNP) for OXTR (chr3:8767079.8788481) from the HapMap database (http://hapmap.ncbi.nlm.nih.gov/). All SNP were genotyped using the TaqMan 5’-exonuclease assay. Deviations from Hardy–Weinberg equilibrium (HWE) and allelic associations were tested using Haploview v4.2 (http://www.broad institute.org/scientific-community/science/programs/medicaland-population-genetics/haploview/haploview). Fixed effect model meta-analysis was performed using catmap (http:// cran.r-project.org/src/contrib/Archive/catmap/). A power calculation was performed using the Genetic Power Calculator (http://pngu.mgh.harvard.edu/~purcell/gpc/). In the case–control sample, the genotypes of rs2139184 could not be determined, and HWE deviations were observed in rs11131149 in both patients and controls and in rs237889 in controls (Table S1). These three SNP were excluded from further analysis. There were no significant associations between any of the 14 SNP examined and schizophrenia in both the case–control and trio samples (Table S2). A metaanalysis using both samples indicated a nominally significant association between rs9840864 and schizophrenia (odds ratio, 1.21; 95% confidence interval: 1.03–1.41, P = 0.0176). The statistical power of the case–control and of the trio samples was 0.56 and 0.14, respectively, assuming a disease prevalence of 0.01, a risk allele frequency of 0.3 and a genotypic relative risk of 1.2 for heterozygous risk allele carriers under the multiplicative model of inheritance. Although rs9840864 was tentatively associated with schizophrenia, the present study did not provide supportive evidence for the contribution of OXTR to susceptibility to schizophrenia. To draw a definitive conclusion, further studies using larger sample sizes should be carried out in a range of ethnic populations.

Rare UNC13B variations and risk of schizophrenia: Whole-exome sequencing in a multiplex family and follow-up resequencing and a case-control study.

Rare genomic variations inherited in multiplex schizophrenia families are suggested to play a role in the genetic etiology of the disease. To identify rare variations with large effects on the risk of developing schizophrenia, we performed whole‐exome sequencing (WES) in two affected and one unaffected individual of a multiplex family with 10 affected individuals. We also performed follow‐up resequencing of the unc‐13 homolog B (Caenorhabditis elegans) (UNC13B) gene, a potential risk gene identified by WES, in the multiplex family and undertook a case–control study to investigate association between UNC13B and schizophrenia. UNC13B coding regions (39 exons) from 15 individuals of the multiplex family and 111 affected offspring for whom parental DNA samples were available were resequenced. Rare missense UNC13B variations identified by resequencing were further tested for association with schizophrenia in two independent case–control populations comprising a total of 1,753 patients and 1,602 controls. A rare missense variation (V1525M) in UNC13B was identified by WES in the multiplex family; this variation was present in five of six affected individuals, but not in eight unaffected individuals or one individual of unknown disease status. Resequencing UNC13B coding regions identified five rare missense variations (T103M, M813T, P1349T, I1362T, and V1525M). In the case–control study, there was no significant association between rare missense UNC13B variations and schizophrenia, although single‐variant meta‐analysis indicated that M813T was nominally associated with schizophrenia. These results do not support a contribution of rare missense UNC13B variations to the genetic etiology of schizophrenia in the Japanese population.

Rare heterozygous truncating variations and risk of autism spectrum disorder: Whole-exome sequencing of a multiplex family and follow-up study in a Japanese population

Rare heterozygous truncating variations in multiplex families with autism spectrum disorder (ASD) are suggested to play a major role in the genetic etiology of ASD. To further investigate the role of rare heterozygous truncating variations, we performed whole‐exome sequencing (WES) in a multiplex ASD family with four affected individuals (two siblings and two maternal cousins), and a follow‐up case–control study in a Japanese population.

Neural activity in the posterior superior temporal region during eye contact perception correlates with autistic traits

The present study investigated the relationship between neural activity associated with gaze processing and autistic traits in typically developed subjects using magnetoencephalography. Autistic traits in 24 typically developed college students with normal intelligence were assessed using the Autism Spectrum Quotient (AQ). The Minimum Current Estimates method was applied to estimate the cortical sources of magnetic responses to gaze stimuli. These stimuli consisted of apparent motion of the eyes, displaying direct or averted gaze motion. Results revealed gaze-related brain activations in the 150-250 ms time window in the right posterior superior temporal sulcus (pSTS), and in the 150-450 ms time window in medial prefrontal regions. In addition, the mean amplitude in the 150-250 ms time window in the right pSTS region was modulated by gaze direction, and its activity in response to direct gaze stimuli correlated with AQ score. pSTS activation in response to direct gaze is thought to be related to higher-order social processes. Thus, these results suggest that brain activity linking eye contact and social signals is associated with autistic traits in a typical population.

Novel Rare Missense Variations and Risk of Autism Spectrum Disorder: Whole-Exome Sequencing in Two Families with Affected Siblings and a Two-Stage Follow-Up Study in a Japanese Population

Rare inherited variations in multiplex families with autism spectrum disorder (ASD) are suggested to play a major role in the genetic etiology of ASD. To further investigate the role of rare inherited variations, we performed whole-exome sequencing (WES) in two families, each with three affected siblings. We also performed a two-stage follow-up case-control study in a Japanese population. WES of the six affected siblings identified six novel rare missense variations. Among these variations, CLN8 R24H was inherited in one family by three affected siblings from an affected father and thus co-segregated with ASD. In the first stage of the follow-up study, we genotyped the six novel rare missense variations identified by WES in 241 patients and 667 controls (the Niigata sample). Only CLN8 R24H had higher mutant allele frequencies in patients (1/482) compared with controls (1/1334). In the second stage, this variation was further genotyped, yet was not detected in a sample of 309 patients and 350 controls (the Nagoya sample). In the combined Niigata and Nagoya samples, there was no significant association (odds ratio = 1.8, 95% confidence interval = 0.1–29.6). These results suggest that CLN8 R24H plays a role in the genetic etiology of ASD, at least in a subset of ASD patients.

Rare genetic variants in CX3CR1 and their contribution to the increased risk of schizophrenia and autism spectrum disorders

CX3CR1, a G protein-coupled receptor solely expressed by microglia in the brain, has been repeatedly reported to be associated with neurodevelopmental disorders including schizophrenia (SCZ) and autism spectrum disorders (ASD) in transcriptomic and animal studies but not in genetic studies. To address the impacts of variants in CX3CR1 on neurodevelopmental disorders, we conducted coding exon-targeted resequencing of CX3CR1 in 370 Japanese SCZ and 192 ASD patients using next-generation sequencing technology, followed by a genetic association study in a sample comprising 7054 unrelated individuals (2653 SCZ, 574 ASD and 3827 controls). We then performed in silico three-dimensional (3D) structural modeling and in vivo disruption of Akt phosphorylation to determine the impact of the detected variant on CX3CR1-dependent signal transduction. We detected a statistically significant association between the variant Ala55Thr in CX3CR1 with SCZ and ASD phenotypes (odds ratio=8.3, P=0.020). A 3D structural model indicated that Ala55Thr could destabilize the conformation of the CX3CR1 helix 8 and affect its interaction with a heterotrimeric G protein. In vitro functional analysis showed that the CX3CR1-Ala55Thr mutation inhibited cell signaling induced by fractalkine, the ligand for CX3CR1. The combined data suggested that the variant Ala55Thr in CX3CR1 might result in the disruption of CX3CR1 signaling. Our results strengthen the association between microglia-specific genes and neurodevelopmental disorders.

Mutation screening of GRIN2B in schizophrenia and autism spectrum disorder in a Japanese population

N-methyl-d-aspartate receptors (NMDARs) play a critical role in excitatory synaptic transmission and plasticity in the central nervous systems. Recent genetics studies in schizophrenia (SCZ) show that SCZ is susceptible to NMDARs and the NMDAR signaling complex. In autism spectrum disorder (ASD), several studies report dysregulation of NMDARs as a risk factor for ASD. To further examine the association between NMDARs and SCZ/ASD development, we conducted a mutation screening study of GRIN2B which encodes NR2B subunit of NMDARs, to identify rare mutations that potentially cause diseases, in SCZ and ASD patients (n = 574 and 152, respectively). This was followed by an association study in a large sample set of SCZ, ASD, and normal healthy controls (n = 4145, 381, and 4432, respectively). We identified five rare missense mutations through the mutation screening of GRIN2B. Although no statistically significant association between any single mutation and SCZ or ASD was found, one of its variant, K1292R, is found only in the patient group. To further examine the association between mutations in GRIN2B and SCZ/ASD development, a larger sample size and functional experiments are needed.

Rare truncating variations and risk of schizophrenia: Whole-exome sequencing in three families with affected siblings and a three-stage follow-up study in a Japanese population

Rare inherited variations in multiplex families with schizophrenia are suggested to play a role in the genetic etiology of schizophrenia. To further investigate the role of rare inherited variations, we performed whole-exome sequencing (WES) in three families, each with two affected siblings. We also performed a three-stage follow-up case-control study in a Japanese population with a total of 2617 patients and 2396 controls. WES identified 15 rare truncating variations that were variously present in the two affected siblings in each family. These variations did not necessarily segregate with schizophrenia within families, and they were different in each family. In the follow-up study, four variations (NWD1 W169X, LCORL R7fsX53, CAMK2B L497fsX497, and C9orf89 Q102X) had a higher mutant allele frequency in patients compared with controls, although these associations were not significant in the combined population, which comprised the first-, second- and third-stage populations. These results do not support a contribution of the rare truncating variations identified in the three families to the genetic etiology of schizophrenia.