title: Whole-exome sequencing and neurite outgrowth analysis in autism spectrum disorder.
abstract: Autism spectrum disorder (ASD) is a complex group of clinically heterogeneous neurodevelopmental disorders with unclear etiology and pathogenesis. Genetic studies have identified numerous candidate genetic variants, including de novo mutated ASD-associated genes; however, the function of these de novo mutated genes remains unclear despite extensive bioinformatics resources. Accordingly, it is not easy to assign priorities to numerous candidate ASD-associated genes for further biological analysis. Here we developed a convenient system for identifying an experimental evidence-based annotation of candidate ASD-associated genes. We performed trio-based whole-exome sequencing in 30 sporadic cases of ASD and identified 37 genes with de novo single-nucleotide variations (SNVs). Among them, 5 of those 37 genes, POGZ, PLEKHA4, PCNX, PRKD2 and HERC1, have been previously reported as genes with de novo SNVs in ASD; and consultation with in silico databases showed that only HERC1 might be involved in neural function. To examine whether the identified gene products are involved in neural functions, we performed small hairpin RNA-based assays using neuroblastoma cell lines to assess neurite development. Knockdown of 8 out of the 14 examined genes significantly decreased neurite development (P<0.05, one-way analysis of variance), which was significantly higher than the number expected from gene ontology databases (P=0.010, Fisher's exact test). Our screening system may be valuable for identifying the neural functions of candidate ASD-associated genes for further analysis and a substantial portion of these genes with de novo SNVs might have roles in neuronal systems, although further detailed analysis might eliminate false positive genes from identified candidate ASD genes.
authors.affiliation: Molecular Research Center for Children's Mental Development, United Graduate School of Child Development, Osaka University, Osaka, Japan.|Department of Psychiatry, Osaka University Graduate School of Medicine, Osaka, Japan.
authors.forename: Ryota
authors.identifier: http://orcid.org/0000-0002-5941-4238
authors.initials: R
authors.lastname: Hashimoto
authors.affiliation: iPS Cell-Based Research Project on Brain Neuropharmacology and Toxicology, Graduate School of Pharmaceutical Sciences, Osaka University, Osaka, Japan.
authors.forename: Takanobu
authors.initials: T
authors.lastname: Nakazawa
authors.affiliation: Department of Human Genetics, Yokohama City University Graduate School of Medicine, Yokohama, Japan.
authors.forename: Yoshinori
authors.initials: Y
authors.lastname: Tsurusaki
authors.affiliation: Department of Psychiatry, Osaka University Graduate School of Medicine, Osaka, Japan.
authors.forename: Yuka
authors.initials: Y
authors.lastname: Yasuda
authors.affiliation: iPS Cell-Based Research Project on Brain Neuropharmacology and Toxicology, Graduate School of Pharmaceutical Sciences, Osaka University, Osaka, Japan.
authors.forename: Kazuki
authors.initials: K
authors.lastname: Nagayasu
authors.affiliation: Laboratory of Molecular Neuropharmacology, Graduate School of Pharmaceutical Sciences, Osaka University, Osaka, Japan.
authors.forename: Kensuke
authors.initials: K
authors.lastname: Matsumura
authors.affiliation: Genomic Science Laboratories, Sumitomo Dainippon Pharma Co., Ltd., Osaka, Japan.
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Kawashima
authors.affiliation: Department of Psychiatry, Osaka University Graduate School of Medicine, Osaka, Japan.
authors.forename: Hidenaga
authors.initials: H
authors.lastname: Yamamori
authors.affiliation: Department of Psychiatry, Osaka University Graduate School of Medicine, Osaka, Japan.
authors.forename: Michiko
authors.initials: M
authors.lastname: Fujimoto
authors.affiliation: Department of Psychiatry, Osaka University Graduate School of Medicine, Osaka, Japan.
authors.forename: Kazutaka
authors.initials: K
authors.lastname: Ohi
authors.affiliation: Department of Molecular Neuropsychiatry, Osaka University Graduate School of Medicine, Osaka, Japan.
authors.forename: Satomi
authors.initials: S
authors.lastname: Umeda-Yano
authors.affiliation: Division of Cerebral Integration, National Institute of Physiological Sciences, Aichi, Japan.
authors.forename: Masaki
authors.initials: M
authors.lastname: Fukunaga
authors.affiliation: Graduate School of Human Sciences, Osaka University, Osaka, Japan.
authors.forename: Haruo
authors.initials: H
authors.lastname: Fujino
authors.affiliation: Laboratory of Molecular Neuropharmacology, Graduate School of Pharmaceutical Sciences, Osaka University, Osaka, Japan.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Kasai
authors.affiliation: Molecular Research Center for Children's Mental Development, United Graduate School of Child Development, Osaka University, Osaka, Japan.|Laboratory of Molecular Neuropharmacology, Graduate School of Pharmaceutical Sciences, Osaka University, Osaka, Japan.
authors.forename: Atsuko
authors.initials: A
authors.lastname: Hayata-Takano
authors.affiliation: Laboratory of Molecular Neuropharmacology, Graduate School of Pharmaceutical Sciences, Osaka University, Osaka, Japan.
authors.forename: Norihito
authors.identifier: http://orcid.org/0000-0002-7850-9064
authors.initials: N
authors.lastname: Shintani
authors.affiliation: Molecular Research Center for Children's Mental Development, United Graduate School of Child Development, Osaka University, Osaka, Japan.|Department of Psychiatry, Osaka University Graduate School of Medicine, Osaka, Japan.
authors.forename: Masatoshi
authors.initials: M
authors.lastname: Takeda
authors.affiliation: Department of Human Genetics, Yokohama City University Graduate School of Medicine, Yokohama, Japan.
authors.forename: Naomichi
authors.initials: N
authors.lastname: Matsumoto
authors.affiliation: Molecular Research Center for Children's Mental Development, United Graduate School of Child Development, Osaka University, Osaka, Japan.|iPS Cell-Based Research Project on Brain Neuropharmacology and Toxicology, Graduate School of Pharmaceutical Sciences, Osaka University, Osaka, Japan.|Laboratory of Molecular Neuropharmacology, Graduate School of Pharmaceutical Sciences, Osaka University, Osaka, Japan.
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Hashimoto
mesh_terms: D000328:Adult; D000818:Animals; D000067877:Autism Spectrum Disorder / Q000235:genetics*; D002460:Cell Line; D059472:Exome; D005260:Female; D020022:Genetic Predisposition to Disease; D006801:Humans; D008297:Male; D051379:Mice; D008875:Middle Aged; D016501:Neurites; D017421:Sequence Analysis; D055815:Young Adult