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

<div><p>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, <i>CLN8</i> 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 <i>CLN8</i> 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 <i>CLN8</i> R24H plays a role in the genetic etiology of ASD, at least in a subset of ASD patients.</p></div

Genomic structure of the ceroid-lipofuscinosis, neuronal 8 (epilepsy, progressive with mental retardation) (<i>CLN8</i>) gene.

<p><i>CLN8</i> spans approximately 22.9 kb and has three exons (rectangles). Black and white rectangles represent coding and untranslated regions, respectively. A horizontal arrow shows the orientation of transcription. Vertical arrows indicate locations of rare non-synonymous variations identified by resequencing.</p

Pedigrees of two families, each with three autism spectrum disorder siblings.

<p>(A) Family #1. All three siblings (II-1, II-2, and II-3) were diagnosed with Asperger’s disorder. (B) Family #2. There were four affected individuals: a proband (II-1) with Asperger’s disorder, his brother (II-2) with Asperger’s disorder, his brother (II-3) with Asperger’s disorder and borderline intellectual functioning, and their father (I-1) with pervasive developmental disorder not otherwise specified. Shaded and unshaded symbols indicate affected and unaffected individuals, respectively. Squares and circles represent males and females, respectively.</p

Clinical phenotypes of the six individuals heterozygous for <i>CLN8</i> R24H.

<p>IQ, intelligence quotient; PDD-NOS, pervasive developmental disorder not otherwise specified.</p><p>Clinical phenotypes of the six individuals heterozygous for <i>CLN8</i> R24H.</p

WES quality report summary.

<p>WES, whole-exome sequencing.</p><p>WES quality report summary.</p

Six novel rare missense variations identified by WES in two families, each with three ASD siblings.

<p>WES, whole-exome sequencing; ASD, autism spectrum disorder; GERP, Genomic Evolutionary Rate Profiling.</p><p>^aPosition according to GRCh37.</p><p>^bReference/mutant allele.</p><p>Six novel rare missense variations identified by WES in two families, each with three ASD siblings.</p

Genotyping of the six novel rare missense variations in the follow-up study.

<p>ASD, autism spectrum disorder; CI, confidence interval; MAF, mutant allele frequency; OR, odds ratio.</p><p>^aGenotypes: reference and mutant alleles are denoted by 1 and 2, respectively.</p><p>Genotyping of the six novel rare missense variations in the follow-up study.</p