Genomic analysis of autism spectrum disorder and identification of a novel disease gene NEO1

Abstract

Being a highly heritable neurodevelopmental disease, autism spectrum disorder (ASD) affects a staggering number of individuals globally with considerable impairments in social interactions and communications. With the immense clinical and genetic heterogeneity, elucidating the underlying pathogenesis of ASD is deemed to be exceedingly challenging. The advances in high throughput next-generation sequencing (NGS) and the increasing utilisation of chromosomal microarray in the recent decade result in tremendous developments in the understanding on the genetic basis of ASD. In this study, these two technologies were applied to dissect the intriguing genetic aetiology of ASD. A cohort of 68 Chinese patients from Hong Kong was recruited consisting both adult and paediatric subjects. Array comparative genomic hybridization (CGH) was performed for the cohort identifying eight clinically significant copy number variants (CNV). The overall diagnostic yield of array CGH was 11.8%. Microduplication at chromosome 16p13.11 was most frequently detected with four positive cases contributing to 5.9% of the cohort. Among the clinically significant CNV, microdeletion at chromosome 15q24 was detected in one subject and this is a recurrent genomic aberration reported in individuals with autism spectrum disorder. To decipher the culprit of ASD in this region, a targeted PCR-based enrichment method was developed using next-generation sequencing to decode all the genes in 15q24 region that listed in the Online Mendelian Inheritance in Man database. Of note, a missense variant and an intronic duplication were identified in NEO1 gene that encodes for neogenin with critical role in interneuron migration and exon guidance, the key pathways in the pathogenesis of ASD. Essentially, the missense variant c.3388C>T (p.Arg1130Cys) impaired the nuclear translocation of neogenin whereas the intronic duplication c.2204-14_2204-2dup predicted to create a cryptic lariat branch point lowering the splicing efficiency. Seemingly, these results provided the first evidence establishing the link between NEO1 and a human phenotype of autism. Apart from the identification of novel disease genes, several methods were developed to study the diverse aetiology of ASD. Notably, the link between mitochondrial disorders and autistic features is widely recognised. Using next-generation sequencing, a whole mitochondrial genome sequencing method using long-range PCR as enrichment was established. The method was validated in DNA samples extracted from blood and urine, which was deemed to be a robust and comprehensive approach to identify mitochondrial mutations. Furthermore, a targeted gene panel consisting key genes that evidently linked to autism were developed using NGS. The genes in the panel include MECP2, PTEN, NLGN3, NRXN1, CNTNAP2 and SHANK3. The method allowed simultaneous analysis of multiple ASD genes in a single analytical run. By and large, the implications of the findings from the present study can be enormous. The CNV spectrum of ASD in the Hong Kong Chinese population was delineated. NEO1 was identified to be the novel disease gene of ASD. These results certainly enhanced the understanding of the genetic basis underpinning the pathogenesis of ASD and promoted the prospect of innovative diagnostics and therapeutics.