Autism genome-wide copy number variation reveals ubiquitin and neuronal genes

Abstract

Autism spectrum disorders (ASDs) are childhood neurodevelopmental disorders with complex genetic origins. Previous studies focusing on candidate genes or genomic regions have identified several copy number variations (CNVs) that are associated with an increased risk of ASDs. Here we present the results from a whole-genome CNV study on a cohort of 859 ASD cases and 1,409 healthy children of European ancestry who were genotyped with approximately 550,000 single nucleotide polymorphism markers, in an attempt to comprehensively identify CNVs conferring susceptibility to ASDs. Positive findings were evaluated in an independent cohort of 1,336 ASD cases and 1,110 controls of European ancestry. Besides previously reported ASD candidate genes, such as NRXN1 (ref. 10) and CNTN4 (refs 11, 12), several new susceptibility genes encoding neuronal cell-adhesion molecules, including NLGN1 and ASTN2, were enriched with CNVs in ASD cases compared to controls (P = 9.5 x 10(-3)). Furthermore, CNVs within or surrounding genes involved in the ubiquitin pathways, including UBE3A, PARK2, RFWD2 and FBXO40, were affected by CNVs not observed in controls (P = 3.3 x 10(-3)). We also identified duplications 55 kilobases upstream of complementary DNA AK123120 (P = 3.6 x 10(-6)). Although these variants may be individually rare, they target genes involved in neuronal cell-adhesion or ubiquitin degradation, indicating that these two important gene networks expressed within the central nervous system may contribute to the genetic susceptibility of ASD.

Figure 1. AK123120: example of overrepresented CNVs

AK123120 chromosome 2 (chr2): 12,986,750–13,291,000 divided into subsections with headers for ACC CNVs, AGRE CNVs, AGRE Affymetrix validation CNVs, and control CNVs. The AGRE Affymetrix Replication track is on the basis of genome-wide 5.0 SNP genotyping data from the Broad Institute (see Supplementary Methods and Acknowledgements), and were generated using the PennCNV-Affy algorithm (see Supplementary Methods), to serve as a further means to validate the Illumina-based CNV calls. SNP and copy number (CN) probe coverage are shown as blue lines across the top. Produced with custom tracks listing CNV calls uploaded to http://genome.ucsc.edu. Figures for all loci are included in Supplementary Information.

Figure 2. Independent validation using qPCR and MLPA

Fluorescent probe-based qPCR assays using Roche Universal probe library and/or MLPA were designed to validate every candidate CNV with a completely independent test (representative series shown for each locus). Error bars denote the s.d. of quadruplicate runs. bp, base pairs; Del, deletion; Dup, duplication.