Multiple studies have confirmed the contribution of rare de novo copy number variations (CNVs) to the risk for Autism Spectrum Disorders (ASD).1-3 While de novo single nucleotide variants (SNVs) have been identified in affected individuals,4 their contribution to risk has yet to be clarified. Specifically, the frequency and distribution of these mutations has not been well characterized in matched unaffected controls, data that are vital to the interpretation of de novo coding mutations observed in probands. Here we show, via whole-exome sequencing of 928 individuals, including 200 phenotypically discordant sibling pairs, that highly disruptive (nonsense and splice-site) de novo mutations in brain-expressed genes are associated with ASD and carry large effects (OR=5.65; CI: 1.44-22.2; p=0.01 asymptotic test). Based on mutation rates in unaffected individuals, we demonstrate that multiple independent de novo SNVs in the same gene among unrelated probands reliably identifies risk alleles, providing a clear path forward for gene discovery. Among a total of 279 identified de novo coding mutations, there is a single instance in probands, and none in siblings, in which two independent nonsense variants disrupt the same gene, SCN2A (Sodium Channel, Voltage-Gated, Type II, Alpha Subunit), a result that is highly unlikely by chance (p=0.005).

Summary Given prior evidence for the contribution of rare copy number variations (CNVs) to autism spectrum disorders (ASD), we studied these events in 4,457 individuals from 1,174 simplex families, composed of parents, a proband and, in most kindreds, an unaffected sibling. We find significant association of ASD with de novo duplications of 7q11.23, where the reciprocal deletion causes Williams-Beuren syndrome, featuring a highly social personality. We identify rare recurrent de novo CNVs at five additional regions including two novel ASD loci, 16p13.2 (including the genes USP7 and C16orf72) and Cadherin13, and implement a rigorous new approach to evaluating the statistical significance of these observations. Overall, we find large de novo CNVs carry substantial risk (OR=3.55; CI =2.16-7.46, p=6.9 × 10−6); estimate the presence of 130-234 distinct ASD-related CNV intervals across the genome; and, based on data from multiple studies, present compelling evidence for the association of rare de novo events at 7q11.23, 15q11.2-13.1, 16p11.2, and Neurexin1.

An increasing number of genetic variants have been implicated in autism spectrum disorders (ASD), and the functional study of such variants will be critical for the elucidation of autism pathophysiology. Here, we report a de novo balanced translocation disruption of TRPC6, a cation channel, in a non-syndromic autistic individual. Using multiple models, such as dental pulp cells, iPSC-derived neuronal cells and mouse models, we demonstrate that TRPC6 reduction or haploinsufficiency leads to altered neuronal development, morphology, and function. The observed neuronal phenotypes could then be rescued by TRPC6 complementation and by treatment with IGF1 or hyperforin, a TRPC6-specific agonist, suggesting that ASD individuals with alterations in this pathway might benefit from these drugs. We also demonstrate that MeCP2 levels affect TRPC6 expression. Mutations in MeCP2 cause Rett syndrome, revealing common pathways among ASDs. Genetic sequencing of TRPC6 in 1041 ASD individuals and 2872 controls revealed significantly more nonsynonymous mutations in the ASD population, and identified loss-of-function mutations with incomplete penetrance in two patients. Taken together, these findings suggest that TRPC6 is a novel predisposing gene for ASD that may act in a multiple-hit model. This is the first study to use iPSC-derived human neurons to model non-syndromic ASD and illustrate the potential of modeling genetically complex sporadic diseases using such cells.

SummaryEndophilin-A, a well-characterized endocytic adaptor essential for synaptic vesicle recycling, has recently been linked to neurodegeneration. We report here that endophilin-A deficiency results in impaired movement, age-dependent ataxia, and neurodegeneration in mice. Transcriptional analysis of endophilin-A mutant mice, complemented by proteomics, highlighted ataxia- and protein-homeostasis-related genes and revealed upregulation of the E3-ubiquitin ligase FBXO32/atrogin-1 and its transcription factor FOXO3A. FBXO32 overexpression triggers apoptosis in cultured cells and neurons but, remarkably, coexpression of endophilin-A rescues it. FBXO32 interacts with all three endophilin-A proteins. Similarly to endophilin-A, FBXO32 tubulates membranes and localizes on clathrin-coated structures. Additionally, FBXO32 and endophilin-A are necessary for autophagosome formation, and both colocalize transiently with autophagosomes. Our results point to a role for endophilin-A proteins in autophagy and protein degradation, processes that are impaired in their absence, potentially contributing to neurodegeneration and ataxia.