REPORT De Novo Pathogenic SCN8A Mutation Identified by Whole-Genome Sequencing of a Family Quartet Affected by Infantile Epileptic Encephalopathy and SUDEP Krishna R. Veeramah, 1 Janelle E. O’Brien, 4 Miriam H. Meisler, 4 Xiaoyang Cheng, 5 Sulayman D. Dib-Hajj, 5 Stephen G. Waxman, 5 Dinesh Talwar, 6,7,9 Santhosh Girirajan, 10 Evan E. Eichler, 10 Linda L. Restifo, 2,7,8 Robert P. Erickson, 3,6 and Michael F. Hammer 1, * Individuals with severe, sporadic disorders of infantile onset represent an important class of disease for which discovery of the under- lying genetic architecture is not amenable to traditional genetic analysis. Full-genome sequencing of affected individuals and their parents provides a powerful alternative strategy for gene discovery. We performed whole-genome sequencing (WGS) on a family quartet containing an affected proband and her unaffected parents and sibling. The 15-year-old female proband had a severe epileptic enceph- alopathy consisting of early-onset seizures, features of autism, intellectual disability, ataxia, and sudden unexplained death in epilepsy. We discovered a de novo heterozygous missense mutation (c.5302A>G [p.Asn1768Asp]) in the voltage-gated sodium-channel gene SCN8A in the proband. This mutation alters an evolutionarily conserved residue in Nav1.6, one of the most abundant sodium channels in the brain. Analysis of the biophysical properties of the mutant channel demonstrated a dramatic increase in persistent sodium current, incomplete channel inactivation, and a depolarizing shift in the voltage dependence of steady-state fast inactivation. Current-clamp analysis in hippocampal neurons transfected with p.Asn1768Asp channels revealed increased spontaneous firing, parox- ysmal-depolarizing-shift-like complexes, and an increased firing frequency, consistent with a dominant gain-of-function phenotype in the heterozygous proband. This work identifies SCN8A as the fifth sodium-channel gene to be mutated in epilepsy and demonstrates the value of WGS for the identification of pathogenic mutations causing severe, sporadic neurological disorders. Massively-parallel-sequencing technologies are revolu- tionizing the process of discovering genetic variants that cause disease. 1 Neurodevelopmental disorders such as epilepsy, autism spectrum disorders (ASDs), intellectual disability (ID), and schizophrenia represent a considerable challenge for molecular genetic analysis because of marked genetic heterogeneity, environmental effects on the severity of symptoms, and the frequent co-occurrence of seizures, autism, and cognitive phenotypes. Studies of copy-number variation (CNV) have demonstrated the contribution of de novo variants in these disorders. 2,3 However, CNVs only appear to contribute to between 10% and 25% of affected cases. 4 It is hypothesized that rare or novel point mutations might contribute to many of the remaining cases under the CD/MRV (common disease/multiple rare variant) model. 5 When the observed phenotype is particularly severe and there is no prior family history of the disorder, it is reasonable to consider a disease model that involves a dominant de novo muta- tion. Support for this model comes from studies of epileptic encephalopathies, in which de novo mutations of the sodium-channel gene SCN1A (MIM 182389) are a major cause of Dravet Syndrome (MIM 607208), 6 whereas de novo mutations in STXBP1 (MIM 602926) and ARX (MIM 300382) have been found in a number of individuals with early infantile epileptic encephalopathy (MIM 308350). 7 When such mutations arise, they are expected to be quickly removed by strong purifying selection (because affected individuals rarely reproduce) and hence would be extremely rare or unique in the population. Although the human mutation rate is on the order of 1 3 10 -8 to 2 3 10 8 per site per generation, 8,9 thousands of genes are potentially involved in neurodevelopment, 10 suggesting that the number of de novo pathogenic muta- tions could be substantial. Thus, although each individual is expected to have only ~1 de novo mutation per exome, 11 a model of rare mutations across many genes might explain why severe neurological disorders are rela- tively common. 12 Whole-exome sequencing of parent-offspring trios offers a cost-effective method for screening coding regions for mutations and has been successful in identifying candi- date de novo variants in sporadic cases of ID, 12 ASDs, 13 and schizophrenia. 14 However, the limitations of current exome capture and sequencing methodologies include incomplete or variable coverage of exons and the inability to infer ploidy across the genome or survey regulatory vari- ation. Whole-genome sequencing (WGS) studies are not limited by these aspects, and when they are implemented in a quartet framework, they have many attractive analyt- ical advantages. For example, it is possible to precisely infer haplotype phase and the location of recombination 1 Arizona Research Laboratories Division of Biotechnology, 2 Department of Neuroscience, 3 Department of Molecular and Cellular Biology, University of Arizona, Tucson, AZ, 85721, USA; 4 Department of Human Genetics, University of Michigan, Ann Arbor, MI 48109-5618, USA; 5 Department of Neurology, Yale School of Medicine, New Haven, CT 06520-8018, USA; 6 Department of Pediatrics, 7 Department of Neurology, 8 Department of Cellular and Molecular Medicine, Arizona Health Science Center, Tucson, AZ 85724, USA; 9 Center for Neurosciences, Tucson, AZ 85718, USA; 10 Department of Genome Sciences, Howard Hughes Medical Institute, University of Washington, Seattle, WA 98195, USA *Correspondence: mfh@email.arizona.edu DOI 10.1016/j.ajhg.2012.01.006. Ó2012 by The American Society of Human Genetics. All rights reserved. 502 The American Journal of Human Genetics 90, 502–510, March 9, 2012