Abstracts

Rationale: Over 700 mutations have been identified in the voltage-gated sodium channel genes SCN1A and SCN2A in human epilepsies, including Genetic Epilepsy with Febrile Seizures Plus (GEFS+), Dravet Syndrome (DS) and Benign Familial Neonatal-Infantile Seizures (BFNIS). Family members with the same mutation frequently display variation in the clinical severity of the disease, suggesting that other factors modify the primary mutation. Several mouse models have been generated to study the genetic basis of epilepsy. A common feature of these epilepsy models is that genetic strain background alters the disease phenotype, suggestive of genetic modifiers in epilepsy. The Scn2aQ54 transgenic mouse model has an epilepsy phenotype that varies depending on the genetic strain background. Scn2aQ54 mice congenic on the C57Bl/6J (B6) strain exhibit delayed seizure onset and improved survival compared to (B6xSJL/J)F1.Q54 mice. Two dominant modifier loci of Scn2aQ54 seizure susceptibility were mapped and designated Moe1 and Moe2. We performed fine mapping and candidate analysis to identify genes that alter seizure susceptibility at Moe1.Methods: Interval specific congenic (ISC) lines were utilized to fine map Moe1. ISC lines were crossed to B6.Q54 to generate F1.Q54 offspring that were heterozygous or homozygous for B6 alleles in Moe1. Spontaneous seizure frequency was measured during 30 minute observations at several ages. To refine Moe1, seizure frequency was compared between mice carrying differing segments of the Moe1 interval. RNA-Seq transcriptome analysis was utilized to evaluate differences in transcript expression and coding sequence between the B6 and SJL strains. Whole brain total RNA was isolated from 6 week old male and female B6 and SJL mice (n=4 per group). Samples were sequenced on the Illumina HiSeq 2000 platform, according to standard protocols. RNA-Seq data was analyzed in collaboration with the Vanderbilt Computational Genomics Core using Tuxedo Tools.Results: Fine mapping suggests that the Moe1 interval contains at least two modifier loci. In the proximal Moe1 interval, B6 homozygosity is protective, while the SJL allele increases susceptibility. Conversely, in the distal interval B6 homozygosity is permissive, whereas the SJL allele is protective. RNA-Seq analysis identified several candidate genes for seizure susceptibility, including top candidates, Cacna1g and NeuroD2. Future transgenic transfer experiments will test the modifier potential of Cacna1g and NeuroD2. Transgenic animals carrying Cacna1g or NeuroD2 BACs will be crossed to Scn2aQ54 mice to assess the effect on phenotype severity. Altered phenotype severity is indicative that the candidate is an epilepsy modifier gene.Conclusions: These experiments evaluate potential modifier genes that may influence the clinical course of epilepsy caused by a sodium channel mutation. In addition to ion channels, potential modifiers include genes involved in axonal transport and neural development. Identification of epilepsy modifier genes may suggest new targets for improved treatment of epilepsy.