De novo mutations in synaptic transmission genes including DNM1 cause epileptic encephalopathies

Abstract

Emerging evidence indicates that epileptic encephalopathies are genetically highly heterogeneous, underscoring the need for large cohorts of well-characterized individuals to further define the genetic landscape. Through a collaboration between two consortia (EuroEPINOMICS and Epi4K/EPGP), we analyzed exome-sequencing data of 356 trios with the "classical" epileptic encephalopathies, infantile spasms and Lennox Gastaut syndrome, including 264 trios previously analyzed by the Epi4K/EPGP consortium. In this expanded cohort, we find 429 de novo mutations, including de novo mutations in DNM1 in five individuals and de novo mutations in GABBR2, FASN, and RYR3 in two individuals each. Unlike previous studies, this cohort is sufficiently large to show a significant excess of de novo mutations in epileptic encephalopathy probands compared to the general population using a likelihood analysis (p = 8.2 × 10(-4)), supporting a prominent role for de novo mutations in epileptic encephalopathies. We bring statistical evidence that mutations in DNM1 cause epileptic encephalopathy, find suggestive evidence for a role of three additional genes, and show that at least 12% of analyzed individuals have an identifiable causal de novo mutation. Strikingly, 75% of mutations in these probands are predicted to disrupt a protein involved in regulating synaptic transmission, and there is a significant enrichment of de novo mutations in genes in this pathway in the entire cohort as well. These findings emphasize an important role for synaptic dysregulation in epileptic encephalopathies, above and beyond that caused by ion channel dysfunction.

Figure 1

Hot-Zone Analysis Illustrates where de novo single-nucleotide mutations reside along the variant-level vector (x axis: PolyPhen-2 HumVar score) and the gene-level vector (y axis: RVIS percentile score). We identified 411 controls (A, adapted from Zhu et al.³⁴) and 232 epileptic encephalopathy cases (B) that reported at least one single-nucleotide de novo mutation. For each of the cases and controls, we plot the single most damaging single-nucleotide de novo mutation, as defined by the shortest Euclidian distance from the most damaging coordinate [1,0]. The hot zone (red shading) is defined as the region that reflects a PolyPhen-2 score ≥0.95 and a RVIS percentile score ≤25%. Details for all hot-zone de novo mutations are available in Table S1. Among the case hot-zone insert we flag with red the hot-zone de novo mutations that occur in the epileptic encephalopathy gene list as described in the panel.

Figure 2

Primary Protein-Protein Interaction Network of 139 Interconnected Proteins This reflects 134 proteins that are affected by de novo substitutions identified among the 356 epileptic encephalopathy probands reported here and 5 literature-introduced epileptic encephalopathy proteins (marked with an asterisk). Ingenuity Pathway Analysis (IPA) annotated “synaptic junction transmission” proteins are marked in yellow. Known epileptic encephalopathy genes, including the newly identified DNM1, are circled in gray. The geometric shapes reflect differing protein roles, as defined by IPA: enzyme, rhombus; ion channel, vertical rectangle; kinase, inverted triangle; ligand-dependent nuclear receptor, horizontal rectangle; phosphatase, triangle; transcription regulator, horizontal oval; transmembrane receptor, vertical oval; transporter, trapezoid; and unknown, circle.

Figure 3

Schematic Representation of DNM1 with Location of Substitutions and Conservation of Substitution Sites (A) Structure of the DNM1 protein with indication of the different domains and the G1–G4 motifs. Substitutions identified in this study are shown below the figure, the substitution in the fitful mouse above the figure. PH, pleckstrin homology; GED, GTPase effector domain; PRD, proline-rich domain. (B) Sequence alignment in different species of the regions of the substitutions found in this study. Substitutions are highlighted in red.