De novo GABRG2mutations associated with epileptic encephalopathies

Abstract

Epileptic encephalopathies are a devastating group of severe childhood onset epilepsies with medication-resistant seizures and poor developmental outcomes. Many epileptic encephalopathies have a genetic aetiology and are often associated with de novo mutations in genes mediating synaptic transmission, including GABA receptor subunit genes. Recently, we performed next generation sequencing on patients with a spectrum of epileptic encephalopathy phenotypes, and we identified five novel (A106T, I107T, P282S, R323W and F343L) and one known (R323Q) de novo GABRG2 pathogenic variants (mutations) in eight patients. To gain insight into the molecular basis for how these mutations contribute to epileptic encephalopathies, we compared the effects of the mutations on the properties of recombinant α1β2γ2L GABA receptors transiently expressed in HEK293T cells. Using a combination of patch clamp recording, immunoblotting, confocal imaging and structural modelling, we characterized the effects of these GABRG2 mutations on GABA receptor biogenesis and channel function. Compared with wild-type α1β2γ2L receptors, GABA receptors containing a mutant γ2 subunit had reduced cell surface expression with altered subunit stoichiometry or decreased GABA-evoked whole-cell current amplitudes, but with different levels of reduction. While a causal role of these mutations cannot be established directly from these results, the functional analysis together with the genetic information suggests that these GABRG2 variants may be major contributors to the epileptic encephalopathy phenotypes. Our study further expands the GABRG2 phenotypic spectrum and supports growing evidence that defects in GABAergic neurotransmission participate in the pathogenesis of genetic epilepsies including epileptic encephalopathies.

Critical roles of αII spectrin in brain development and epileptic encephalopathy

“…Gephyrin is required for the organization, stability, and clustering of GABA A Rs (37,38). Genetic mutations in both GABA A Rs and gephyrin have been identified in different epilepsy syndromes, including severe epileptic encephalopathies (39)(40)(41)(42)(43)(44). Our data suggest that defective gephyrin clustering and GABAergic innervation of cortical pyramidal neurons contribute to the pathogenesis of EIEE5, providing further evidence that inhibitory transmission plays critical roles in EIEEs.…”

Expanding the phenotypic spectrum of GABRG2 variants: a recurrent GABRG2 missense variant associated with a severe phenotype

“…Furthermore, variable movement disorder, dysmorphic features, and visual impairment and ocular issues have also been seen in most of these patients (Table 1). While these features are highly variable, they have not been recognized in other GABRG2 -associated cases published prior to 2016, but are consistent with a recent report of five pathogenic/likely pathogenic missense variants in GABRG2 (Shen et al, 2017). …”

“…A recent in vitro functional study from Shen et.al demonstrated that not only did the p.A106T variant decrease macroscopic GABA-evoked currents in p.A106T transfected HEK293T cells by 30%, but it also reduced surface levels of mutant γ2L subunits by 26% compared to co-expressed wild-type γ2L subunits. These results all suggest that the p.A106T variant reduces biogenesis of GABA type A receptors (Shen et al, 2017). …”

“…To date, pathogenic variants in the GABRG2 gene, including 11 truncating variants (four nonsense, four frame-shifts, two splice-sites, and one large deletion) and 16 missense variants, have been reported in a subset of families and individuals with a variety of phenotypes ranging from epileptic encephalopathies (including Dravet syndrome) to genetic (generalized) epilepsy with febrile seizures plus (GEFS+), to febrile seizures associated with childhood absence epilepsy (CAE) and milder simple febrile seizures (FS) (Baulac et al, 2001; Wallace et al, 2001; Harkin et al, 2002; Kananura et al, 2002; Audenaert et al, 2006; Hirose, 2006; Sun et al, 2008; Macdonald et al, 2010; Shi et al, 2010; Cantarín-Extremera et al, 2011; Lachance-Touchette et al, 2011; Balan et al, 2013; Carvill et al, 2013; Tian et al, 2013; Johnston et al, 2014; Boillot et al, 2015; Reinthaler et al, 2015; Shen et al, 2017). In these studies, the majority of pathogenic variants in GABRG2 segregated predominantly with a FS phenotype.…”

“…Some individuals with GABRG2 pathogenic variants may never develop seizures, indicating incomplete penetrance (Wallace et al, 2001; Hirose, 2006). Only recently, a case series revealed evidence of GABRG2 missense variants being responsible for a more severe epileptic encephalopathy phenotype (Shen et al, 2017). Here we describe a recurrent pathogenic missense variant (c.316 G>A; p.A106T) in the GABRG2 gene in five unrelated patients with severe phenotypes which include significant motor and speech delays, intellectual disability, hypotonia, movement disorder, dysmorphic features, and visual impairment and other ocular issues, in addition to variable early-onset seizures.…”