Abstract
Synaptic inhibition is essential for shaping the dynamics of neuronal networks, and aberrant inhibition plays an important role in neurological disorders. Gephyrin is a central player at inhibitory postsynapses, directly binds and organizes GABAA and glycine receptors (GABAARs and GlyRs), and is thereby indispensable for normal inhibitory neurotransmission. Additionally, gephyrin catalyzes the synthesis of the molybdenum cofactor (MoCo) in peripheral tissue. We identified a de novo missense mutation (G375D) in the gephyrin gene (GPHN) in a patient with epileptic encephalopathy resembling Dravet syndrome. Although stably expressed and correctly folded, gephyrin-G375D was non-synaptically localized in neurons and acted dominant-negatively on the clustering of wild-type gephyrin leading to a marked decrease in GABAAR surface expression and GABAergic signaling. We identified a decreased binding affinity between gephyrin-G375D and the receptors, suggesting that Gly375 is essential for gephyrin-receptor complex formation. Surprisingly, gephyrin-G375D was also unable to synthesize MoCo and activate MoCo-dependent enzymes. Thus, we describe a missense mutation that affects both functions of gephyrin and suggest that the identified defect at GABAergic synapses is the mechanism underlying the patient's severe phenotype.
Highlights
With a burden of 68 million people affected worldwide, epilepsy is one of the most common neurological disorders (Ngugi et al, 2010)
The screening of follow-up cohorts using the Multiplex Amplification of Specific Targets for Resequencing (MASTR) and the molecular inversion probes (MIP) technology did not lead to the identification of additional cases with mutations in GPHN
The decreased stoichiometry of 6His-G375D/GlyR b-loop residues 378–426 (GlyR-bL) interaction suggests a much lower occupancy for gephyrin-G375D compared to WT-gephyrin (Fig 6E). These results suggest that Gly375 is essential for gephyrin–receptor interaction and the replacement of Gly375 to aspartate severely impairs the interaction between gephyrin and GABAARs/GlyRs
Summary
With a burden of 68 million people affected worldwide, epilepsy is one of the most common neurological disorders (Ngugi et al, 2010). The epilepsy is refractory and developmental delay occurs soon after seizure onset (Dravet, 2011). Besides structural and metabolic defects, EEs can be caused by genetic alterations. Most of these EEs present as monogenic disorders due to de novo mutations (EuroEPINOMICS-RES Consortium et al, 2014; Allen et al, 2013; Veeramah et al, 2013). About 80% of patients with Dravet syndrome, for example, carry a de novo mutation in the gene SCN1A (Parihar & Ganesh, 2013). We performed whole exome sequencing (WES) on a patient with a Dravet-like syndrome (not carrying an SCN1A mutation) and his parents and identified a heterozygous de novo missense mutation in the gephyrin gene (GPHN)
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