GABAA Receptor Coupling Junction and Pore GABRB3 Mutations are Linked to Early-Onset Epileptic Encephalopathy

Ciria C Hernandez,Robert L Macdonald,Dingding Shen,Weijing Kong,Yujia Zhang,Xiaoyan Liu,Ningning Hu,Yuwu Jiang,Wangzhen Shen

doi:10.1038/s41598-017-16010-3

Ciria C Hernandez, Robert L Macdonald + Show 7 more

Open Access

https://doi.org/10.1038/s41598-017-16010-3

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Abstract

GABAA receptors are brain inhibitory chloride ion channels. Here we show functional analyses and structural simulations for three de novo missense mutations in the GABAA receptor β3 subunit gene (GABRB3) identified in patients with early-onset epileptic encephalopathy (EOEE) and profound developmental delay. We sought to obtain insights into the molecular mechanisms that might link defects in GABAA receptor biophysics and biogenesis to patients with EOEE. The mutant residues are part of conserved structural domains such as the Cys-loop (L170R) and M2-M3 loop (A305V) that form the GABA binding/channel gating coupling junction and the channel pore (T288N), which are functionally coupled during receptor activation. The mutant coupling junction residues caused rearrangements and formation of new hydrogen bonds in the open state, while the mutant pore residue reshaped the pore cavity. Whereas mutant coupling junction residues uncoupled during activation and caused gain of function, the mutant pore residue favoured low conductance receptors and differential sensitivity to diazepam and loss of function. These data reveal novel molecular mechanisms by which EOEE-linked mutations affect GABAA receptor function.

Highlights

Onset epileptic encephalopathies (EOEE) are a group of heterogeneous epilepsy disorders that are almost invariable associated with poor prognosis and are treatment-refractory[4]
Since GABRB3 has been reported to have a variety of mutations associated with diverse forms of epileptic encephalopathy syndromes, we speculated that the occurrence of the pathogenic mutant residues within conserved structural domains of GABAA receptors correlates with the dysfunction and the severity of the epileptic phenotype
We found that the impacts of the β3 subunit mutant residues L170R, T288N, and A305V on GABAA receptor function and biogenesis were quite different, and it was entirely dependent on the location of the mutant residue in the highly conserved regions of the GABAA receptor

Summary

Introduction

Onset epileptic encephalopathies (EOEE) are a group of heterogeneous epilepsy disorders that are almost invariable associated with poor prognosis and are treatment-refractory[4]. Since GABRB3 has been reported to have a variety of mutations associated with diverse forms of epileptic encephalopathy syndromes, we speculated that the occurrence of the pathogenic mutant residues within conserved structural domains of GABAA receptors correlates with the dysfunction and the severity of the epileptic phenotype. In vitro studies reported that the GABAA receptor function was differentially disrupted by the de novo β3 subunit mutant residues, D120N11, E180G11, Y184H9, L256Q9 and Y302C9,11, located at the GABA-binding interface and transmembrane domain that underlies channel activation, and these in turn were associated with the most severe forms of the EOEE epilepsy spectrum. We found that the mutant residues in this region uncouple channel activation mainly through perturbations in the coupling junction and the pore, and that these are the molecular mechanisms that underlie the epilepsy syndrome phenotype

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