Distinct Functional Alterations and Therapeutic Options of Two Pathological De Novo Variants of the T292 Residue of GABRA1 Identified in Children with Epileptic Encephalopathy and Neurodevelopmental Disorders.

Wenlin Chen,Yu Tian Wang,Yang Ge,Romi Juneja,Lidong Liu,Joshua Melo,Yee Wah So,Jie Lu

doi:10.3390/ijms23052723

Wenlin Chen, Yu Tian Wang + Show 6 more

Open Access

https://doi.org/10.3390/ijms23052723

Copy DOI

Abstract

Mutations of GABAAR have reportedly led to epileptic encephalopathy and neurodevelopmental disorders. We have identified a novel de novo T292S missense variant of GABRA1 from a pediatric patient with grievous global developmental delay but without obvious epileptic activity. This mutation coincidentally occurs at the same residue as that of a previously reported GABRA1 variant T292I identified from a pediatric patient with severe epilepsy. The distinct phenotypes of these two patients prompted us to compare the impacts of the two mutants on the receptor function and to search for suitable therapeutics. In this study, we used biochemical techniques and patch-clamp recordings in HEK293 cells overexpressing either wild-type or mutated rat recombinant GABAARs. We found that the α1T292S variant significantly increased GABA-evoked whole-cell currents, shifting the dose–response curve to the left without altering the maximal response. In contrast, the α1T292I variant significantly reduced GABA-evoked currents, shifting the dose–response curve to the right with a severely diminished maximum response. Single-channel recordings further revealed that the α1T292S variant increased, while the α1T292I variant decreased the GABAAR single-channel open time and open probability. Importantly, we found that the T292S mutation-induced increase in GABAAR function could be fully normalized by the negative GABAAR modulator thiocolchicoside, whereas the T292I mutation-induced impairment of GABAAR function was largely rescued with a combination of the GABAAR positive modulators diazepam and verapamil. Our study demonstrated that α1T292 is a critical residue for controlling GABAAR channel gating, and mutations at this residue may produce opposite impacts on the function of the receptors. Thus, the present work highlights the importance of functionally characterizing each individual GABAAR mutation for ensuring precision medicine.

Highlights

Licensee MDPI, Basel, Switzerland.Epilepsy is a neurological disorder characterized by recurrent seizures that occur due to abnormally excessive electrical discharges of the cerebral neurons as a result of the disruption of the excitation–inhibition balance (E/I balance) in the brain [1–3]
Through screening several clinically approved drugs that reportedly act on GABAA Rs, we found that the GABAA R allosteric inhibitor thiocolchicoside (TCC) [30] could reduce the gain of function caused by the T292S mutation, thereby normalizing the receptor function
Through whole-exome sequencing (WES), we identified a novel de novo T292S (C875G) missense variant of GABRA1

Summary

Introduction

Epilepsy is a neurological disorder characterized by recurrent seizures that occur due to abnormally excessive electrical discharges of the cerebral neurons as a result of the disruption of the excitation–inhibition balance (E/I balance) in the brain [1–3]. Recent studies have identified several hundred genes associated with epilepsy, among which genes encoding ion channels/receptors predominate [3,5]. Synaptic neurotransmission in the central nervous system (CNS), the γ-aminobutyric acid (GABA) type A receptor (GABAA R) plays a critical role in maintaining the neuronal E/I balance in the CNS and is considered an important genetic risk factor for epilepsy. The dysfunction of the receptor underlies the pathogenesis of many neurological diseases. Accumulating evidence suggests that genetic variants of the GABAA R may cause conditions including epileptic encephalopathy (EE) [6–9], developmental delay [9,10], Fragile X, Rett Syndrome, and Dravet Syndrome [11–13]

Methods

Results

Discussion

Conclusion