CHRNA2 and Nocturnal Frontal Lobe Epilepsy: Identification and Characterization of a Novel Loss of Function Mutation

Abstract

Mutations in genes coding for subunits of the neuronal nicotinic acetylcholine receptor (nAChR) have been involved in familial sleep-related hypermotor epilepsy (also named autosomal dominant nocturnal frontal lobe epilepsy, ADNFLE). Most of these mutations reside in CHRNA4 and CHRNB2 genes, coding for the α4 and β2 nAChR subunits, respectively. Two mutations with contrasting functional effects were also identified in the CHRNA2 gene coding for the α2 subunit. Here, we report the third mutation in the CHRNA2 , found in a patient showing ADNFLE. The patient was examined by scalp EEG, contrast-enhanced brain magnetic resonance imaging (MRI), and nocturnal video-polysomnographic recording. All exons and the exon-intron boundaries of CHRNA2 , CHRNA4 , CHRNB2 , CRH , KCNT1 were amplified and Sanger sequenced. In the proband, we found a c.754T>C (p.Tyr252His) missense mutation located in the N-terminal ligand-binding domain and inherited from the mother. Functional studies were performed by transient co-expression of α2 and α2 Tyr 252 His , with either β2 or β4, in human embryonic kidney (HEK293) cells. Equimolar amounts of subunits expression were obtained by using F2A-based multi-cistronic constructs encoding for the genes relative to the nAChR subunits of interest and for the enhanced green fluorescent protein. The mutation reduced the maximal currents by approximately 80% in response to saturating concentrations of nicotine in homo- and heterozygous form, in both the α2β4 and α2β2 nAChR subtypes. The effect was accompanied by a strong right-shift of the concentration-response to nicotine. Similar effects were observed using ACh. Negligible effects were produced by α2 Tyr252His on the current reversal potential. Moreover, binding of (±)-[ 3 H]Epibatidine revealed an approximately 10-fold decrease of both K d and B max (bound ligand in saturating conditions), in cells expressing α2 Tyr252His . The reduced B max and whole-cell currents were not caused by a decrease in mutant receptor expression, as minor effects were produced by α2 Tyr252His on the level of transcripts and the membrane expression of α2β4 nAChR. Overall, these results suggest that α2 Tyr252His strongly reduced the number of channels bound to the agonist, without significantly altering the overall channel expression. We conclude that mutations in CHRNA2 are more commonly linked to ADNFLE than previously thought, and may cause a loss-of-function phenotype.

Cytisine Exerts an Anti-Epileptic Effect via α7nAChRs in a Rat Model of Temporal Lobe Epilepsy

“…Recently, increasing data have suggested that acetylcholine (ACh) and cholinergic neurotransmission participate in the pathophysiology of epilepsy ( Ghasemi and Hadipour-Niktarash, 2015 ; Biagioni et al, 2019 ; Meller et al, 2019 ). Genetic studies have also demonstrated that mutations in neuronal nicotinic ACh receptors (nAChRs) are responsible for some specific forms of epileptic disorders, such as juvenile myoclonic epilepsy (JME) and autosomal-dominant nocturnal frontal lobe epilepsy (ADNFLE) ( Rozycka et al, 2009 ; Rozycka et al, 2013 ; Weltzin et al, 2016 ; Villa et al, 2019 ). Neural nAChRs are extensively distributed in the CNS and represent a large family of ligand-gated ion channels constructed from combinations of α (α2–α10) and β (β2–β4) subunits ( Gotti et al, 2006 ).…”

“…Among them, homomeric α7 and heteromeric α4β2 pentameric nAChRs are predominant subtypes in the brain ( Gotti et al, 2006 ). It has been reported that α7 and α4β2 subtypes of nAChRs might be linked to some idiopathic types of epilepsy, including JME and ADNFLE ( Rozycka et al, 2009 ; Rozycka et al, 2013 ; Weltzin et al, 2016 ; Villa et al, 2019 ). Therefore, nAChR abnormalities may serve as a biomarker in epilepsy with a genetic background ( Garibotto et al, 2019 ), suggesting that nAChRs may also be a potential target for the treatment of epilepsy.…”

Germline Mutation Enrichment in Pathways Controlling Endothelial Cell Homeostasis in Patients with Brain Arteriovenous Malformation: Implication for Molecular Diagnosis

“…In the end, we focused our attention on the GO terms regarding the response to hypoxia (GO:0001666, GO:0036293, GO:0070482) with the patient as a carrier of rs55685423 (c.1191G>C, p.Gln397His) in the CHRNB2 locus. This gene encodes for the cholinergic receptor nicotinic beta 2 subunit [ 40 ]. Its expression was demonstrated in HBMECs, where it contributes to capillary network formation and to angiogenic response to inflammation [ 41 ].…”

Nicotine: A Targeted Therapy for Epilepsy Due to nAChR Gene Variants

“…23 Recent data have noted CHRNA2 that some mutations associated with an autosomal dominant sleep-related hypermotor epilepsy phenotype have a physiologic loss of function of the nAChR. 24 It is unclear if nicotine therapy would be beneficial in this population, and potential targeted therapies should be further explored.…”