Characterization of a novel missense mutation in the α2 subunit of the neuronal nicotinic acetylcholine receptor linked to sleep-related generalized seizures with cognitive deficit

Aims: Mutations in the CHRNA4, CHRNB2, and CHRNA2 genes, respectively encoding the α4, β2 and α2 subunits of the neuronal nicotinic receptors (nAChRs) are linked to autosomal dominant sleep-related hypermotor epilepsy. The pathogenesis is uncertain, as different nAChR subunits exert complex physiological roles in the mature neocortex as well as during synaptogenesis.Methods: We generated different F2A-based tri-cistronic constructs for equimolar expression of α2/β4 nAChR subunits and carried out functional tests by patch-clamp recording on HEK 293 cells. Whole-cell currents were elicited by fast application of nicotine or ACh, at -70/-80 mV.Results: a novel missense mutation in the CHRNA2 gene (c.926T>C; p.Val309Ala) was detected in heterozygosity in two members of a family characterized by sleep-related tonic-clonic epileptic seizures accompanied by cognitive deficit. The mutation is located in a highly conserved region of the third transmembrane domain (M3), which closely interact with the pore-lining M2 segments during the gating conformational rearrangement operated by ACh binding. The mutation is described as potentially deleterious by PolyPhen-2, SIFT and MutationTaster predictors. In homo- and heterozygous condition, mutant α2 decreased the nAChR response to nicotine and ACh. The peak current density tested with 100 µM nicotine was 143 ± 11.7 pA/pF in WT and decreased by approximately 75% in both homo- and heterozygous condition. Similar effects were produced by 100 µM ACh (WT: 7.6 ± 1.25 pA/pF, n=13; homozygote: 2 ± 0.32 pA/pF, n=27; p < 0.001, Student t-test).Conclusions: loss-of-function mutations in CHRNA2 could contribute to sleep-related tonic-clonic seizures associated with cognitive deficit.