Abstracts

Rationale: KCNA2 encodes voltage-gated potassium channel Kv1.2, abundantly expressed in the brain. De novo mutations in this gene have been associated with epileptic encephalopathy and hereditary spastic paraplegia. A family with a spectrum of milder epilepsy has also been reported. Furthermore, KCNA2 was shown as one of the genes harbored by microdeletions that were detected in genetic generalized epilepsy (GGE) patients. We studied functionally three KCNA2 variants detected in patients with GGE, which were recruited through the multicentre Epilepsy Phenome/Genome Project and Epi4K consortium. None of the three variants, G60E, W150C, and V339G, was found in the Exome Aggregation Consortium (ExAC) database or the Genome Aggregation Database (gnomAD). Methods: Mutations were introduced in WT KCNA2 cDNA inserted into the pcDNA3 vector, which served as a template for cRNA preparation. Equal amounts of WT and mutant cRNA were injected into Xenopus laevis oocytes on the same day and automated two-microelectrode voltage clamp recordings were performed in parallel on days 2 and 3 after injection. Results: Two of the analyzed mutations, V60E and W150C, showed a prominent loss of function by reducing the maximum current amplitudes to about 5 % and 20 % of the WT current amplitudes, respectively. In contrast, the V339G variant caused a 5-fold increase of the maximum current amplitude compared to the WT. In addition, a 20 mV leftward shift in the voltage-dependence of activation was also observed for this variant, indicating an overall gain of function pathomechanism. Conclusions: Our results indicate that the GGE-associated variants cause both gain and loss of function of the Kv1.2 channel. Further analysis is necessary to understand how these functional alterations may lead to seizures and GGE. Funding: National Health and Medical Research Council of Australia through a Program grant 10915693.