title: A recurrent KCNQ2 pore mutation causing early onset epileptic encephalopathy has a moderate effect on M current but alters subcellular localization of Kv7 channels.
abstract: Mutations in the KCNQ2 gene encoding the voltage-dependent potassium M channel Kv7.2 subunit cause either benign epilepsy or early onset epileptic encephalopathy (EOEE). It has been proposed that the disease severity rests on the inhibitory impact of mutations on M current density. Here, we have analyzed the phenotype of 7 patients carrying the p.A294V mutation located on the S6 segment of the Kv7.2 pore domain (Kv7.2(A294V)). We investigated the functional and subcellular consequences of this mutation and compared it to another mutation (Kv7.2(A294G)) associated with a benign epilepsy and affecting the same residue. We report that all the patients carrying the p.A294V mutation presented the clinical and EEG characteristics of EOEE. In CHO cells, the total expression of Kv7.2(A294V) alone, assessed by western blotting, was only 20% compared to wild-type. No measurable current was recorded in CHO cells expressing Kv7.2(A294V) channel alone. Although the total Kv7.2(A294V) expression was rescued to wild-type levels in cells co-expressing the Kv7.3 subunit, the global current density was still reduced by 83% compared to wild-type heteromeric channel. In a configuration mimicking the patients' heterozygous genotype i.e., Kv7.2(A294V)/Kv7.2/Kv7.3, the global current density was reduced by 30%. In contrast to Kv7.2(A294V), the current density of homomeric Kv7.2(A294G) was not significantly changed compared to wild-type Kv7.2. However, the current density of Kv7.2(A294G)/Kv7.2/Kv7.3 and Kv7.2(A294G)/Kv7.3 channels were reduced by 30% and 50% respectively, compared to wild-type Kv7.2/Kv7.3. In neurons, the p.A294V mutation induced a mislocalization of heteromeric mutant channels to the somato-dendritic compartment, while the p.A294G mutation did not affect the localization of the heteromeric channels to the axon initial segment. We conclude that this position is a hotspot of mutation that can give rise to a severe or a benign epilepsy. The p.A294V mutation does not exert a dominant-negative effect on wild-type subunits but alters the preferential axonal targeting of heteromeric Kv7 channels. Our data suggest that the disease severity is not necessarily a consequence of a strong inhibition of M current and that additional mechanisms such as abnormal subcellular distribution of Kv7 channels could be determinant.
authors.affiliation: Aix-Marseille Université, GMGF, Marseille, France; INSERM, UMR_S 910, Marseille, France.
authors.forename: Affef
authors.initials: A
authors.lastname: Abidi
authors.affiliation: Aix-Marseille Université, CNRS, CRN2M-UMR7286, Marseille, France.
authors.forename: Jérôme J
authors.initials: JJ
authors.lastname: Devaux
authors.affiliation: Aix-Marseille Université, Institut de Neurobiologie de la Méditerranée (INMED), Marseille, France; INSERM, UMR_S 901, Marseille, France.
authors.forename: Florence
authors.initials: F
authors.lastname: Molinari
authors.affiliation: Aix-Marseille Université, GMGF, Marseille, France; INSERM, UMR_S 910, Marseille, France.
authors.forename: Gisèle
authors.initials: G
authors.lastname: Alcaraz
authors.affiliation: Aix-Marseille Université, Institut de Neurobiologie de la Méditerranée (INMED), Marseille, France; INSERM, UMR_S 901, Marseille, France.
authors.forename: François-Xavier
authors.initials: FX
authors.lastname: Michon
authors.affiliation: Aix-Marseille Université, GMGF, Marseille, France; INSERM, UMR_S 910, Marseille, France; APHM, Hôpital d'Enfants de la Timone, Service de neurologie pédiatrique, Marseille, France.
authors.forename: Julie
authors.initials: J
authors.lastname: Sutera-Sardo
authors.affiliation: Aix-Marseille Université, Institut de Neurobiologie de la Méditerranée (INMED), Marseille, France; INSERM, UMR_S 901, Marseille, France.
authors.forename: Hélène
authors.initials: H
authors.lastname: Becq
authors.affiliation: Aix-Marseille Université, GMGF, Marseille, France; INSERM, UMR_S 910, Marseille, France; APHM, Hôpital d'enfants de la Timone, Département de génétique médicale et de biologie cellulaire, Marseille France.
authors.forename: Caroline
authors.initials: C
authors.lastname: Lacoste
authors.affiliation: CHU Besançon, Service de génétique et neuropédiatrie, Besançon, France.
authors.forename: Cécilia
authors.initials: C
authors.lastname: Altuzarra
authors.affiliation: Université Pierre et Marie Curie, Groupe de Recherche Clinique « Déficiences Intellectuelles de Causes Rares », Paris, France; APHP, service de neurologie pédiatrique, Hôpital Trousseau, Paris, France.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Afenjar
authors.affiliation: APHP, Service de Génétique Médicale et Centre de Références « Déficiences Intellectuelles de Causes Rares », Groupe Hospitalier Pitié-Salpêtrière, Paris, France; Université Pierre et Marie Curie, Groupe de Recherche Clinique « Déficiences Intellectuelles de Causes Rares », Paris, France.
authors.forename: Cyril
authors.initials: C
authors.lastname: Mignot
authors.affiliation: APHP, service de neurologie pédiatrique, Hôpital Trousseau, Paris, France.
authors.forename: Diane
authors.initials: D
authors.lastname: Doummar
authors.affiliation: CHU de Nantes, Service de génétique médicale, Nantes, France.
authors.forename: Bertrand
authors.initials: B
authors.lastname: Isidor
authors.affiliation: CHU d'Angers, Service de neurologie pédiatrique, Angers, France.
authors.forename: Sylvie N
authors.initials: SN
authors.lastname: Guyen
authors.affiliation: CHU d'Angers, Département de Biochimie et Génétique, Angers, France.
authors.forename: Estelle
authors.initials: E
authors.lastname: Colin
authors.affiliation: CHU de Tours, Service de neurologie pédiatrique, Tours, France.
authors.forename: Sabine
authors.initials: S
authors.lastname: De La Vaissière
authors.affiliation: CHU de Tours, Service de génétique, Tours, France.
authors.forename: Damien
authors.initials: D
authors.lastname: Haye
authors.affiliation: CHU de Lille, Service de neurologie pédiatrique, Lille, France.
authors.forename: Adeline
authors.initials: A
authors.lastname: Trauffler
authors.affiliation: Aix-Marseille Université, GMGF, Marseille, France; INSERM, UMR_S 910, Marseille, France; APHM, Hôpital d'enfants de la Timone, Département de génétique médicale et de biologie cellulaire, Marseille France.
authors.forename: Catherine
authors.initials: C
authors.lastname: Badens
authors.affiliation: CHU de Saint Etienne, Service de génétique médicale, France.
authors.forename: Fabienne
authors.initials: F
authors.lastname: Prieur
authors.affiliation: Hospices Civils de Lyon, Service de génétique, Lyon, France.
authors.forename: Gaetan
authors.initials: G
authors.lastname: Lesca
authors.affiliation: Aix-Marseille Université, GMGF, Marseille, France; INSERM, UMR_S 910, Marseille, France.
authors.forename: Laurent
authors.initials: L
authors.lastname: Villard
authors.affiliation: Aix-Marseille Université, GMGF, Marseille, France; INSERM, UMR_S 910, Marseille, France; APHM, Hôpital d'Enfants de la Timone, Service de neurologie pédiatrique, Marseille, France. Electronic address: mathieu.milh@ap-hm.fr.
authors.forename: Mathieu
authors.initials: M
authors.lastname: Milh
authors.affiliation: Aix-Marseille Université, Institut de Neurobiologie de la Méditerranée (INMED), Marseille, France; INSERM, UMR_S 901, Marseille, France. Electronic address: laurent.aniksztejn@inserm.fr.
authors.forename: Laurent
authors.initials: L
authors.lastname: Aniksztejn
mesh_terms: D000818:Animals; D001921:Brain / Q000378:metabolism / Q000503:physiopathology*; D016466:CHO Cells; D002478:Cells, Cultured; D003412:Cricetulus; D004827:Epilepsy / Q000175:diagnosis / Q000235:genetics* / Q000503:physiopathology; D006624:Hippocampus / Q000378:metabolism; D006801:Humans; D051658:KCNQ2 Potassium Channel / Q000235:genetics* / Q000378:metabolism / Q000502:physiology*; D009154:Mutation; D009474:Neurons / Q000378:metabolism; D010641:Phenotype
chemical_list: D051658:KCNQ2 Potassium Channel; C496914:KCNQ2 protein, human
keywords: Early epileptic encephalopathy; Kv7 channels; M-current; Subcellular channel expression; p.A294G mutation; p.A294V mutation