title: Characterization of a de novo SCN8A mutation in a patient with epileptic encephalopathy.
abstract: OBJECTIVE
  Recently, de novo SCN8A missense mutations have been identified as a rare dominant cause of epileptic encephalopathies (EIEE13). Functional studies on the first described case demonstrated gain-of-function effects of the mutation. We describe a novel de novo mutation of SCN8A in a patient with epileptic encephalopathy, and functional characterization of the mutant protein.
  
  
  METHODS
  Whole exome sequencing was used to discover the variant. We generated a mutant cDNA, transfected HEK293 cells, and performed Western blotting to assess protein stability. To study channel functional properties, patch-clamp experiments were carried out in transfected neuronal ND7/23 cells.
  
  
  RESULTS
  The proband exhibited seizure onset at 6 months of age, diffuse brain atrophy, and more profound developmental impairment than the original case. The mutation p.Arg233Gly in the voltage sensing transmembrane segment D1S4 was present in the proband and absent in both parents. This mutation results in a temperature-sensitive reduction in protein expression as well as reduced sodium current amplitude and density and a relative increased response to a slow ramp stimulus, though this did not result in an absolute increased current at physiological temperatures.
  
  
  CONCLUSIONS
  The new de novo SCN8A mutation is clearly deleterious, resulting in an unstable protein with reduced channel activity. This differs from the gain-of-function attributes of the first SCN8A mutation in epileptic encephalopathy, pointing to heterogeneity of mechanisms. Since Nav1.6 is expressed in both excitatory and inhibitory neurons, a differential effect of a loss-of-function of Nav1.6 Arg223Gly on inhibitory interneurons may underlie the epilepsy phenotype in this patient.
authors.affiliation: Department of Medical Genetics, University Medical Center Utrecht, Utrecht, The Netherlands. Electronic address: c.dekovel@umcutrecht.nl.
authors.forename: Carolien G F
authors.initials: CG
authors.lastname: de Kovel
authors.affiliation: Department of Human Genetics, University of Michigan, Ann Arbor, MI 48109-5618, USA; Department of Neurology, University of Michigan, Ann Arbor, MI 48109-5618, USA.
authors.forename: Miriam H
authors.initials: MH
authors.lastname: Meisler
authors.affiliation: Department of Medical Genetics, University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: Eva H
authors.initials: EH
authors.lastname: Brilstra
authors.affiliation: Department of Pediatrics, University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: Frederique M C
authors.initials: FM
authors.lastname: van Berkestijn
authors.affiliation: Department of Medical Genetics, University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: Ruben
authors.initials: R
authors.lastname: van 't Slot
authors.affiliation: Department of Medical Genetics, University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: Stef
authors.initials: S
authors.lastname: van Lieshout
authors.affiliation: Department of Medical Genetics, University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: Isaac J
authors.initials: IJ
authors.lastname: Nijman
authors.affiliation: Department of Human Genetics, University of Michigan, Ann Arbor, MI 48109-5618, USA.
authors.forename: Janelle E
authors.initials: JE
authors.lastname: O'Brien
authors.affiliation: Arizona Research Laboratories, Division of Biotechnology, University of Arizona, Tucson, AZ 85721, USA.
authors.forename: Michael F
authors.initials: MF
authors.lastname: Hammer
authors.affiliation: Department of Neurology, Center for Neuroscience and Regeneration Research, Yale University School of Medicine, New Haven, CT 06510, USA; Rehabilitation Research Center, Veterans Affairs Connecticut Healthcare System, West Haven, CT 06516, USA.
authors.forename: Mark
authors.initials: M
authors.lastname: Estacion
authors.affiliation: Department of Neurology, Center for Neuroscience and Regeneration Research, Yale University School of Medicine, New Haven, CT 06510, USA; Rehabilitation Research Center, Veterans Affairs Connecticut Healthcare System, West Haven, CT 06516, USA.
authors.forename: Stephen G
authors.initials: SG
authors.lastname: Waxman
authors.affiliation: Department of Neurology, Center for Neuroscience and Regeneration Research, Yale University School of Medicine, New Haven, CT 06510, USA; Rehabilitation Research Center, Veterans Affairs Connecticut Healthcare System, West Haven, CT 06516, USA.
authors.forename: Sulayman D
authors.initials: SD
authors.lastname: Dib-Hajj
authors.affiliation: Department of Medical Genetics, University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: Bobby P C
authors.initials: BP
authors.lastname: Koeleman
mesh_terms: D001120:Arginine / Q000235:genetics; D002675:Child, Preschool; D004827:Epilepsy / Q000150:complications / Q000235:genetics*; D005260:Female; D005998:Glycine / Q000235:genetics; D057809:HEK293 Cells; D006801:Humans; D008564:Membrane Potentials / Q000235:genetics; D008831:Microcephaly / Q000150:complications / Q000235:genetics; D009154:Mutation / Q000235:genetics*; D062557:NAV1.6 Voltage-Gated Sodium Channel / Q000235:genetics*; D018408:Patch-Clamp Techniques; D013696:Temperature; D014162:Transfection
chemical_list: D062557:NAV1.6 Voltage-Gated Sodium Channel; C100681:SCN8A protein, human; D001120:Arginine; D005998:Glycine
keywords: Epileptic encephalopathy; Exome sequencing; Nav1.6; Patch-clamp; SCN8A