title: De novo GABRG2 mutations associated with epileptic encephalopathies.
abstract: Epileptic encephalopathies are a devastating group of severe childhood onset epilepsies with medication-resistant seizures and poor developmental outcomes. Many epileptic encephalopathies have a genetic aetiology and are often associated with de novo mutations in genes mediating synaptic transmission, including GABAAreceptor subunit genes. Recently, we performed next generation sequencing on patients with a spectrum of epileptic encephalopathy phenotypes, and we identified five novel (A106T, I107T, P282S, R323W and F343L) and one known (R323Q) de novo GABRG2 pathogenic variants (mutations) in eight patients. To gain insight into the molecular basis for how these mutations contribute to epileptic encephalopathies, we compared the effects of the mutations on the properties of recombinant α1β2γ2L GABAAreceptors transiently expressed in HEK293T cells. Using a combination of patch clamp recording, immunoblotting, confocal imaging and structural modelling, we characterized the effects of these GABRG2 mutations on GABAAreceptor biogenesis and channel function. Compared with wild-type α1β2γ2L receptors, GABAAreceptors containing a mutant γ2 subunit had reduced cell surface expression with altered subunit stoichiometry or decreased GABA-evoked whole-cell current amplitudes, but with different levels of reduction. While a causal role of these mutations cannot be established directly from these results, the functional analysis together with the genetic information suggests that these GABRG2 variants may be major contributors to the epileptic encephalopathy phenotypes. Our study further expands the GABRG2 phenotypic spectrum and supports growing evidence that defects in GABAergic neurotransmission participate in the pathogenesis of genetic epilepsies including epileptic encephalopathies.
authors.affiliation: 1 The Graduate Program of Neuroscience, Vanderbilt University, Nashville, TN 37232, USA.
authors.forename: Dingding
authors.initials: D
authors.lastname: Shen
authors.affiliation: 2 Department of Neurology, Vanderbilt University, Nashville, TN 37240, USA.
authors.forename: Ciria C
authors.initials: CC
authors.lastname: Hernandez
authors.affiliation: 2 Department of Neurology, Vanderbilt University, Nashville, TN 37240, USA.
authors.forename: Wangzhen
authors.initials: W
authors.lastname: Shen
authors.affiliation: 2 Department of Neurology, Vanderbilt University, Nashville, TN 37240, USA.
authors.forename: Ningning
authors.initials: N
authors.lastname: Hu
authors.affiliation: 3 Epilepsy Genetics Program and the Department of Neurology, Boston Children's Hospital, Boston, MA 02115, USA.|4 Harvard Medical School, Boston, MA 02115, USA.
authors.forename: Annapurna
authors.initials: A
authors.lastname: Poduri
authors.affiliation: 3 Epilepsy Genetics Program and the Department of Neurology, Boston Children's Hospital, Boston, MA 02115, USA.
authors.forename: Beth
authors.initials: B
authors.lastname: Shiedley
authors.affiliation: 3 Epilepsy Genetics Program and the Department of Neurology, Boston Children's Hospital, Boston, MA 02115, USA.
authors.forename: Alex
authors.initials: A
authors.lastname: Rotenberg
authors.affiliation: 5 Division of Pediatric Neurology and Developmental Medicine, University of Basel Children's Hospital, Basel 4056, Switzerland.
authors.forename: Alexandre N
authors.initials: AN
authors.lastname: Datta
authors.affiliation: 6 Clinic for Children and Adolescents Dritter Orden, Divison of Neuropediatrics, München, 80638 Germany.
authors.forename: Steffen
authors.initials: S
authors.lastname: Leiz
authors.affiliation: 7 Clinic for Children and Adolescents, Halle/Saale, 06097 Germany.
authors.forename: Steffi
authors.initials: S
authors.lastname: Patzer
authors.affiliation: 8 Department of Pediatric Neurology, Kiel University, Kiel 24118 Germany; Northern German Epilepsy Centre for Children and Adolescents, Schwentinental - Raisdorf, 24223 Germany.
authors.forename: Rainer
authors.initials: R
authors.lastname: Boor
authors.affiliation: 9 Center for Rare Childhood Disorders, Translational Genomics Research Institute, Phoenix, 85004 AZ, USA.
authors.forename: Kerri
authors.initials: K
authors.lastname: Ramsey
authors.affiliation: 10 Departments of Neurology and Paediatrics, Division of Child Neurology, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, PA 19104, USA.|11 Children's Hospital of Philadelphia, Philadelphia, PA 19104, USA.
authors.forename: Ethan
authors.initials: E
authors.lastname: Goldberg
authors.affiliation: 10 Departments of Neurology and Paediatrics, Division of Child Neurology, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, PA 19104, USA.|11 Children's Hospital of Philadelphia, Philadelphia, PA 19104, USA.
authors.forename: Ingo
authors.initials: I
authors.lastname: Helbig
authors.affiliation: 10 Departments of Neurology and Paediatrics, Division of Child Neurology, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, PA 19104, USA.|11 Children's Hospital of Philadelphia, Philadelphia, PA 19104, USA.
authors.forename: Xilma R
authors.initials: XR
authors.lastname: Ortiz-Gonzalez
authors.affiliation: 12 Institute of Human Genetics, University of Leipzig Hospitals and Clinics, Leipzig, 04103 Germany.
authors.forename: Johannes R
authors.initials: JR
authors.lastname: Lemke
authors.affiliation: 10 Departments of Neurology and Paediatrics, Division of Child Neurology, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, PA 19104, USA.|11 Children's Hospital of Philadelphia, Philadelphia, PA 19104, USA.
authors.forename: Eric D
authors.initials: ED
authors.lastname: Marsh
authors.affiliation: 2 Department of Neurology, Vanderbilt University, Nashville, TN 37240, USA robert.macdonald@vanderbilt.edu.
authors.forename: Robert L
authors.initials: RL
authors.lastname: Macdonald
mesh_terms: D002648:Child; D002675:Child, Preschool; D000069279:Drug Resistant Epilepsy / Q000235:genetics* / Q000503:physiopathology*; D055724:Electrophysiological Phenomena; D004827:Epilepsy / Q000235:genetics* / Q000503:physiopathology*; D059472:Exome; D005260:Female; D057809:HEK293 Cells; D006801:Humans; D008297:Male; D009154:Mutation; D018408:Patch-Clamp Techniques; D010641:Phenotype; D011963:Receptors, GABA-A / Q000235:genetics*
chemical_list: C546279:GABRG2 protein, human; D011963:Receptors, GABA-A
keywords: GABAA receptor; GABRG2; de novo mutation; epileptic encephalopathy; next generation sequencing