title: Disturbed neuronal ER-Golgi sorting of unassembled glycine receptors suggests altered subcellular processing is a cause of human hyperekplexia.
abstract: Recent studies on the pathogenic mechanisms of recessive hyperekplexia indicate disturbances in glycine receptor (GlyR) α1 biogenesis. Here, we examine the properties of a range of novel glycine receptor mutants identified in human hyperekplexia patients using expression in transfected cell lines and primary neurons. All of the novel mutants localized in the large extracellular domain of the GlyR α1 have reduced cell surface expression with a high proportion of receptors being retained in the ER, although there is forward trafficking of glycosylated subpopulations into the ER-Golgi intermediate compartment and cis-Golgi compartment. CD spectroscopy revealed that the mutant receptors have proportions of secondary structural elements similar to wild-type receptors. Two mutants in loop B (G160R, T162M) were functional, but none of those in loop D/β2-3 were. One nonfunctional truncated mutant (R316X) could be rescued by coexpression with the lacking C-terminal domain. We conclude that a proportion of GlyR α1 mutants can be transported to the plasma membrane but do not necessarily form functional ion channels. We suggest that loop D/β2-3 is an important determinant for GlyR trafficking and functionality, whereas alterations to loop B alter agonist potencies, indicating that residues here are critical elements in ligand binding.
authors.affiliation: Institute for Clinical Neurobiology, Julius-Maximilians-University of Würzburg, 97078 Würzburg, Germany.
authors.forename: Natascha
authors.initials: N
authors.lastname: Schaefer
authors.affiliation: Institute of Biochemistry, Department of Biochemistry and Molecular Medicine, Friedrich-Alexander-University Erlangen-Nürnberg, 91054 Erlangen, Germany.
authors.forename: Christoph J
authors.initials: CJ
authors.lastname: Kluck
authors.affiliation: Department of Biochemistry, University of Cambridge, Cambridge CB2 1QW, United Kingdom.
authors.forename: Kerry L
authors.initials: KL
authors.lastname: Price
authors.affiliation: Institute of Biochemistry, Bioinformatics Department, Friedrich-Alexander-University Erlangen-Nürnberg, 91054 Erlangen, Germany.
authors.forename: Heike
authors.initials: H
authors.lastname: Meiselbach
authors.affiliation: Institute for Clinical Neurobiology, Julius-Maximilians-University of Würzburg, 97078 Würzburg, Germany.
authors.forename: Nadine
authors.initials: N
authors.lastname: Vornberger
authors.affiliation: Research Center for Bio-Macromolecules and Department of Biopolymers, University Bayreuth, 95447 Bayreuth, Germany.
authors.forename: Stephan
authors.initials: S
authors.lastname: Schwarzinger
authors.affiliation: Institute of Biochemistry, Department of Biochemistry and Molecular Medicine, Friedrich-Alexander-University Erlangen-Nürnberg, 91054 Erlangen, Germany.
authors.forename: Stephanie
authors.initials: S
authors.lastname: Hartmann
authors.affiliation: Institute for Clinical Neurobiology, Julius-Maximilians-University of Würzburg, 97078 Würzburg, Germany.
authors.forename: Georg
authors.initials: G
authors.lastname: Langlhofer
authors.affiliation: Center of Human Genetics and.
authors.forename: Solveig
authors.initials: S
authors.lastname: Schulz
authors.affiliation: Department of Neuropediatrics, Jena University Hospital, 07743 Jena, Germany.
authors.forename: Nadja
authors.initials: N
authors.lastname: Schlegel
authors.affiliation: Interdisciplinary Pediatric Center for Children with Developmental Disabilities and Severe Chronic Disorders, University Medical Center, Georg August University, 37075 Göttingen, Germany, and.
authors.forename: Knut
authors.initials: K
authors.lastname: Brockmann
authors.affiliation: Temple Street Children's University Hospital, Dublin, Dublin1, Ireland.
authors.forename: Bryan
authors.initials: B
authors.lastname: Lynch
authors.affiliation: Institute of Biochemistry, Department of Biochemistry and Molecular Medicine, Friedrich-Alexander-University Erlangen-Nürnberg, 91054 Erlangen, Germany.
authors.forename: Cord-Michael
authors.initials: CM
authors.lastname: Becker
authors.affiliation: Department of Biochemistry, University of Cambridge, Cambridge CB2 1QW, United Kingdom.
authors.forename: Sarah C R
authors.initials: SC
authors.lastname: Lummis
authors.affiliation: Institute for Clinical Neurobiology, Julius-Maximilians-University of Würzburg, 97078 Würzburg, Germany, Villmann_C@ukw.de.
authors.forename: Carmen
authors.initials: C
authors.lastname: Villmann
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002648:Child; D002522:Chlorocebus aethiops; D004721:Endoplasmic Reticulum / Q000235:genetics / Q000378:metabolism*; D005260:Female; D006056:Golgi Apparatus / Q000235:genetics / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D007223:Infant; D042541:Intracellular Space / Q000378:metabolism*; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D009474:Neurons / Q000378:metabolism*; D010375:Pedigree; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D018009:Receptors, Glycine / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D016750:Stiff-Person Syndrome / Q000175:diagnosis / Q000235:genetics / Q000378:metabolism*
chemical_list: D018009:Receptors, Glycine
keywords: assembly; biogenesis; glycine receptor; human hyperekplexia; rescue of function; subcompartimentalization