title: A novel hyperekplexia-causing mutation in the pre-transmembrane segment 1 of the human glycine receptor alpha1 subunit reduces membrane expression and impairs gating by agonists.
abstract: In this study, we have compared the functional consequences of three mutations (R218Q, V260M, and Q266H) in the alpha(1) subunit of the glycine receptor (GlyRA1) causing hyperekplexia, an inherited neurological channelopathy. In HEK-293 cells, the agonist EC(50s) for glycine-activated Cl(-) currents were increased from 26 microm in wtGlyRA1, to 5747, 135, and 129 microm in R218Q, V260M, and Q266H GlyRA1 channels, respectively. Cl(-) currents elicited by beta-alanine and taurine, which behave as agonists at wtGlyRA1, were decreased in V260M and Q266H mutant receptors and virtually abolished in GlyRA1 R218Q receptors. Gly-gated Cl(-) currents were similarly antagonized by low concentrations of strychnine in both wild-type (wt) and R218Q GlyRA1 channels, suggesting that the Arg-218 residue plays a crucial role in GlyRA1 channel gating, with only minor effects on the agonist/antagonist binding site, a hypothesis supported by our molecular model of the GlyRA1 subunit. The R218Q mutation, but not the V260M or the Q266H mutation, caused a marked decrease of receptor subunit expression both in total cell lysates and in isolated plasma membrane proteins. This decreased expression does not seem to explain the reduced agonist sensitivity of GlyRA1 R218Q channels since no difference in the apparent sensitivity to glycine or taurine was observed when wtGlyRA1 receptors were expressed at levels comparable with those of R218Q mutant receptors. In conclusion, multiple mechanisms may explain the dramatic decrease in GlyR function caused by the R218Q mutation, possibly providing the molecular basis for its association with a more severe clinical phenotype.
authors.affiliation: Division of Pharmacology, Department of Neuroscience, School of Medicine, University of Naples Federico II, 80131 Naples, Italy.
authors.forename: Pasqualina
authors.initials: P
authors.lastname: Castaldo
authors.forename: Patrizia
authors.initials: P
authors.lastname: Stefanoni
authors.forename: Francesco
authors.initials: F
authors.lastname: Miceli
authors.forename: Giangennaro
authors.initials: G
authors.lastname: Coppola
authors.forename: Emanuele Miraglia
authors.initials: EM
authors.lastname: Del Giudice
authors.forename: Giulia
authors.initials: G
authors.lastname: Bellini
authors.forename: Antonio
authors.initials: A
authors.lastname: Pascotto
authors.forename: James R
authors.initials: JR
authors.lastname: Trudell
authors.forename: Neil L
authors.initials: NL
authors.lastname: Harrison
authors.forename: Lucio
authors.initials: L
authors.lastname: Annunziato
authors.forename: Maurizio
authors.initials: M
authors.lastname: Taglialatela
mesh_terms: D001927:Brain Diseases / Q000235:genetics*; D006801:Humans; D015640:Ion Channel Gating; D008958:Models, Molecular; D009154:Mutation; D018009:Receptors, Glycine / Q000737:chemistry / Q000235:genetics* / Q000502:physiology; D013216:Reflex, Startle; D013329:Structure-Activity Relationship; D013331:Strychnine / Q000494:pharmacology
chemical_list: D018009:Receptors, Glycine; D013331:Strychnine