NovelGLRA1Missense Mutation (P250T) in Dominant Hyperekplexia Defines an Intracellular Determinant of Glycine Receptor Channel Gating

Abstract

Missense mutations as well as a null allele of the human glycine receptor alpha1 subunit gene GLRA1 result in the neurological disorder hyperekplexia [startle disease, stiff baby syndrome, Mendelian Inheritance in Man (MIM) #149400]. In a pedigree showing dominant transmission of hyperekplexia, we identified a novel point mutation C1128A of GLRA1. This mutation encodes an amino acid substitution (P250T) in the cytoplasmic loop linking transmembrane regions M1 and M2 of the mature alpha1 polypeptide. After recombinant expression, homomeric alpha1(P250T) subunit channels showed a strong reduction of maximum whole-cell chloride currents and an altered desensitization, consistent with a prolonged recovery from desensitization. Apparent glycine binding was less affected, yielding an approximately fivefold increase in Ki values. Topological analysis predicts that the substitution of proline 250 leads to the loss of an angular polypeptide structure, thereby destabilizing open channel conformations. Thus, the novel GLRA1 mutant allele P250T defines an intracellular determinant of glycine receptor channel gating.

New Hyperekplexia Mutations Provide Insight into Glycine Receptor Assembly, Trafficking, and Activation Mechanisms

“…Consistent with previous studies (10,(37)(38)(39)(40), dominant mutations were expressed at the cell surface, thereby causing changes to the glycine sensitivity, conductance, and/or open probability. In contrast, recessive and compound heterozygous mutations mainly affected cell surface trafficking and insertion of receptors into the membrane (10,(41)(42)(43).…”

Allosteric and hyperekplexic mutant phenotypes investigated on an α 1 glycine receptor transmembrane structure

“…At the bottom of the channel, P250T confers reduced glycine sensitivity and increases the rate of desensitization on the α 1 GlyR (Table S2) (27). First, on Lily, P250T produces a very similar phenotype characterized by (i) a marked increase in desensitization kinetics, with current traces displaying a 20% vs. an 80% decay upon 1-s application of proton at pH 4 for WT and P250T, respectively (Fig.…”

Mechanisms for Picrotoxinin and Picrotin Blocks of α2 Homomeric Glycine Receptors

“…However, both hypotheses are equally plausible because an interaction between GlyR desensitization and channel gating is likely to occur. Indeed, the GlyR desensitization mechanism implies interactions between the M1 and M2 intracellular loop and the M2 transmembrane domain forming the channel pore of the receptor (33,34). It is likely that M1-M2 and M2-M3 loops act in parallel to control channel activation by acting as hinges governing allosteric control of the M2 domain (35).…”