The GLRA1 Missense Mutation W170S Associates Lack of Zn²⁺Potentiation with Human Hyperekplexia

Abstract

Hyperekplexia is a neurological disorder associated primarily with mutations in the ␣1 subunit of glycine receptors (GlyRs) that lead to dysfunction of glycinergic inhibitory transmission. To date, most of the identified mutations result in disruption of surface expression or altered channel properties of ␣1-containing GlyRs. Little evidence has emerged to support an involvement of allosteric GlyR modulation in human hyperekplexia. Here, we report that recombinant human GlyRs containing ␣1 or ␣1␤ subunits with a missense mutation in the ␣1 subunit (W170S), previously identified from familial hyperekplexia, caused remarkably reduced potentiation and enhanced inhibition by Zn 2ϩ . Interestingly, mutant ␣1 W170S ␤ GlyRs displayed no significant changes in potency or maximum response to glycine, taurine, or ␤-alanine. By temporally separating the potentiating and the inhibitory effects of Zn 2ϩ , we found that the enhancement of Zn 2ϩ inhibition resulted from a loss of Zn 2ϩ -mediated potentiation. The W170S mutation on the background of H107N, which was previously reported to selectively disrupt Zn 2ϩ inhibition, showed remarkable attenuation of Zn 2ϩ -mediated potentiation and thus indicated that W170 is an important residue for the Zn 2ϩ -mediated GlyR potentiation. Moreover, overexpressing the ␣1 W170S subunit in cultured rat neurons confirmed the results from heterologous expression. Together, our results reveal a new zinc potentiation site on ␣1 GlyRs and a strong link between Zn 2ϩ modulation and human disease.

Tonic zinc inhibits spontaneous firing in dorsal cochlear nucleus principal neurons by enhancing glycinergic neurotransmission

“…Moreover, recent studies have linked mutations that abolish zinc-mediated allosteric GlyR potentiation with human hyperekplexia (Zhou et al, 2013). We propose that zinc mediated allosteric enhancement of GlyRs may be involved in tinnitus.…”

Physiological properties of Glycinergic synapses with defined subunit compositions

“…Physiological levels of zinc potentiate GlyRs (38,39), and the loss of zinc potentiation via the a1 D80A mutation accelerated the IPSC decay rate and thus caused hyperekplexia in a knock-in mouse model (32). Thus, it was reasonable to hypothesise that the W170S-mediated hyperekplexia phenotype was due to a diminished capacity of IPSCs to transfer chloride ions (19). However, in the present study we found that the decay time constant of IPSCs mediated by a1…”

“…The Y128C mutation was found to induce a sufficiently large spontaneous leak current as to degrade HEK293 cell viability (17) and thus we did not study it further. The V280M mutation significantly reduced both the glycine EC 50 and the I max values, although the W170S, Q226E and R414H mutations had little effect of these parameters (16,19). The Q226E, V280M and R414H mutations have previously been shown to induce spontaneous single channel activity (16,24).…”

“…b GlyR showed that zinc potentiation was abolished (19). Physiological levels of zinc potentiate GlyRs (38,39), and the loss of zinc potentiation via the a1 D80A mutation accelerated the IPSC decay rate and thus caused hyperekplexia in a knock-in mouse model (32).…”

Excessive Startle with Novel GLRA1 Mutations in 4 Chinese Patients and a Literature Review of GLRA1-Related Hyperekplexia

“…31 Studies showed that the mutant residues which is to be the binding sites for Zn 2+ may involve in the loss of zinc potentiation. 27,32 Except for the loss-of-function mechanism above, some gain-of-function mutations can also cause HPX. To date, four dominant mutations (Y156C, Q254E, V308M, R442H) have been demonstrated to induce spontaneous GlyR activation, which result in enhanced sodium and calcium influx rates, and direct-…”