Mutations in GNAL: a novel cause of craniocervical dystonia

Abstract

Importance: Mutations in the GNAL gene have recently been shown to cause primary torsion dystonia. The GNAL-encoded protein (Gαolf) is important for dopamine D1 receptor function and odorant signal transduction. We sequenced all 12 exons of GNAL in 461 patients from Germany, Serbia, and Japan, including 318 patients with dystonia (190 with cervical dystonia), 51 with hyposmia and Parkinson disease, and 92 with tardive dyskinesia or acute dystonic reactions.

Observations: We identified the following two novel heterozygous putative mutations in GNAL: p.Gly213Ser in a German patient and p.Ala353Thr in a Japanese patient. These variants were predicted to be pathogenic in silico, were absent in ethnically matched control individuals, and impaired Gαolf coupling to D1 receptors in a bioluminescence energy transfer (BRET) assay. Two additional variants appeared to be benign because they behaved like wild-type samples in the BRET assay (p.Ala311Thr) or were detected in ethnically matched controls (p.Thr92Ala). Both patients with likely pathogenic mutations had craniocervical dystonia with onset in the fifth decade of life. No pathogenic mutations were detected in the patients with hyposmia and Parkinson disease, tardive dyskinesias, or acute dystonic reactions.

Conclusions and relevance: Mutations in GNAL can cause craniocervical dystonia in different ethnicities. The BRET assay may be a useful tool to support the pathogenicity of identified variants in the GNAL gene.

Figure 1. Schematic Representation of Mutations in the GNAL Gene

Tan boxes indicate previously reported mutations^,; gray boxes, putatively pathogenic mutations detected in this study. Electropherograms show the wild-type sequence (above), the mutation (middle) at DNA level, and the complementary DNA (cDNA) sequence (below). Forward and reverse cDNA strands are shown for individuals L4486 and DYT_family7, indicating equal expression of the wild-type and mutant alleles (red boxes). The cross-species sequence alignment of stimulatory α subunit Gα_olf obtained from Mutation Taster software (http://www.mutationtaster.org) is also shown.

Figure 2. Functional Effects of GNAL Mutations in a Cell-Based Bioluminescence Energy Transfer (BRET) Assay

A, Time course of changes in BRET signal (R) after stimulation of cells expressing dopamine D₁ receptor with dopamine and subsequent deactivation by haloperidol. B, Change in the BRET ratio from basal (R₀) to maximal response (R_max), reflecting the extent of the stimulatory α subunit Gα_olf activation. C, Basal BRET ratios calculated before the application of dopamine, reflecting the extent of Gα_olf association with the G_βγ subunits. Two of the mutations (p.Gly213Ser and p.Ala353Thr) had greatly diminished amplitudes of the BRET response after dopamine application (shown in parts A and B), with responses virtually indistinguishable from the random baseline fluctuations seen in the absence of Gα_olf, suggesting that these mutations lead to a complete loss of Gα_olf function. In contrast, the p.Ala311Thr variant performed similarly to the wild type. Error bars indicate SEM values. One-way analysis of variance followed by the Holm-Sidak method was performed to determine statistically significant differences relative to wild-type control. ^aP < .001.