A loss-of-function variant in canine GLRA1 associates with a neurological disorder resembling human hyperekplexia

Abstract

Hereditary hyperekplexia is a rare neuronal disorder characterized by an exaggerated startle response to sudden tactile or acoustic stimuli. In this study, we present a Miniature Australian Shepherd family showing clinical signs, which have genetic and phenotypic similarities with human hereditary hyperekplexia: episodes of muscle stiffness that could occasionally be triggered by acoustic stimuli. Whole genome sequence data analysis of two affected dogs revealed a 36-bp deletion spanning the exon-intron boundary in the glycine receptor alpha 1 (GLRA1) gene. Further validation in pedigree samples and an additional cohort of 127 Miniature Australian Shepherds, 45 Miniature American Shepherds and 74 Australian Shepherds demonstrated complete segregation of the variant with the disease, according to an autosomal recessive inheritance pattern. The protein encoded by GLRA1 is a subunit of the glycine receptor, which mediates postsynaptic inhibition in the brain stem and spinal cord. The canine GLRA1 deletion is located in the signal peptide and is predicted to cause exon skipping and subsequent premature stop codon resulting in a significant defect in glycine signaling. Variants in GLRA1 are known to cause hereditary hyperekplexia in humans; however, this is the first study to associate a variant in canine GLRA1 with the disorder, establishing a spontaneous large animal disease model for the human condition.

Fig. 1

Most muscle fibers of both muscles showed a high load of partially confluent dark blue lipid droplets in between muscle filaments (a: white frame) and within the subsarcolemmal perinuclear cytoplasm (a: bound by arrows). At the ultrastructural level (b, c), the droplets present as homogenous semi electron dense conglomerates, typical for neutral fat (b, c: asterisks). They are located close to mitochondria (a, b: arrowheads), many showing disruption of cristae architecture and trilaminar inclusions (b: white arrowheads). Staining/contrasting: a: toluidine blue–safranin O; b, c uranyl acetate–lead citrate. Magnification: see scale bars

Fig. 2

a The exon–intron structure of the GLRA1 gene. The 36-bp deletion (position marked with dashed line) encompasses part of the intron 1 and exon 2. b GLRA1 protein domains with the signal peptide (SP), N-terminus (N), four transmembrane domains (TM1–TM4) and C-terminus (C). The variant (highlighted with light blue) deletes a part encoding the signal peptide. The resulting protein is predicted to contain a part of the normal signal peptide followed by 19 abnormal amino acids (highlighted with red). c The structure of the glycine receptor alpha-1 subunit. d Heteromeric glycine receptors are formed by five subunits: three α subunits and two β subunits

Fig. 3

Sanger sequencing chromatograms for dogs homozygous for the reference allele (wt), affected individuals homozygous for the variant (hom), and heterozygous carriers (het). The deletion region is highlighted in gray in the reference allele in the uppermost image