Homozygous deletion of glutamate receptor gene GRID2 causes new hotfoot mutant phenotype, characterized by early-onset cerebellar ataxia and retinal dystrophy

Abstract

Purpose: To identify the genetic cause of early-onset autosomal recessive cerebellar ataxia (ARCA) associated with retinal dystrophy in a consanguineous family.

Methods: An affected six-months old child underwent neurological and ophthalmological examinations. Following genetic analyses were conducted: homozygosity mapping, copy number analysis, conventional PCR, Sanger sequencing, qPCR, whole exome sequencing (WES). Expression analysis of GRID2 was performed by qPCR and immunohistochemistry, in murine and human retina.

Results: The child had truncal and appendicular ataxia, tonic upgaze and nystagmus. Magnetic resonance imaging showed cerebellar atrophy, particularly of the vermis region. Fundoscopy revealed no pigmentary abnormalities and electroretinography demonstrated reduced scotopic and photopic amplitudes, pointing to early-onset retinal dystrophy. A homozygous deletion of exon 2 of GRID2 (p.Gly30_Glu81del) was found in the proband, compatible with mouse hotfoot mutant ho15J. GRID2 encodes an ionotropic glutamate receptor known to be selectively expressed in cerebellar Purkinje cells. Here, we demonstrated GRID2 mRNA expression in human adult retina and retinal pigment epithelium. In addition, Grid2 expression was demonstrated in different stages of murine retinal development. GRID2 protein expression was observed in both murine and human retina. More specifically, GRID2 localized to photoreceptor inner segments, the outer plexiform layer and ganglion cell layer in human and mouse. Faint Grid2 immunoreactivity was also observed at the inner nuclear layer/inner plexiform layer margin in the mouse retina. In order to rule out involvement of mutations in another gene, WES was conducted but did not reveal any other disease-causing mutations, supporting the phenotype observed here represents a single clinical entity.

Conclusions: We identified GRID2 as underlying disease gene of early-onset ARCA with retinal dystrophy, expanding the clinical spectrum of GRID2 hotfoot deletion mutants in humans. We demonstrated, for the first time, GRID2 mRNA and protein expression in human and murine retina, providing evidence for a novel functional role of GRID2 in the retina. Finally, to the best of our knowledge GRID2 is the second glutamate receptor gene, apart from GRM6, leading to retinal disease when mutated.

Keywords: 539 genetics • 604 mutations • 688 retina