Identification of a mutation in ASRGL1 as the underlying cause of early-onset recessive Retinal Degeneration (RD).

Abstract

Purpose: To understand the molecular pathology underlying early-onset retinal degeneration in a large consanguineous pedigree.

Methods: A thorough ophthalmic evaluation including measurement of visual acuity, visual fields, electroretinography (ERGs), and fundus photography was carried out. Blood samples were collected from 6 affected and 7 unaffected family members and genomic DNA was isolated from white blood cells. Linkage and haplotype analyses were performed using microsatellite markers. Exome capture was preformed using Agilent V5+UTRs kit and sequenced on an Illumina HiSeq 2000 genome analyzer. Sequence reads were mapped against the hg19 version of the human genome and analyzed using Genome Analysis Toolkit software package. Exome variants were analyzed using exomeSuite and verified by segregation analysis. The expression, and sub-cellular localization of the wild type and mutant ASRGL1 proteins were investigated by RT-PCR, immunohistochemistry and immunocytochemistry. ASRGL1 was expressed in E.Coli and COS-7 cells to examine the enzymatic activity of the wild type and mutant protein.

Results: The ophthalmic examination was suggestive of non-syndromic early onset retinal degeneration. Linkage and haplotype analyses mapped the disease locus to chromosome 11. Exome sequencing and segregation analysis identified a novel missense change resulting in p.G178R alteration in the Asaparaginase like 1 (ASRGL1) gene as the underlying cause of RD. In vitro analysis revealed that the mutation impairs the autocatalytic activity of ASRGL1. In addition, the unprocessed protein forms high molecular weight aggregates. In sharp contrast to the localization pattern of wild type ASRGL1 that distributes through the cell, mutant ASRGL1 shows perinuclear localization and re-organization of cytoskeleton in COS-7 cells. Immunohistochemistry localized the ASRGL1 protein to photoreceptor layer in the retina of adult mice.

Conclusions: The Linkage-coupled exome sequencing implicates the photoreceptor expressed gene ASRGL1 in retinal degeneration. We present evidence that the G178R allele has impaired autocatalytic activity, which results in the loss of active ASRGL1. In addition the mutant ASRGL1 forms aggresomes, that may lead to photoreceptor degeneration.