Identification and characterization of the VAX2 p.Leu139Arg variant: possible involvement of VAX2 in cone dystrophy

<b>Objective</b>: This study was undertaken with the objective to investigate the potential involvement of VAX2 in retinal degeneration. <b>Methods</b>: A cohort of macular and cone dystrophy patients (<i>n</i> = 70) was screened for variant identification. Polymerase chain reaction (PCR) products were purified using ExoSAP-IT. Direct sequencing of PCR products was performed using BigDye 3.1 on the ABI 3730 DNA Analyzer and analyzed using DNASTAR software tool. Search for known variant was performed using the following platforms: 1000 Genomes Project, Ensembl, UCSC, ExAc, and dbSNP. The <i>VAX2</i> mutants were generated using the GeneArt® Site-Directed Mutagenesis kit. <i>In vitro</i> analysis was performed in hTERTRPE-1 (RPE-1) cell line. Cells were photographed using a Zeiss AXIOVERT S100 microscope. Images were analyzed using Photoshop CS4 software. <b>Results</b>: Here, we report the identification of a heterozygous non-synonymous variant (c.416T&gt;G; p.Leu139Arg) in one cone dystrophy proband. Functional characterization of this variant <i>in vitro</i> revealed an aberrant phenotype seen as protein mislocalization to cytoplasm/nucleus and aggregates undergoing degradation or forming aggresomes. The cellular phenotype suggests protein loss-of-function. Analysis of the <i>VAX2</i> p.Leu139Met, a variant present in the normal population, showed a phenotype similar to the wild-type, further supporting the hypothesis for the Leucine 139 to Arginine change to be damaging. <b>Conclusions</b>: This study raises the interesting possibility for evaluating <i>VAX2</i> as a candidate gene for cone dystrophy.