Mitochondrial cysteinyl-tRNA synthetase 2 protects against excitotoxic retinal cell death via enhanced supersulfide production

Mitochondrial cysteinyl-transfer RNA synthetase 2 (CARS2) is involved not only in the ligation of cysteine to transfer RNA but also in the synthesis of intracellular supersulfides. In this study, we investigated the role of CARS2 in the survival of retinal ganglion cells (RGCs) under excitotoxic conditions. Immunohistochemical analysis showed strong expression of CARS2 in RBPMS-positive RGCs in the mouse retina. Overexpression of exogenous human CARS2 (hCARS2) in mouse retinas and in the rat-derived retinal cell line R28 did not affect endogenous CARS2 mRNA levels. Adeno-associated virus 2-mediated overexpression of hCARS2 in RGCs significantly reduced cell death induced by excitotoxicity following intravitreal injection of N-methyl-D-aspartate. Similarly, hCARS2 overexpression decreased glutamate-induced excitotoxic cell death in R28 cells. Quantitative reverse transcription polymerase chain reaction analysis demonstrated a significant increase in CARS2 expression in R28 cells treated with glutamate. Using specific probes, we found that hCARS2-overexpressing R28 cells treated with glutamate exhibited higher intracellular levels of sulfane sulfur species and lower levels of reactive oxygen species (ROS) than control cells with basal CARS2 expression. Moreover, the oxidative stress marker gene Hmox1 was significantly downregulated in CARS2-overexpressing R28 cells compared with control cells. Taken together, these findings suggest that CARS2 plays a critical role in protecting retinal cells from excitotoxic cell death by increasing sulfane sulfur production and decreasing ROS accumulation. Given that CARS2 is predominantly expressed in RGCs among retinal cells, it may serve as a preemptive defense mechanism that enhances antioxidative activity at basal expression levels to support RGC survival. • CARS2 protein was strongly localized in retinal ganglion cells (RGCs). • Overexpression of CARS2 attenuated RGC death in an experimental model of excitotoxic injury. • In the retina-derived R28 cell line, elevated CARS2 expression reduced cell death following glutamate exposure. • Increased CARS2 expression scavenged glutamate-induced reactive oxygen species in R28 cells. • The expression level of an antioxidative stress marker gene was decreased in glutamate-treated R28 cells with enhanced CARS2 expression.