DJ-1-dependent regulation of oxidative stress in the retinal pigment epithelium (RPE)
Authors: Vera L Bonilha
Karen G Shadrach
Mary E Rayborn
Joe G Hollyfield
Journal: PLoS ONE (2013)
Discipline: Multidisciplinary
Keywords: DJ-1; oxidative stress; RPE cells; ROS generation; AMD
Affiliations: Department of Ophthalmic Research, The Cole Eye Institute, Cleveland Clinic Lerner College of Medicine, Cleveland, Ohio, United States of America.

Abstract:  Background: DJ-1 is found in many tissues, including the brain, where it has been extensively studied due to its association with Parkinson's disease. DJ-1 functions as a redox-sensitive molecular chaperone and transcription regulator that robustly protects cells from oxidative stress. Methodology: Retinal pigment epithelial (RPE) cultures were treated with H2O2 for various times followed by biochemical and immunohistological analysis. Cells were transfected with adenoviruses carrying the full-length human DJ-1 cDNA and a mutant construct, which has the cysteine residues at amino acid 46, 53 and 106 mutated to serine (C to S) prior to stress experiments. DJ-1 localization, levels of expression and reactive oxygen species (ROS) generation were also analyzed in cells expressing exogenous DJ-1 under baseline and oxidative stress conditions. The presence of DJ-1 and oxidized DJ-1 was evaluated in human RPE total lysates. The distribution of DJ-1 was assessed in AMD and non-AMD cryosectionss and in isolated human Bruch's membrane (BM)/choroid from AMD eyes. Principal findings: DJ-1 in RPE cells under baseline conditions, displays a diffuse cytoplasmic and nuclear staining. After oxidative challenge, more DJ-1 was associated with mitochondria. Increasing concentrations of H2O2 resulted in a dose-dependent increase in DJ-1. Overexpression of DJ-1 but not the C to S mutant prior to exposure to oxidative stress led to significant decrease in the generation of ROS. DJ-1 and oxDJ-1 intensity of immunoreactivity was significantly higher in the RPE lysates from AMD eyes. More DJ-1 was localized to RPE cells from AMD donors with geographic atrophy and DJ-1 was also present in isolated human BM/choroid from AMD eyes. Conclusions/significance: DJ-1 regulates RPE responses to oxidative stress. Most importantly, increased DJ-1 expression prior to oxidative stress leads to decreased generation of ROS, which will be relevant for future studies of AMD since oxidative stress is a known factor affecting this disease. 

Retracted: No

PubPeer comments:
 This article has two corrections. * July 2014 correction republication because Figure 2 was left out of the paper: https: journals.plos.org plosone article?id=10.1371 journal.pone.0104292 * October 2017 correction, found here: https: journals.plos.org plosone article?id=10.1371 journal.pone.0185834  *The authors would like to correct Figs 2 and 4, as errors were introduced in the preparation of these figures for publication. The GAPDH control load in Fig 2A (D407 +H2O2 1h) had been duplicated by mistake in Fig 2B (ARPE-19 + H2O2 18hr). These same ARPE-19 +H2O2 blots were stripped and re-probed in Fig 4 with the oxDJ-1 antibody. The authors checked the original images obtained and the GAPDH panels in Figs 2 and 4 were corrected. The correct versions of Figs 2 and 4 are provided here, along with their corrected figure legends. The authors confirm that these changes do not alter their findings. The authors have provided the underlying images as Supporting Information.* Duplications of GAPDH panels as reported to journal in 2015 (2 years before correction):  I thank the authors for correcting this duplication. Image uploaded here for Image Forensics training purposes. ,