Abstract
Normal function of the retinal pigment epithelium (RPE) is essential for maintaining the structural integrity of retinal photoreceptors and the visual process. Sustained oxidative damage of the RPE due to aging and other risk factors contributes to the development of age-related macular degeneration (AMD). The transcription factor NF-E2-related factor 2 (Nrf2) is a central regulator of cellular antioxidant and detoxification responses. Enhancing Nrf2 function protects RPE cells from oxidation-related apoptosis and cell death. Previously, we demonstrated that Nrf2 activation can be induced by endoplasmic reticulum (ER) stress; however, the mechanisms are not fully understood. In the present study, we examined the role of X box-binding protein 1 (XBP1), an ER stress-inducible transcription factor, in regulation of Nrf2 in the RPE. We found that RPE-specific XBP1 conditional knockout (cKO) mice exhibit a significant reduction in Nrf2 mRNA and protein levels, along with decreased expression of major Nrf2 target genes, in the RPE/choroid complex. Using primary RPE cells isolated from XBP1 cKO mice and human ARPE-19 cell line, we confirmed that loss of XBP1 gene or pharmacological inhibition of XBP1 splicing drastically reduces Nrf2 levels in the RPE. Conversely, overexpression of spliced XBP1 results in a modest but significant increase in cytosolic and nuclear Nrf2 protein levels without affecting the transcription of Nrf2 gene. Moreover, induction of ER stress by tunicamycin and thapsigargin markedly increases Nrf2 expression, which is abolished in cells pretreated with XBP1 splicing inhibitors 4μ8C and quinotrierixin. Mechanistic studies indicate that quinotrierixin reduces Nrf2 expression likely through inhibition of protein translation. Finally, we demonstrate that overexpression of Nrf2 protected RPE cells against oxidative injury but appeared to be insufficient to rescue from XBP1 deficiency-induced cell death. Taken together, our results indicate that XBP1 modulates Nrf2 activity in RPE cells and that XBP1 deficiency contributes to oxidative injury of the RPE.
Highlights
The retinal pigment epithelium (RPE), a monolayer of pigmented epithelial cells in the eye, plays a crucial role in the nourishment and detoxification of photoreceptors (Strauss, 2005)
We examined whether the induction of NF-E2-related factor 2 (Nrf2) by endoplasmic reticulum (ER) stress is affected in X box-binding protein 1 (XBP1)-deficient RPE
We found that Nrf2 expression was drastically decreased by 70% in XBP1 conditional knockout (cKO) RPE/choroid complex compared to the controls after tunicamycin treatment (Figure 1D)
Summary
The retinal pigment epithelium (RPE), a monolayer of pigmented epithelial cells in the eye, plays a crucial role in the nourishment and detoxification of photoreceptors (Strauss, 2005). To cope with the oxidative environment, the RPE has evolved effective defenses against oxidative stress It is rich in antioxidants such as vitamin E, as well as enzymes and proteins that neutralize free radicals, including superoxide dismutase (SOD), catalase, and glutathione (Shamsi and Boulton, 2001). Mice lacking Nrf develop drusen-like deposits, accumulation of lipofuscin, spontaneous choroidal neovascularization (CNV) and sub-RPE deposition of inflammatory proteins, which resemble human AMD (Zhao et al, 2011). These findings, along with compelling evidence from other published studies, support the importance of Nrf signaling in the anti-oxidant defense system of the RPE
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