Calpain Activation Is the Major Cause of Cell Death in Photoreceptors Expressing a Rhodopsin Misfolding Mutation
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The majority of mutations in rhodopsin (RHO) cause misfolding of the protein and has been linked to degeneration of photoreceptor cells in the retina. A lot of attention has been set on targeting ER stress for the development of new therapies for inherited retinal degeneration caused by mutations in the RHO gene. Nevertheless, the cell death pathway activated by RHO misfolded protein is still debated. In this study, we analyzed the retina of the knock-in mouse expressing the P23H misfolded mutant RHO. We found persistent unfolded protein response (UPR) during degeneration. Interestingly, long-term stimulation of the PERK branch of ER stress had a protective effect by phosphorylating nuclear factor erythroid 2–related factor 2 (NRF2) transcription factor, associated with antioxidant responses. Otherwise, we provide evidence that increased intracellular calcium and activation of calpains strongly correlated with rod photoreceptor cell death. By blocking calpain activity, we significantly decreased the activation of caspase-7 and apoptosis-inducing factor (AIF), two cell death effectors, and cell demise, and effectively protected the retina from degeneration caused by the P23H dominant mutation in RHO.
KeywordsadRP Rod eIF2α Spectrin PD150606 Z-VAD-FMK GSK2606414A
The authors acknowledge the Cell-lab Facility and CSSI of the University of Modena and Reggio Emilia for the cytofluorimetric analysis and animal husbandry assistance.
A.C. and D.S. performed experimental procedures and contributed to the writing of the manuscript. M.M. performed flow cytometry analysis. V. M designed and supervised the experiments and wrote the manuscript.
V.M. was supported by the research grant Fondazione Roma (call for proposals 2013 on Retinitis Pigmentosa), European Union (transMed, MSCA-ITN-2017-765441), and Fondazione Telethon (grant numbers GGP11210, GGP14180).
Compliance with Ethical Standards
All procedures on mice were conducted at CSSI (Centro Servizi Stabulario Interdipartimentale), approved by the Ethical Committee of University of Modena and Reggio Emilia and by the Italian Ministero della Salute (346/2015-PR), and were in accordance with the ARVO Statement for the Use of Animals in Ophthalmic and Vision Research.
Conflict of Interest
The authors declare that they have no conflict of interest.
Informed consent was obtained from all individual participants included in the study.
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