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Martial Mbefo, Yvan Arsenijevic; Epigenetic modifications associated with photoreceptor cell death in retinopathies.. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):3637.
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© ARVO (1962-2015); The Authors (2016-present)
Retinitis Pigmentosa (RP) is a form of aggressive and inherited visual disorder resulting from a progressive and selective degeneration of photoreceptors cells in the retinal outer layer. However, the exact molecular mechanisms mediating the photoreceptors cell death remain elusive. In order to better understanding the chronology of the cascade of events controlling the photoreceptor cell loss in retinopathies, we have recently demonstrated that the cell death in various models of retinal degeneration is mediated by key regulators of the cell cycle such as CDK4, E2F1 and the Polycomb repressive complex 1 (PrC1) component BMIi-1. Genetic suppression of Bmi-1 generates the most powerful protection never obtained so far, and demonstrates more than 70% rescue of photoreceptor cells in KO animal as compared to that of WT. Here we investigated whether other Polycomb partners preceeding Bmi-1 action are also involved in the cell death process
Freshly extracted retinal tissues obtained from FVB-Crx GFP RD1 mice were immediately processed in calpain buffer. Following dissociation, photoreceptors were purified by cell sorting and the crx-GFP positive cells were snap frozen at -80°C until the day of analysis. For Biochemistry, the samples were homogenized in lysis buffer, follow by co-immunoprecipitation before the immunoblotting. In parallel, paraformaldehyde-fixed sections were immunostained against various antigens, mounted and analyzed by confocal microscopy.
Photoreceptors cells purified from the RD1 mouse retina demonstrated a significant epigenetic change as compared to the WT mice. Immunohistochemical analysis of frozen tissues sections showed a dramatic hypertrimethylation of histone 3 Lys27 (H3K27me3) in cells expressing an early marker of photoreceptor cell death, and an up-regulation of EZH2 in RD1 but not in WT photoreceptors. Interestingly however, such changes in H3K27me3 mark were progressively lost during the late phase of the disease. Furthermore, the changes associated with H3K27me3 levels predominantly occur at disease onset and correlate with the retinal degeneration kinetic previously reported with the cell cycle proteins markers in our animal model.
All together, these findings suggest that epigenetic mark reorganization may be involved in the process of photoreceptor cell death by controlling the reactivation of cell cycle proteins.
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