Genetic deletion of S-opsin prevents rapid cone photoreceptor degeneration in the Guanylate Cyclase-1 knockout model of Leber congenital amaurosis

摘要

Purpose : Retinal guanylate cyclase-1 (GC1), an outer segment protein expressed in both rods and cones, is responsible for synthesis of cyclic GMP (cGMP). Mutations in GC1 are associated with severe, early-onset Leber congenital amaurosis-1(LCA1). Previous studies have shown that both M-and S-opsins are mislocalized in Lrata??/a??, a murine model for LCA1. In addition, ventral and central cones degenerate more rapidly than dorsal cones in GC1-/- mouse. We have shown previously that S-opsin is more prone to aggregation than M-opsin and is likely to cause the rapid ventral/central cone degeneration. The purpose of this study was to investigate the role of S-opsin in rapid cone degeneration of GC1-/- by genetically deleting S-opsin. Methods : We bred GC1a??/a?? mice with S-opsina??/a?? mice to generate GC1a??/a??S-opsina??/a?? mice. We compared cone density and morphology between GC1a??/a??, GC1a??/a??S-opsina??/a??, and WT mouse lines by histology. We also evaluated expression and subcellular localization of cone-specific proteins, including M- and S-opsin, by Western blotting and immunohistochemistry, respectively. Results : Whereas more than 70% and 50% of cones in the central and ventral regions of GC1a??/a?? retina, respectively, degenerated within eight weeks (wks), deletion of S-opsin (GC1a??/a??S-opsina??/a??) maintained central cone density at WT level for at least 10 wks. Ventral cone density was maintained above 80% of WT level for at least 10 wks. To determine whether mistrafficked cone opsins were differentially processed, as in Lrata??/a??, we conducted Western blots at three stages of cone degeneration: (i) 2 wks postnatal, predegeneration; (ii) 5 wks, early degeneration; and (iii) 6 wks, ongoing degeneration. At 2 wks, protein levels of both M-opsin and S-opsin were similar to WT. However, at 5 and 6 wks, M-opsin protein level was markedly reduced while S-opsin level remained the same compared with WT. Assuming that protein synthesis for cone opsins is minimally affected in early GC1a??/a?? cone degeneration, our results suggest that mislocalized S-opsin was more resistant to proteasome degradation than mislocalized M-opsin. Conclusions : Our results suggest that S-opsin accumulation is responsible for rapid cone degeneration in central and ventral retina of GC1a??/a?? mice. This could be a general mechanism for rapid cone degeneration in a number of animal models with cone opsin mislocalization. This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.