کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
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4011802 | 1261165 | 2010 | 9 صفحه PDF | دانلود رایگان |
![عکس صفحه اول مقاله: S-opsin protein is incompletely modified during N-glycan processing in Rpe65−/− mice S-opsin protein is incompletely modified during N-glycan processing in Rpe65−/− mice](/preview/png/4011802.png)
Retinal pigment epithelium-specific protein 65 kDa (RPE65) is a key enzyme for the visual cycle in the eye. Rpe65−/− mice lack 11-cis-retinal, and show early cone degeneration and mislocalization of cone opsins. The present study investigated whether abnormal modification of cone opsins at the protein level is present in Rpe65−/− mice. Retina–RPE–choroids of Rpe65−/− mice at 3, 5 and 7 weeks old were used. Immunohistochemistry of opsins was performed using cryosections and retinal flatmounts. We evaluated levels of mRNA for cone and rod opsin genes by RT-PCR and levels of proteins by western blotting. To examine modification patterns of N-glycan in Rpe65−/− mice, cone opsins were digested with peptide-N-glycosidase (PNGase) F. S-opsin protein was detected at ∼40-kDa as a major band in wild-type mice, whereas ∼42-kDa S-opsin protein was detected in Rpe65−/− mice. After PNGase F treatment, mobility of S-opsin protein in wild-type and Rpe65−/− mice on SDS-PAGE was similar. In addition, ∼25-kDa S-opsin polypeptide was notably detected in Rpe65−/− mice. Conversely, M-opsin proteins were not observed by immunohistochemistry or western blotting in Rpe65−/− mice, but expression of M-opsin mRNA in Rpe65−/− mice did not differ significantly from that in wild-type mice at 3 and 5 weeks. Mobility of M-opsin protein in Rpe65−/− mice was unchanged. Our data suggest that S-opsin protein is incompletely modified during N-glycan processing in Rpe65−/− mice, whereas M-opsin protein is severely reduced by posttranslational degradation in the absence of incomplete N-glycan processing in Rpe65−/− mice.
Journal: Experimental Eye Research - Volume 91, Issue 1, July 2010, Pages 54–62