Differential Expression of Cope During Light–Induced Retinal Damage in Rats

Abstract

Purpose: : We have identified coatomer protein complex subunit epsilon (COPE) from a differential cross screen of degenerative retina in a rat animal model and further characterized its expression in the degenerative profile and in human retina and RPE using quantitative RT–PCR. The COPE gene encodes the epsilon subunit of the coatomer protein complex which is involved in retrograde Golgi to ER transport of dilysine–tagged proteins.

Methods: : We use a green light–induced retinal degeneration (LIRD) model in dark reared Sprague Dawley rats to synchronize light and oxidative damage induced stress in the retina. Differential cross screens using cDNA probes derived from RNA from untreated and light treated rat retinae were performed on cDNA libraries representative of light treated rat retinae. Macroarrays were used as a tertiary screening method. One clone representing an interesting expression profile was further characterized by DNA sequencing, bioinformatic analyses and quantitative RT–PCR.

Results: : We identified a cDNA clone that matched the predicted rat COPE gene. Bioinformatic analyses revealed that the rat COPE mRNA sequence is similar to mouse and human COPE sequences. At the genomic level, COPE gene structure is maintained and conserved from human through fugu and zebrafish. Virtual northerns and published array data suggest that COPE is expressed at high levels in many tissues including a variety of ganglion cell types. Quantitative RT–PCR confirmed rat retinal expression of COPE over a time course of light exposures to rats. There is an initial decrease in COPE mRNA levels through 8 hours of light exposure with levels returning to unexposed levels at 16 hours. Further quantitative RT–PCR analysis demonstrated expression of COPE in human retina and fovea.

Conclusions: : The coatomer complex may be involved in recycling of ER derived vesicles during the production of rhodopsin and the reduction of rhodopsin production during LIRD might decrease the need for coatomer complexes in photoreceptor cells. However, rhodopsin mRNA levels are low at 16 hours of light exposure while COPE expression increases from 8 hours suggesting another role for COPE. It is thus important to determine which cells in the retina express COPE during LIRD. We are now using immunohistochemistry to identify which retinal cells express high levels of coatomer complex proteins.

Keywords: gene/expression • stress response • retinal degenerations: cell biology