Circadian Gene Expression Profiles in Rat Retina: What are the Crystallins Doing?

Abstract

Abstract: : Purpose: To better understand the endogenous factors related to circadian-dependant retinal light damage we determined gene expression profiles in rat retina at various times of the day and night. Methods: Weanling male Sprague-Dawley rats were reared in either dim cyclic light or darkness. At P60 all rats were dark adapted and retinas excised from 3 separate groups of 3 rats each at 0100, 0900 and 1700. mRNA was extracted and gene expression profiles determined using Affymetrix rat genome U34A Genechips®. For selected genes their expression profiles were confirmed by real time PCR and the presence of proteins determined by 2D gel electrophoresis and Western analysis. Results: The expression of a number of retinal genes changed in a time dependent fashion in both cyclic light and dark reared rats. Among these, all classes of crystallin mRNA were expressed. with the lowest levels at 0100. By real time PCR, α and ß crystallin expression was from 2-12 fold greater at 0900 and 1700. Gamma crystallin gene expression was much greater at 0900 in cyclic reared rats, but higher in dark reared animals at 1700. By Western analysis of 2D gel protein profiles we detected the presence of multiple immunoreactive forms of α, ßand γ- crystallins. Conclusions: Retinal crystallin gene expression profiles change in a manner consistent with circadian dependent light damage susceptibility. Multiple forms of the proteins are also present, suggesting that time dependent post translational modifications occur. Retinal cystallins appear to be among the endogenous factors that influence light induced retinal degeneration.

Keywords: circadian rhythms • crystallins • gene microarray