Intense Light– and Time–Dependent Changes in Rat Rod Outer Segment Crystallins

Abstract

Abstract: : Purpose:To determine the presence and expression levels of crystallins in rat retinal ROS samples at various times of the day and night and in response to intense visible light. Methods: Weanling male Sprague–Dawley rats were reared in either dim cyclic light or in darkness. At P 60, animals were dark–adapted for 16 hours and then either exposed to intense visible light at 0100, 0900 or 1700 or not. Light treatments were for 0.5, 3 or 8 hour durations. ROS were isolated from 8 light–exposed or unexposed animals for each time point. ROS proteins were visualized on 2D–gels, and α, ß and γ crystallins were identified by western analysis with individual or class specific antibodies. Individual crystallins and other ROS proteins were excised from gels, digested in situ with trypsin and identified by capillary LC MS/MS. Results: In unexposed rat ROS, αA and αB crystallins were readily detected at 0100, 0900 and 1700, while ß and γ crystallins were less prevalent. Upon exposure to intense light, changes in ROS crystallins were found to depend on the duration of exposure and time of day of light treatment. Mass spectrometric analyses of excised gel spots have revealed a partial 2D proteomic footprint of rat ROS. Conclusions: Crystallins are present in rat ROS and exhibit expression patterns which vary by time of day and intense light exposure. Our data suggests that intense light–induced photoreceptor cell degeneration leads to changes in ROS crystallin proteins and/or post–translational processing which appears to correlate with the duration of light exposure. CR: None. Support: NIH EY–01959 & M. Petticrew, Springfield, OH (DTO); NIH EY06603 and EY014239 (JWC).

Keywords: crystallins • photoreceptors • proteomics