Molecular identification of circadian clock proteins in the rat lens: evidence of circadian regulation of glutathione homeostasis?

Abstract

Purpose : To determine whether the lens acts as a circadian clock that controls the rhythmic expression of genes involved in glutathione (GSH) redox regulation

Methods : Western blotting and immunohistochemistry were used to confirm the expression of circadian clock core proteins (BMAL1 & CLOCK) in rat lenses. Reverse transcription-quantitative PCR (RT- qPCR) was performed on RNA extracted from rat lenses collected every 4 hours over a 24-hour period to determine if there were cyclic differences in the expression of clock component genes (Bmal1, Clock, Per and Cry) and GSH related redox genes involved in GSH synthesis and regeneration.

Results : Western blotting for circadian clock proteins from lens homogenates revealed bands consistent with the predicted molecular weights for BMAL1 and CLOCK. Immunohistochemistry revealed labelling for BMAL1 and CLOCK in the epithelium and fibre cells in the outer cortex. RT-qPCR revealed fluctuations for both the clock component genes and redox genes over a 24-hour period suggestive that the lens may utilize circadian time keeping mechanisms to regulate GSH homeostasis

Conclusions : Collectively, these findings reveal that the lens contains the core molecular components of the circadian clock, and that fluctuations in the expression of clock component genes and GSH related genes occurs during the day/night cycle. This implies that the lens may act as a circadian clock that controls the rhythmic expression of genes involved in GSH homeostasis to provide protection against UV insults. Since circadian rhythms are altered with advancing age, this implies that disruption of the circadian regulation of redox balance may be an initiating factor in age related lens cataract.

This is a 2021 ARVO Annual Meeting abstract.