CLOCK and TIMELESS regulate rhythmic occupancy of the BRAHMA chromatin-remodeling protein at clock gene promoters

Circadian clock and chromatin-remodeling complexes are tightly intertwined systems that regulate rhythmic gene expression. The circadian clock promotes rhythmic expression, timely recruitment, and/or activation of chromatin remodelers, while chromatin remodelers regulate accessibility of clock transcription factors to the DNA to influence expression of clock genes. We previously reported that the BRAHMA (BRM) chromatin-remodeling complex promotes the repression of circadian gene expression in Drosophila. In this study, we investigated the mechanisms by which the circadian clock feeds back to modulate daily BRM activity. Using chromatin immunoprecipitation, we observed rhythmic BRM binding to clock gene promoters despite constitutive BRM protein expression, suggesting that factors other than protein abundance are responsible for rhythmic BRM occupancy at clock-controlled loci. Since we previously reported that BRM interacts with two key clock proteins, CLOCK (CLK) and TIMELESS (TIM), we examined their effect on BRM occupancy to the period (per) promoter. We observed reduced BRM binding to the DNA in clk null flies, suggesting that CLK is involved in enhancing BRM occupancy to initiate transcriptional repression at the conclusion of the activation phase. Additionally, we observed reduced BRM binding to the per promoter in flies overexpressing TIM, suggesting that TIM promotes BRM removal from DNA. These conclusions are further supported by elevated BRM binding to the per promoter in flies subjected to constant light and experiments in Drosophila tissue culture in which the levels of CLK and TIM are manipulated. In summary, this study provides new insights into the reciprocal regulation between the circadian clock and the BRM chromatin-remodeling complex.