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Understanding the role of chromatin remodeling in the regulation of circadian transcription in Drosophila.

Kwok RS, Lam VH, Chiu JC - Fly (Austin) (2015)

Bottom Line: The robust cycling of circadian gene expression is critical for proper timekeeping, and is regulated by transcription factor binding, RNA polymerase II (RNAPII) recruitment and elongation, and post-transcriptional mechanisms.Here, we discuss the characterization of the BRAHMA (BRM) chromatin-remodeling protein in Drosophila in the context of circadian clock regulation.Finally, we divulge ongoing efforts and investigative directions toward a deeper mechanistic understanding of transcriptional regulation of circadian gene expression at the chromatin level.

View Article: PubMed Central - PubMed

Affiliation: a Department of Entomology and Nematology ; University of California Davis ; Davis , CA 95616 , USA.

ABSTRACT
Circadian clocks enable organisms to anticipate daily changes in the environment and coordinate temporal rhythms in physiology and behavior with the 24-h day-night cycle. The robust cycling of circadian gene expression is critical for proper timekeeping, and is regulated by transcription factor binding, RNA polymerase II (RNAPII) recruitment and elongation, and post-transcriptional mechanisms. Recently, it has become clear that dynamic alterations in chromatin landscape at the level of histone posttranslational modification and nucleosome density facilitate rhythms in transcription factor recruitment and RNAPII activity, and are essential for progression through activating and repressive phases of circadian transcription. Here, we discuss the characterization of the BRAHMA (BRM) chromatin-remodeling protein in Drosophila in the context of circadian clock regulation. By dissecting its catalytic vs. non-catalytic activities, we propose a model in which the non-catalytic activity of BRM functions to recruit repressive factors to limit the transcriptional output of CLOCK (CLK) during the active phase of circadian transcription, while the primary function of the ATP-dependent catalytic activity is to tune and prevent over-recruitment of negative regulators by increasing nucleosome density. Finally, we divulge ongoing efforts and investigative directions toward a deeper mechanistic understanding of transcriptional regulation of circadian gene expression at the chromatin level.

No MeSH data available.


Related in: MedlinePlus

The effects of two classes of brm mutants on CLK-activated transcription. (A) Knockdown of brm via RNAi results in the reduction of both catalytic and non-catalytic functions. The decrease in catalytic activity leads to lower nucleosome density as compared to control, and decrease in non-catalytic function impairs recruitment of repressive complexes. As a result, CLK-activated gene expression is elevated. (B) Expression of catalytically-inactive BRMK804R protein results in decreased nucleosome density. The more open chromatin allows for an augmented effect of the non-catalytic function retained by BRMK804R, which may include recruitment of repressors. Experimental data suggests that interactors of BRM have a negative effect on CLK binding to the per promoter, and per expression is downregulated as a result of decreased CLK binding.36 (C) Wild type BRM possesses both non-catalytic and catalytic functions that balance each other and serve to fine-tune CLK-activated transcription. Non-catalytic function of BRM may promote recruitment of proteins that have repressive effects on transcription. The over-recruitment of repressive factors is prevented by the catalytic function of BRM to maintain nucleosome density.
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f0002: The effects of two classes of brm mutants on CLK-activated transcription. (A) Knockdown of brm via RNAi results in the reduction of both catalytic and non-catalytic functions. The decrease in catalytic activity leads to lower nucleosome density as compared to control, and decrease in non-catalytic function impairs recruitment of repressive complexes. As a result, CLK-activated gene expression is elevated. (B) Expression of catalytically-inactive BRMK804R protein results in decreased nucleosome density. The more open chromatin allows for an augmented effect of the non-catalytic function retained by BRMK804R, which may include recruitment of repressors. Experimental data suggests that interactors of BRM have a negative effect on CLK binding to the per promoter, and per expression is downregulated as a result of decreased CLK binding.36 (C) Wild type BRM possesses both non-catalytic and catalytic functions that balance each other and serve to fine-tune CLK-activated transcription. Non-catalytic function of BRM may promote recruitment of proteins that have repressive effects on transcription. The over-recruitment of repressive factors is prevented by the catalytic function of BRM to maintain nucleosome density.

Mentions: The most surprising finding of our study was perhaps the fact that the two classes of brm mutants we examined exhibited opposing effects on circadian gene expression, with brm RNAi displaying increased clock gene expression compared to control and brmK804R showing downregulation of clock gene expression (Fig. 1D). Since both classes of brm mutants were predicted to show a decrease in catalytic activity and exhibited a reduction in nucleosome density at per and tim loci as assayed by histone H3 ChIP-qPCR, our results suggested that the BRM complex normally functions to condense the chromatin at clock gene loci. Moreover, our results strongly suggested that it is likely the intact non-catalytic activity of BRMK804R that led to the differential effects exhibited by the two classes of brm mutants with respect to clock gene expression. We postulate that when brm is knocked down via RNAi, the chromatin is more open due to a loss of nucleosome remodeling catalytic activity, allowing for an aberrant elevation in gene expression that results partly from an increased recruitment of general transcription machinery (Fig. 2A). While this explains the increase in clock gene expression and apparent decrease in RNAPII stalling in flies expressing brm RNAi, the opposite effect in clock gene expression observed in flies expressing the catalytically-inactive brmK804R remains puzzling, especially since the chromatin is also more relaxed in brmK804R flies as compared to wild type (Fig. 2B). Fortunately, investigation of changes in CLK binding to the per and tim promoters promptly provided us with valuable clues to indicate that the non-catalytic activity retained by BRMK804R, which is knocked down in brm RNAi mutant, is the driving force for decreased clock gene expression in flies expressing brmK804R. ChIP-qPCR revealed that CLK binding to per and tim promoters in brmK804R mutants is lower compared to that of control flies, despite the more open chromatin in these mutants (Fig. 2B). This suggests that the catalytically-inactive BRMK804R protein can influence CLK occupancy independent of ATP-dependent modifications in chromatin structure, perhaps through recruitment of repressive factors or factors that negatively impact CLK levels.Figure 2.


Understanding the role of chromatin remodeling in the regulation of circadian transcription in Drosophila.

Kwok RS, Lam VH, Chiu JC - Fly (Austin) (2015)

The effects of two classes of brm mutants on CLK-activated transcription. (A) Knockdown of brm via RNAi results in the reduction of both catalytic and non-catalytic functions. The decrease in catalytic activity leads to lower nucleosome density as compared to control, and decrease in non-catalytic function impairs recruitment of repressive complexes. As a result, CLK-activated gene expression is elevated. (B) Expression of catalytically-inactive BRMK804R protein results in decreased nucleosome density. The more open chromatin allows for an augmented effect of the non-catalytic function retained by BRMK804R, which may include recruitment of repressors. Experimental data suggests that interactors of BRM have a negative effect on CLK binding to the per promoter, and per expression is downregulated as a result of decreased CLK binding.36 (C) Wild type BRM possesses both non-catalytic and catalytic functions that balance each other and serve to fine-tune CLK-activated transcription. Non-catalytic function of BRM may promote recruitment of proteins that have repressive effects on transcription. The over-recruitment of repressive factors is prevented by the catalytic function of BRM to maintain nucleosome density.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4862430&req=5

f0002: The effects of two classes of brm mutants on CLK-activated transcription. (A) Knockdown of brm via RNAi results in the reduction of both catalytic and non-catalytic functions. The decrease in catalytic activity leads to lower nucleosome density as compared to control, and decrease in non-catalytic function impairs recruitment of repressive complexes. As a result, CLK-activated gene expression is elevated. (B) Expression of catalytically-inactive BRMK804R protein results in decreased nucleosome density. The more open chromatin allows for an augmented effect of the non-catalytic function retained by BRMK804R, which may include recruitment of repressors. Experimental data suggests that interactors of BRM have a negative effect on CLK binding to the per promoter, and per expression is downregulated as a result of decreased CLK binding.36 (C) Wild type BRM possesses both non-catalytic and catalytic functions that balance each other and serve to fine-tune CLK-activated transcription. Non-catalytic function of BRM may promote recruitment of proteins that have repressive effects on transcription. The over-recruitment of repressive factors is prevented by the catalytic function of BRM to maintain nucleosome density.
Mentions: The most surprising finding of our study was perhaps the fact that the two classes of brm mutants we examined exhibited opposing effects on circadian gene expression, with brm RNAi displaying increased clock gene expression compared to control and brmK804R showing downregulation of clock gene expression (Fig. 1D). Since both classes of brm mutants were predicted to show a decrease in catalytic activity and exhibited a reduction in nucleosome density at per and tim loci as assayed by histone H3 ChIP-qPCR, our results suggested that the BRM complex normally functions to condense the chromatin at clock gene loci. Moreover, our results strongly suggested that it is likely the intact non-catalytic activity of BRMK804R that led to the differential effects exhibited by the two classes of brm mutants with respect to clock gene expression. We postulate that when brm is knocked down via RNAi, the chromatin is more open due to a loss of nucleosome remodeling catalytic activity, allowing for an aberrant elevation in gene expression that results partly from an increased recruitment of general transcription machinery (Fig. 2A). While this explains the increase in clock gene expression and apparent decrease in RNAPII stalling in flies expressing brm RNAi, the opposite effect in clock gene expression observed in flies expressing the catalytically-inactive brmK804R remains puzzling, especially since the chromatin is also more relaxed in brmK804R flies as compared to wild type (Fig. 2B). Fortunately, investigation of changes in CLK binding to the per and tim promoters promptly provided us with valuable clues to indicate that the non-catalytic activity retained by BRMK804R, which is knocked down in brm RNAi mutant, is the driving force for decreased clock gene expression in flies expressing brmK804R. ChIP-qPCR revealed that CLK binding to per and tim promoters in brmK804R mutants is lower compared to that of control flies, despite the more open chromatin in these mutants (Fig. 2B). This suggests that the catalytically-inactive BRMK804R protein can influence CLK occupancy independent of ATP-dependent modifications in chromatin structure, perhaps through recruitment of repressive factors or factors that negatively impact CLK levels.Figure 2.

Bottom Line: The robust cycling of circadian gene expression is critical for proper timekeeping, and is regulated by transcription factor binding, RNA polymerase II (RNAPII) recruitment and elongation, and post-transcriptional mechanisms.Here, we discuss the characterization of the BRAHMA (BRM) chromatin-remodeling protein in Drosophila in the context of circadian clock regulation.Finally, we divulge ongoing efforts and investigative directions toward a deeper mechanistic understanding of transcriptional regulation of circadian gene expression at the chromatin level.

View Article: PubMed Central - PubMed

Affiliation: a Department of Entomology and Nematology ; University of California Davis ; Davis , CA 95616 , USA.

ABSTRACT
Circadian clocks enable organisms to anticipate daily changes in the environment and coordinate temporal rhythms in physiology and behavior with the 24-h day-night cycle. The robust cycling of circadian gene expression is critical for proper timekeeping, and is regulated by transcription factor binding, RNA polymerase II (RNAPII) recruitment and elongation, and post-transcriptional mechanisms. Recently, it has become clear that dynamic alterations in chromatin landscape at the level of histone posttranslational modification and nucleosome density facilitate rhythms in transcription factor recruitment and RNAPII activity, and are essential for progression through activating and repressive phases of circadian transcription. Here, we discuss the characterization of the BRAHMA (BRM) chromatin-remodeling protein in Drosophila in the context of circadian clock regulation. By dissecting its catalytic vs. non-catalytic activities, we propose a model in which the non-catalytic activity of BRM functions to recruit repressive factors to limit the transcriptional output of CLOCK (CLK) during the active phase of circadian transcription, while the primary function of the ATP-dependent catalytic activity is to tune and prevent over-recruitment of negative regulators by increasing nucleosome density. Finally, we divulge ongoing efforts and investigative directions toward a deeper mechanistic understanding of transcriptional regulation of circadian gene expression at the chromatin level.

No MeSH data available.


Related in: MedlinePlus