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Direct regulation of CLOCK expression by REV-ERB.

Crumbley C, Burris TP - PLoS ONE (2011)

Bottom Line: A REV-ERB response element (RevRE) was identified within this region of the CLOCK gene and was conserved between humans and mice.Additionally, the CLOCK RevRE conferred REV-ERB responsiveness to a heterologous reporter gene.Our data suggests that REV-ERBα plays a dual role in regulation of the activity of the BMAL1/CLOCK heterodimer by regulation of expression of both the BMAL1 and CLOCK genes.

View Article: PubMed Central - PubMed

Affiliation: The Scripps Research Institute, Jupiter, Florida, United States of America.

ABSTRACT
Circadian rhythms are regulated at the cellular level by transcriptional feedback loops leading to oscillations in expression of key proteins including CLOCK, BMAL1, PERIOD (PER), and CRYPTOCHROME (CRY). The CLOCK and BMAL1 proteins are members of the bHLH class of transcription factors and form a heterodimer that regulates the expression of the PER and CRY genes. The nuclear receptor REV-ERBα plays a key role in regulation of oscillations in BMAL1 expression by directly binding to the BMAL1 promoter and suppressing its expression at certain times of day when REV-ERBα expression levels are elevated. We recently demonstrated that REV-ERBα also regulates the expression of NPAS2, a heterodimer partner of BMAL1. Here, we show that REV-ERBα also regulates the expression another heterodimer partner of BMAL1, CLOCK. We identified a REV-ERBα binding site within the 1(st) intron of the CLOCK gene using a chromatin immunoprecipitation - microarray screen. Suppression of REV-ERBα expression resulted in elevated CLOCK mRNA expression consistent with REV-ERBα's role as a transcriptional repressor. A REV-ERB response element (RevRE) was identified within this region of the CLOCK gene and was conserved between humans and mice. Additionally, the CLOCK RevRE conferred REV-ERB responsiveness to a heterologous reporter gene. Our data suggests that REV-ERBα plays a dual role in regulation of the activity of the BMAL1/CLOCK heterodimer by regulation of expression of both the BMAL1 and CLOCK genes.

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Identification of a REV-ERBα binding site within the CLOCK gene.(a) Significant REV-ERBα occupancy was observed within the CLOCK gene within the 1st intron. The genomic structure of CLOCK is shown with REV-ERBα occupancy illustrated as the gray line. The arrow indicates the direction of transcription. The raw ChIP/chip data is shown in a window beneath the gene as is a screen shot from the integrative genome browser. (b) The region of REV-ERBα occupancy was scanned for conserved RevRE using the Evolutionarily Conserved Region Browser and MatInspector. A RevRE was found to be conserved between mice and humans, the alignment of the RevRE is shown. (c) Alignment of the CLOCK RevRE to a characterized RevRE in the Bmal1 promoter.
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pone-0017290-g001: Identification of a REV-ERBα binding site within the CLOCK gene.(a) Significant REV-ERBα occupancy was observed within the CLOCK gene within the 1st intron. The genomic structure of CLOCK is shown with REV-ERBα occupancy illustrated as the gray line. The arrow indicates the direction of transcription. The raw ChIP/chip data is shown in a window beneath the gene as is a screen shot from the integrative genome browser. (b) The region of REV-ERBα occupancy was scanned for conserved RevRE using the Evolutionarily Conserved Region Browser and MatInspector. A RevRE was found to be conserved between mice and humans, the alignment of the RevRE is shown. (c) Alignment of the CLOCK RevRE to a characterized RevRE in the Bmal1 promoter.

Mentions: A ChIP/chip screen was performed to determine regions where REV-ERBα is bound within the genome as previously described [9]. Significant REV-ERBα occupancy was observed with the CLOCK gene, as diagrammed in Figure 1a. Using the Evolutionarily Conserved Region Browser [15], it was determined that a putative RevRE is located within the CLOCK gene that is conserved between mice and humans, as shown in Figure 1b. The same site was detected using MatInspector [16]. An alignment between the CLOCK RevRE site to a characterized RevRE in the BMAL1 promoter is shown in Figure 1c.


Direct regulation of CLOCK expression by REV-ERB.

Crumbley C, Burris TP - PLoS ONE (2011)

Identification of a REV-ERBα binding site within the CLOCK gene.(a) Significant REV-ERBα occupancy was observed within the CLOCK gene within the 1st intron. The genomic structure of CLOCK is shown with REV-ERBα occupancy illustrated as the gray line. The arrow indicates the direction of transcription. The raw ChIP/chip data is shown in a window beneath the gene as is a screen shot from the integrative genome browser. (b) The region of REV-ERBα occupancy was scanned for conserved RevRE using the Evolutionarily Conserved Region Browser and MatInspector. A RevRE was found to be conserved between mice and humans, the alignment of the RevRE is shown. (c) Alignment of the CLOCK RevRE to a characterized RevRE in the Bmal1 promoter.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3066191&req=5

pone-0017290-g001: Identification of a REV-ERBα binding site within the CLOCK gene.(a) Significant REV-ERBα occupancy was observed within the CLOCK gene within the 1st intron. The genomic structure of CLOCK is shown with REV-ERBα occupancy illustrated as the gray line. The arrow indicates the direction of transcription. The raw ChIP/chip data is shown in a window beneath the gene as is a screen shot from the integrative genome browser. (b) The region of REV-ERBα occupancy was scanned for conserved RevRE using the Evolutionarily Conserved Region Browser and MatInspector. A RevRE was found to be conserved between mice and humans, the alignment of the RevRE is shown. (c) Alignment of the CLOCK RevRE to a characterized RevRE in the Bmal1 promoter.
Mentions: A ChIP/chip screen was performed to determine regions where REV-ERBα is bound within the genome as previously described [9]. Significant REV-ERBα occupancy was observed with the CLOCK gene, as diagrammed in Figure 1a. Using the Evolutionarily Conserved Region Browser [15], it was determined that a putative RevRE is located within the CLOCK gene that is conserved between mice and humans, as shown in Figure 1b. The same site was detected using MatInspector [16]. An alignment between the CLOCK RevRE site to a characterized RevRE in the BMAL1 promoter is shown in Figure 1c.

Bottom Line: A REV-ERB response element (RevRE) was identified within this region of the CLOCK gene and was conserved between humans and mice.Additionally, the CLOCK RevRE conferred REV-ERB responsiveness to a heterologous reporter gene.Our data suggests that REV-ERBα plays a dual role in regulation of the activity of the BMAL1/CLOCK heterodimer by regulation of expression of both the BMAL1 and CLOCK genes.

View Article: PubMed Central - PubMed

Affiliation: The Scripps Research Institute, Jupiter, Florida, United States of America.

ABSTRACT
Circadian rhythms are regulated at the cellular level by transcriptional feedback loops leading to oscillations in expression of key proteins including CLOCK, BMAL1, PERIOD (PER), and CRYPTOCHROME (CRY). The CLOCK and BMAL1 proteins are members of the bHLH class of transcription factors and form a heterodimer that regulates the expression of the PER and CRY genes. The nuclear receptor REV-ERBα plays a key role in regulation of oscillations in BMAL1 expression by directly binding to the BMAL1 promoter and suppressing its expression at certain times of day when REV-ERBα expression levels are elevated. We recently demonstrated that REV-ERBα also regulates the expression of NPAS2, a heterodimer partner of BMAL1. Here, we show that REV-ERBα also regulates the expression another heterodimer partner of BMAL1, CLOCK. We identified a REV-ERBα binding site within the 1(st) intron of the CLOCK gene using a chromatin immunoprecipitation - microarray screen. Suppression of REV-ERBα expression resulted in elevated CLOCK mRNA expression consistent with REV-ERBα's role as a transcriptional repressor. A REV-ERB response element (RevRE) was identified within this region of the CLOCK gene and was conserved between humans and mice. Additionally, the CLOCK RevRE conferred REV-ERB responsiveness to a heterologous reporter gene. Our data suggests that REV-ERBα plays a dual role in regulation of the activity of the BMAL1/CLOCK heterodimer by regulation of expression of both the BMAL1 and CLOCK genes.

Show MeSH
Related in: MedlinePlus