Characterization of the Core Mammalian Clock Component, NPAS2, as a REV-ERBα/RORα Target Gene

The mammalian clock is regulated at the cellular level by a transcriptional/translational feedback loop. BMAL1/CLOCK (or NPAS2) heterodimers activate the expression of the PERIOD (PER) and CRYPTOCHROME (CRY) genes acting as transcription factors directed to the PER and CRY promoters via E-box elements. PER and CRY proteins form heterodimers and suppress the activity of the BMAL1/CLOCK (or NPAS2) completing the feedback loop. The circadian expression of BMAL1 is influenced by retinoic acid receptor-related orphan receptor alpha (ROR alpha) and REV-ERB alpha, two nuclear receptors that target a ROR-response element in the promoter of the BMAL1 gene. Given that BMAL1 functions as an obligate heterodimer with either CLOCK or NPAS2, it is unclear how the expression of the partner is coordinated with BMAL1 expression. Here, we demonstrate that NPAS2 is also a ROR alpha and REV-ERB alpha target gene. Using a ChIP/microarray screen, we identified both ROR alpha and REV-ERB alpha occupancy of the NPAS2 promoter. We identified two functional ROREs within the NPAS2 promoter and also demonstrate that both ROR alpha and REV-ERB alpha regulate the expression of NPAS2 mRNA. These data suggest a mechanism by which ROR alpha and REV-ERB alpha coordinately regulate the expression of the positive arm of the circadian rhythm feedback loop.