Relationship between circadian oscillations of Rev-erbalpha expression and intracellular levels of its ligand, heme

Abstract

The nuclear hormone receptors, REV-ERBalpha [NR1D1] and REV-ERBbeta [NR1D1], were recently demonstrated to be receptors for the porphyrin, heme. Heme regulates the ability of these receptors to repress transcription of their target genes via modulation of the affinity of the receptor's ligand binding domain for the corepressor, NCoR. The REV-ERBs function as critical components of the mammalian clock and their expression oscillates in a circadian manner. Here, we show that in NIH3T3 cells intracellular heme levels also oscillate in a circadian fashion. These data are the first to show the temporal relationship of intracellular heme levels to the expression of its receptor, Rev-erbalpha, and suggest that the rapid oscillations in heme levels may an important component regulating REV-ERB transcriptional activity.

Fig. 1

Circadian oscillations of Per2 and Bmal1 expression in NIH3T3 cells following serum shock. Cells were synchronized with a 50% horse serum shock followed by analysis of Per2 and Bmal1 mRNA by qPCR at the times indicated. *P < 0.05.

Fig. 2

Circadian oscillations of Rev-erbα expression and intracellular [Heme] in NIH3T3 cells following serum shock. Cells were synchronized with a 50% horse serum shock followed by analysis of Rev-erbα mRNA by qPCR and intracellular [Heme] at the times indicated. *P < 0.05.

Fig. 3

Circadian oscillations of Per2 expression and intracellular [Heme] in NIH3T3 cells following heme shock. Cells were synchronized with a 30 mM hemin shock followed by analysis of Per2 mRNA by qPCR and intracellular [Heme] at the times indicated. *P < 0.05.

Fig. 4

Model comparing the oscillations in Rev-erbα and intracellular [Heme] suggesting the possibility that there may be variability in the level of receptor occupancy and thus repressor activity while REV-ERBα is elevated during the circadian rhythm.