Two differentially active alternative promoters control the expression of the zebrafish orphan nuclear receptor gene Rev-erbα

The orphan nuclear receptor Rev-erba (NR1D1) plays an important role in the regulation of the circadian pacemaker and its expression has been shown to be regulated with a robust circadian rhythm in zebrafish and mammals. In addition, in zebrafish its expression has been shown to be developmentally regulated. In order to analyze the mechanisms of the zfRev-erb alpha gene regulation, we have isolated its 5'-upstream region. We found that two promoters control the zf Rev-erba expression. The first one (ZfP1) is characterized by a very high degree of sequence identity with the mammalian P1 promoter and contains, as the mammalian P1, a functional Rev-erba-binding site (RevDR2). Inhibition of zfRev-erba activity in zebrafish embryos using antisense-morpholino knockdown results in an increase of zfRev-erba gene expression suggesting that zfRev-erba is repressing its own transcription in vivo. In addition, we show that ROR orphan receptors also regulate in vitro and in vivo zfRev-erba gene expression through the same RevDR2 element. In contrast, the second promoter Zf P2 is strikingly different from the mammalian P2: its sequence is not conserved between zebrafish and mammals and is not regulated by the same transcription factors. Together, these data suggest that ZfP1 is orthologous to the mammalian P1 promoter, whereas zebrafish ZfP2 has no mammalian ortholog and does not function like ZfP1 to control Rev-erba expression.