RAR-mediated epigenetic control of the cytochrome P450 Cyp26a1 in embryocarcinoma cells

Retinoic acid (RA) is a signaling molecule that plays a pivotal role in major cellular processes and vertebrate development. RA action is mediated by specialized transcription factors, the nuclear RA receptors (RARs), which regulate the transcription of genes containing a RA-responsive element (RARE). Here we demonstrate that the genes for the RA-receptor RAR beta 2 and the cytochrome P450 RA-specific hydrolase Cyp26a1 involved in RA catabolism are coordinately regulated by RA. We found that both RAR beta 2 and Cyp26a1 genes are epigenetically silenced in the absence of DNA methylation in RAC65, a P19 embryocarcinoma cell line derivative carrying a dominant-negative RAR alpha mutant and resistant to the growth-inhibitory and differentiation effects of RA. In response to RA, RAR beta 2 transcription is epigenetically regulated by RAR alpha. Similarly, we found that Cyp26a1 transcription is epigenetically regulated by RAR beta 2. Knocking down RAR beta 2 transcription by RNA interference in wild-type P19 cells, with an intact RAR alpha, induced Cyp26a1 transcriptional repression in the absence of DNA methylation. Concomitantly, cells developed RA resistance and did not undergo RA-induced neuron differentiation. Apparently, RAR alpha, RAR beta 2 and Cyp26a1 are components of a RA-regulated gene network. Factors affecting an upstream gene of the network can trigger repressive chromatin changes-which are propagated in a domino fashion-at downstream genes of the network. This study also shows that chromatin inactivity, and consequent transcriptional silencing, can be achieved in the absence of DNA methylation