Epigenetic regulation of the circadian clock: Role of 5-aza-2'-deoxycytidine

Abstract

ABSTRACT We have been investigating transcriptional regulation of the BMAL1 gene, a critical component of the mammalian clock system including DNA methylation. Here, a more detailed analysis of the regulation of DNA methylation of BMAL1 proceeded in RPMI8402 lymphoma cells. We found that CpG islands in the BMAL1 and the PER2 promoters were hyper-and hypo-methylated, respectively and that 5-aza-2'-deoxycytidine (aza-dC) not only enhanced PER2 gene expression but also PER2 oscillation within 24 hours in RPMI8402 cells. That is, such hypermethylation of CpG islands in the BMAL1 promoter restricted PER2 expression that was recovered by aza-dC within one day in these cells. These results suggest that the circadian clock system can be recovered through BMAL1 expression induced by aza-dC within a day. The RPIB9 promoter of RPMI8402 cells, which is a methylation hotspot in lymphoblastic leukemia, was also hypermethylated, and aza-dC gradually recovered RPIB9 expression in three days. In addition, methylation-specific PCR revealed a different degree of aza-dC-induced methylation release by between BMAL1 and RPIB9. These results suggest that the aza-dC-induced recovery of gene expression from DNA methylation is dependent on a gene, for example, the rapid response to demethylation by the circadian system, and thus is of importance to clinical strategies for treating cancer