Epigenetic regulation by RAR alpha maintains ligand-independent transcriptional activity

Retinoic acid receptors (RARs) alpha, beta and gamma are key regulators of embryonic development. Hematopoietic differentiation is regulated by RAR alpha, and several types of leukemia show aberrant RAR alpha activity. Through microarray expression analysis, we identified transcripts differentially expressed between F9 wild-type (Wt) and RAR alpha knockout cells cultured in the absence or presence of the RAR-specific ligand all trans retinoic acid (RA). We validated the decreased Mest, Tex13, Gab1, Bcl11a, Tcfap2a and HMGcs1 transcript levels, and increased Slc38a4, Stmn2, RpL39l, Ref2L, Mobp and Rlf1 transcript levels in the RAR alpha knockout cells. The decreased Mest and Tex13 transcript levels were associated with increased promoter CpG-island methylation and increased repressive histone modifications (H3K9me3) in RAR alpha knockout cells. Increased Slc38a4 and Stmn2 transcript levels were associated with decreased promoter CpG-island methylation and increased permissive histone modifications (H3K9/K14ac, H3K4me3) in RAR alpha knockout cells. We demonstrated specific association of RAR alpha and RXR alpha with the Mest promoter. Importantly, stable expression of a dominant negative, oncogenic PML-RAR alpha fusion protein in F9 Wt cells recapitulated the decreased Mest transcript levels observed in RAR alpha knockout cells. We propose that RAR alpha plays an important role in cellular memory and imprinting by regulating the CpG methylation status of specific promoter regions.