Modulation of retinoic acid receptor alpha activity by lysine methylation in the DNA binding domain

Metabolic labeling and detection with a methylated lysine-specific antibody confirm lysine methylation of RAR alpha in mammalian cells. We previously reported Lys (347) trimethylation of mouse retinoic acid receptor alpha (RAR alpha) in the ligand binding domain (LBD) that affected ligand sensitivity of the dissected LBD. Here we report two monomethylated residues, Lys (109) and Lys (171) identified by LC-ESI-MS/MS in the DNA binding domain (DBD) and the hinge region, which affect retinoic acid (RA) sensitivity, coregulator interaction and heterodimerization with retinoid X receptor (RXR) in the context of the full-length protein. Constitutive negative mutation at Lys (109), but not Lys (171), reduces RA-dependent activation. Methylation at Lys (109) plays a more dominant role than trimethylation at Lys (347) in terms of RA activation of the full-length receptor. Lys (109) is located in a homologous sequence (CEGC K GFFRRS) of the DBD in RARs and is conserved in the nuclear receptor superfamily even across the species boundary. This study uncovers a potential role for monomethylation at Lys (109) in coordinating the synergy between DBD and LBD for ligand-dependent activation of RAR alpha.