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 Lys³⁴⁷ 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, Lys¹⁰⁹ and Lys¹⁷¹ 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 Lys¹⁰⁹, but not Lys¹⁷¹, reduces RA-dependent activation. Methylation at Lys¹⁰⁹ plays a more dominant role than trimethylation at Lys³⁴⁷ in terms of RA activation of the full-length receptor. Lys¹⁰⁹ is located in a homologous sequence (CEGCKGFFRRS) 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 Lys¹⁰⁹ in coordinating the synergy between DBD and LBD for ligand-dependent activation of RAR alpha.