Phosphorylation of the retinoic acid receptor-alpha by protein kinase A.

The phosphorylation of retinoic acid receptor-alpha 1 (RAR alpha 1) by PKA was investigated both in vitro and in vivo. We show that bacterially expressed RAR alpha 1 is phosphorylated in vitro by protein kinase A (PKA) at the unique serine residue 369 located in the C-terminal end of the E region. We also show that RAR alpha 1 overexpressed in COS-1 cells is phosphorylated on multiple serine residues and that phosphorylation at serine 369 occurs only when COS-1 cells are cotransfected with PKA or treated with forskolin. RAR alpha 1 mutants were constructed in which serine 369 was replaced by an alanine (S369A) or a glutamic acid (S369E) residue. Comparison of the tryptic phosphopeptide patterns of wild type and mutated RAR alpha 1 overexpressed in COS-1 cells allowed us to confirm that serine 369 is the unique phosphorylation site for PKA in cultured cells. The DNA-binding efficiency of RAR alpha/retinoid X receptor-alpha (RXR alpha) heterodimers was enhanced in vitro by the S369E mutation. However, in tr...