The conserved amphipatic alpha-helical core motif of RAR gamma and RAR alpha activating domains is indispensable for RA-induced differentiation of F9cells

In monolayers cultures, retinoic acid (RA) induces the differentiation of F 9 embryonal carcinomal (EC) cells into primitive endoderm-like cells, while a combination of RA and dibutyryl cAMP leads to parietal endoderm-like dif ferentiation. Knock out of all RAR gamma isoforms (RAR gamma(-/-) line) dra stically impairs primitive and subsequent parietal endodermal differentiati on and affects the induction of many endogenous RA-responsive genes. Using lines that reexpress RAR gamma 12 or overexpress RAR alpha 1 lacking their AF-2AD core (RAR gamma Delta AF2 and RAR gamma Delta AF2, respectively), we show that this conserved amphipatic alpha-helical motif (helix 12) of the ligand binding domain, and therefore the activation function AF-2 of both r eceptors, is required for the induction of differentiation and target gene expression upon RA treatment of F9 EC cells. We also show that these deleti on mutants behave as dominant negatives.