The conserved amphipatic alpha-helical core motif of RARgamma and RARalpha activating domains is indispensable for RA-induced differentiation of F9 cells.
In monolayers cultures, retinoic acid (RA) induces the differentiation of F9 embryonal carcinomal (EC) cells into primitive endoderm-like cells, while a combination of RA and dibutyryl cAMP leads to parietal endoderm-like differentiation. Knock out of all RARgamma isoforms (RARgamma(-/-) line) drastically impairs primitive and subsequent parietal endodermal differentiation and affects the induction of many endogenous RA-responsive genes. Using lines that reexpress RARgamma2 or overexpress RARalpha1 lacking their AF-2AD core (RARgammadeltaAF2 and RARalphadeltaAF2, 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 receptors, is required for the induction of differentiation and target gene expression upon RA treatment of F9 EC cells. We also show that these deletion mutants behave as dominant negatives.
Plassat
J
J
Institut de Génétique et de Biologie Moléculaire et Cellulaire, CNRS/INSERM/ULP/Collège de France, BP 163, CU de Strasbourg, France.
Penna
L
L
Chambon
P
P
Rochette-Egly
C
C
D000595:Amino Acid Sequence; D000818:Animals; D000970:Antineoplastic Agents / Q000378:metabolism / Q000494:pharmacology*; D018236:Carcinoma, Embryonal; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D002461:Cell Line, Transformed; D017124:Conserved Sequence; D004707:Endoderm / Q000166:cytology / Q000502:physiology; D017353:Gene Deletion; D015972:Gene Expression Regulation, Neoplastic / Q000502:physiology; D016296:Mutagenesis / Q000502:physiology; D011401:Promoter Regions, Genetic / Q000502:physiology; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D018168:Receptors, Retinoic Acid / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D000072482:Retinoic Acid Receptor alpha; D014162:Transfection; D014212:Tretinoin / Q000378:metabolism / Q000494:pharmacology*; D014407:Tumor Cells, Cultured; D000097768:Retinoic Acid Receptor gamma
D000970:Antineoplastic Agents; D018168:Receptors, Retinoic Acid; D000072482:Retinoic Acid Receptor alpha; D014212:Tretinoin; D000097768:Retinoic Acid Receptor gamma