Expression of the vitamin D and the retinoid X receptors in Saccharomyces cerevisiae: alternative in vivo models for ligand-induced transactivation.
The transcription factors of the nuclear hormone receptor family regulate gene expression via a complex network of macromolecular interactions. The ligand dependent activity of the vitamin D receptor is of particular interest because it modulates gene expression by the heterodimeric interaction with retinoid X receptors. We report here that individual functions of the vitamin D receptor including DNA-binding, homo- and heterodimerization and transactivation can be reconstituted in the yeast Saccharomyces cerevisiae. Interestingly, the simultaneous expression of the native vitamin D receptor and the retinoid X receptor beta resulted in a ligand independent transactivation of the lacZ reporter gene coupled to a mouse osteopontin vitamin D response element. However, homodimerization of the vitamin D receptor and heterodimerization were strongly enhanced upon ligand binding, when the receptors were expressed as fusion proteins with the Gal4 transcription factor in a yeast two-hybrid system. Furthermore, transactivating activity of a Gal4-fused vitamin D receptor was induced by vitamin D in a one-hybrid system devoid of retinoid X receptors. In addition, both Gal4-based systems behaved similar with regard to their dose-dependent response to vitamin D and related compounds when compared to the transcriptional activity of the vitamin D receptor in transiently transfected MCF-7 cells. Our results point out that specific ligands strongly enhanced receptor dimerization and induced transactivation in yeast and in MCF-7 cells. The constitutive transactivation by vitamin D receptor-retinoid X receptor heterodimers in yeast, depending on DNA binding of the receptors, strongly argues for the existence of cofactors, which are absent in yeast, but play a fundamental role in gene regulation in higher eukaryotic organisms.
Berghöfer-Hochheimer
Y
Y
Department of Cell and Molecular Biology, Hans-Knöll-Institut für Naturstoff-Forschung e.V., Jena, Germany.
Zurek
C
C
Langer
G
G
Munder
T
T
D000818:Animals; D001943:Breast Neoplasms; D002135:Calcium-Binding Proteins / Q000378:metabolism; D037282:Calreticulin; D019281:Dimerization; D006801:Humans; D008024:Ligands; D051379:Mice; D010957:Plasmids; D018167:Receptors, Calcitriol / Q000096:biosynthesis* / Q000235:genetics*; D018168:Receptors, Retinoic Acid / Q000096:biosynthesis* / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D047488:Retinoid X Receptors; D012261:Ribonucleoproteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D014157:Transcription Factors / Q000096:biosynthesis* / Q000235:genetics*; D015533:Transcriptional Activation; D014407:Tumor Cells, Cultured
D002135:Calcium-Binding Proteins; D037282:Calreticulin; D008024:Ligands; D018167:Receptors, Calcitriol; D018168:Receptors, Retinoic Acid; D011993:Recombinant Fusion Proteins; D047488:Retinoid X Receptors; D012261:Ribonucleoproteins; D014157:Transcription Factors