Promoter of FGF8 reveals a unique regulation by unliganded RARalpha.
We previously reported that retinoids were inducing a complete switch in the expression of two isoforms from the fgf8 gene. In order to gain insight into the transcriptional mechanisms possibly involved in this regulation, we cloned and sequenced a fragment of genomic DNA encompassing 6 kb of the region 5' upstream of the fgf8 coding sequence and investigated its promoter elements. A comprehensive series of biochemical and cellular experiments determined two distinct functional regions cis-responsive to retinoids and/or their receptors: (i) a canonical RARE (type DR2) which is the cis target of a RARalpha-RXRalpha liganded heterodimer; and (ii) a completely novel type of response element composed of two half-binding sites separated by 87 nucleotides, which we demonstrate to be the target of an unliganded RARalpha homodimer phosphorylated on the Ser77 residue. Combined activities of these cis and trans-acting factors support a model of a complex regulation of fgf8 expression: by alternative binding to two distinct promoter elements, phosphorylated-unliganded-RARalpha homodimer or its liganded form have two distinct and mutually exclusive trans-activating activities, explaining the expression of two different isoforms of fgf8.
Brondani
Vincent
V
Oncology Research, Novartis Pharma AG, CH-4002 Basel, Switzerland.
Klimkait
Thomas
T
Egly
Jean-Marc
JM
Hamy
François
F
D016475:3T3 Cells; D000495:Allosteric Site; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D004247:DNA / Q000235:genetics / Q000378:metabolism; D018983:DNA Footprinting; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D003850:Deoxyribonuclease I / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D051524:Fibroblast Growth Factor 8; D005346:Fibroblast Growth Factors / Q000737:chemistry / Q000235:genetics*; D005786:Gene Expression Regulation; D006801:Humans; D008024:Ligands; D051379:Mice; D008969:Molecular Sequence Data; D010766:Phosphorylation; D010768:Phosphoserine / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011485:Protein Binding; D020033:Protein Isoforms / Q000737:chemistry / Q000235:genetics; D018168:Receptors, Retinoic Acid / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D020218:Response Elements / Q000235:genetics; D000072482:Retinoic Acid Receptor alpha; D047488:Retinoid X Receptors; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D015533:Transcriptional Activation
D004268:DNA-Binding Proteins; C496456:FGF8 protein, human; C496457:Fgf8 protein, mouse; D008024:Ligands; D020033:Protein Isoforms; C000607071:RARA protein, human; C000607072:Rara protein, mouse; D018168:Receptors, Retinoic Acid; D000072482:Retinoic Acid Receptor alpha; D047488:Retinoid X Receptors; D014157:Transcription Factors; D051524:Fibroblast Growth Factor 8; D010768:Phosphoserine; D005346:Fibroblast Growth Factors; D004247:DNA; D003850:Deoxyribonuclease I