title: Retinoid receptors cause distortion of the retinoic acid response element in the phosphoenolpyruvate carboxykinase gene promoter.
abstract: Functional retinoic acid response elements (RAREs) have been described wherein the direct repeats are separated by 1, 2 or 5 bp (termed DR1, DR2 and DR5 respectively). We have previously shown that retinoic acid receptor/retinoid X receptor (RAR/RXR) binds a DR1 RARE within the phosphoenolpyruvate carboxykinase (PEPCK) gene promoter and is the trans-acting complex that mediates the retinoic acid (RA) response. However, the mechanism of trans-activation is unknown. The consequences of RAR/RXR binding to the PEPCK RARE were examined using a circular permutation analysis as a first step to explore the possible role of DNA conformational changes in the RA response. The RAR/RXR heterodimer produced a distortion angle of 78 degrees. The DNA distortion was shown to be at the centre of the PEPCK RARE; RA did not affect the severity of the distortion angle or the location of the distortion centre. Monomers and homodimers of RAR also distorted the DNA, but to a lesser extent than did RAR/RXR. The results of a phasing analysis demonstrated that RAR/RXR heterodimers did not induce a static DNA bend, in either the presence or the absence of RA. A cyclization kinetics assay was employed to show that RAR/RXR binding affected DNA ring closure in a phase-sensitive, RA-insensitive, manner. Taken together, these observations support the idea that RAR/RXR heterodimers distort the structure of the PEPCK RARE, at least in part, by altering DNA flexibility. The conformational change in the PEPCK RARE upon RAR/RXR binding has implications for how RAR/RXR heterodimers recognize various RARE structures.
authors.affiliation: Department of Molecular Physiology and Biophysics, Vanderbilt University School, Nashville, TN 37232, USA.
authors.forename: D K
authors.initials: DK
authors.lastname: Scott
authors.forename: R K
authors.initials: RK
authors.lastname: Hall
authors.forename: D K
authors.initials: DK
authors.lastname: Granner
mesh_terms: D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004926:Escherichia coli; D006801:Humans; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010729:Phosphoenolpyruvate Carboxykinase (GTP) / Q000235:genetics*; D010957:Plasmids; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D018168:Receptors, Retinoic Acid / Q000096:biosynthesis / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D015183:Restriction Mapping; D000072482:Retinoic Acid Receptor alpha; D047488:Retinoid X Receptors; D012176:Retinoids / Q000378:metabolism; D014157:Transcription Factors / Q000096:biosynthesis / Q000378:metabolism*
chemical_list: D017931:DNA Primers; D004268:DNA-Binding Proteins; C000607071:RARA protein, human; D018168:Receptors, Retinoic Acid; D011994:Recombinant Proteins; D000072482:Retinoic Acid Receptor alpha; D047488:Retinoid X Receptors; D012176:Retinoids; D014157:Transcription Factors; D010729:Phosphoenolpyruvate Carboxykinase (GTP)