Characteristics of vitamin D3 receptor (VDR) binding to the vitamin D response element (VDRE) in rat bone sialoprotein gene promoter.
Bone sialoprotein (BSP) is a mineralised tissue-specific protein that is highly expressed during the initial formation of bone and cementum. Expression of BSP is suppressed by the osteotropic hormone, 1,25-dihydroxyvitamin D3 (vitamin D3), which regulates bone remodelling. In previous studies, we have identified a vitamin D response element (VDRE) that is integrated with a novel inverted TATA box in the rat BSP promoter which mediates the suppression of BSP transcription (1). Although the nucleotide sequences of VDREs in different genes conform to a direct (hexamer) repeat, spaced by three nucleotides, the precise sequences are unique for each VDRE. To determine whether the nucleotide differences in the VDRE influence VDR binding, we have compared interactions of VDR proteins with various VDREs using gel mobility shift analysis. Both natural and recombinant VDRs bound to rat BSP and both mouse and porcine osteopontin (OPN) VDRE oligonucleotides in a concentration-dependent manner with a strong preference for dimer formation, whereas equal amounts of dimer and monomer were bound to the human osteocalcin VDRE. However, whereas a truncated VDR comprising the DNA binding domain alone bound the mouse osteopontin VDRE, it failed to interact with the porcine OPN and rat BSP VDREs. VDR binding to the BSP was sequence specific, as shown by mutagenesis analysis, and could be abolished by heat and VDR antibody. These studies demonstrate that subtle differences in the nucleotide sequence of VDREs affect VDR binding, which mediates the vitamin D3 response.
Li
J J
JJ
Medical Research Council Group in Periodontal Physiology, Faculty of Dentistry, University of Toronto, Ontario, Canada. jack.li@utoronto.ca
Kim
R H
RH
Zhang
Q
Q
Ogata
Y
Y
Sodek
J
J
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D002117:Calcitriol / Q000494:pharmacology; D002460:Cell Line; D004247:DNA / Q000235:genetics / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects; D006801:Humans; D058574:Integrin-Binding Sialoprotein; D051379:Mice; D008957:Models, Genetic; D009805:Odontogenesis / Q000235:genetics; D053495:Osteopontin; D011401:Promoter Regions, Genetic; D051381:Rats; D018167:Receptors, Calcitriol / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion; D012795:Sialoglycoproteins / Q000235:genetics*; D013552:Swine; D016385:TATA Box; D014162:Transfection
C546871:IBSP protein, human; C546872:Ibsp protein, mouse; C546873:Ibsp protein, rat; D058574:Integrin-Binding Sialoprotein; D018167:Receptors, Calcitriol; D011994:Recombinant Proteins; C506895:SPP1 protein, human; D012795:Sialoglycoproteins; C506896:Spp1 protein, mouse; C506897:Spp1 protein, rat; D053495:Osteopontin; D004247:DNA; D002117:Calcitriol