BIOCHEMISTRY AND GENETICS OF VITAMIN D RECEPTOR ACTIONS

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DESCRIPTION (Adapted from the Applicant's Abstract): The vitamin D receptor (VDR) binds to the vitamin D response element (VDRE) as a heterodimeric complex with the retinoid X receptor (RXR) and mediates the effects of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] on gene expression. It is well-established that additional nuclear proteins, termed receptor or nuclear auxiliary factors, are required for high affinity binding to the VDREs. The principal investigator used a yeast two-hybrid system to clone cDNAs which code for VDR-interacting proteins, two of them in the absence of 1,25(OH)2D3 and nine in the presence of 1,25(OH)2D3. She hypothesizes that the vitamin D interacting proteins obtained from the yeast two-hybrid system are transcription factors for 1,25(OH)2D3 mediated gene regulation. In support of this hypothesis, she found that one of them, human steroid receptor coactivator-1 (SRC-1), a known transcription factor for other steroid hormones, interacts with the VDR in a ligand-dependent manner as shown by (-galactosidase production. Interaction of VDR and SRC-1 takes place at physiologic concentrations of 1,25(OH)2D3. Deletion-mutation analysis of the VDR established that the ligand-dependent activation domain (AF-2) of the receptor is required for interaction with SRC-1 and deletion-mutation analysis of SRC-1 demonstrated that 27 amino acids are required for interaction with the VDR. Further, antisense studies indicate that SRC-1 is required for stimulation of alkaline phosphatase production by human osteosarcoma cells in response to 1,25(OH)2D3. The principal investigator proposes to extend these studies to investigate whether and to what extent SRC-1 and other putative factors are involved in 1,25(OH)2D3 mediated gene regulation. The results are predicted to provide important new information about the molecular biology of 1,25(OH)2D3-mediated effects on cellular function.