Osteoporosis is a metabolic bone disorder characterized by low bone mass and microstructural deterioration of bone tissue, culminating in increased bone fragility and increased risk of fracture. Two principle steroid hormones which have strong links to the osteoporotic disease process are estrogen (E2) and 1alpha,25-dihydroxyvitamin D3 (1,25(OH)2D3). The biological effects of E2 and 1,25(OH)2D3 are mediated through nuclear receptors (the ER and the VDR, respectively) which function as ligand activated transcription factors that control the expression of specific genes or gene networks in bone cells. However, the precise mechanisms through which these receptors influence the transcriptional process are largely unknown. In preliminary studies, we have isolated a novel, human cDNA clone that encodes a 62,000 Da nuclear protein that interacts selectively with several nuclear receptors, including the VDR and the ER. Importantly, coexpression of this cDNA clone in a VDR-responsive gene expression system dramatically augments l,25(OH)2D3-/VDR-mediated transcription. The clone has been designated as NCoA-62, for Nuclear Receptor CoActivator of 62,000 Da molecular mass. The long-term goal of this proposal is to understand the molecular mechanism of VDR- and ER- mediated gene expression in bone cells and in the short-term, this research proposal focuses on determining the functional role of NCoA-62 in vitamin D and estrogen regulated transcription. Our hypothesis states that NCoA-62 is a bridging protein that serves as an important communication link between the promoter-bound receptor and the transcription preinitiation complex. To test this hypothesis, we propose four specific aims that: 1. establish whether NCoA62 is involved in ER-mediated transcription in mammalian cells 2. identify the protein domains that are important for NCoA62 coactivator activity 3. examine the essentiality of NCoA-62 in 1,25(OH)2D3- and E2-mediated transcription in osteoblast cell lines 4. identify and characterize osteoblast nuclear proteins that interact with NCoA62. These studies are designed to improve our understanding of the basic mechanisms involved in vitamin D and estrogen dependent transcription in bone and to identify the important functional components of this complex process. Through an acquired understanding of this complex process, rational therapeutic strategies will be realized in the treatment and prevention of debilitating bone disorders such as osteoporosis.
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