1alpha, 25-dihydroxyvitamin D3 (1,25OH2D3) and estrogen (E2) are steroid hormones that are essential for normal skeletal development, and for maintaining bone integrity. These hormones function through nuclear receptors to regulate the transcription of specific genes or gene networks. The continuing long-term goal of this proposal centers on defining the molecular mechanisms through which these two ligand-activated receptors regulate the transcription machinery in the skeletal system. This continuation proposal focuses on NR comodulatory proteins and, more specifically, on our discovery of a novel coactivator, NCoA62, that is involved in VDR-, ER-, and other NR-mediated transactivation systems. Most importantly, NCoA62 is unrelated to other NR comodulators, such as the SRC family of coactivators, and the mechanisms through which NCoA62 functions are poorly understood. In the past 4 years of funding, we have made considerable progress in defining the significance and potential molecular mechanisms involved in NCoA62 coactivation. The most significant findings include the preferential interaction of NCoA62 with the VDR-RXR heterodimer, ternary complex formation between VDR-NCoA62-SRCs, and the cooperative effects of NCoA62 and SRCs in VDR-activated transcription. These studies imply an intimate interplay between NCoA62 and distinct coactivator proteins in VDR-mediated transcription. On the basis of our past 4 years on this project, we propose the general hypothesis that NCoA62 is a novel VDR coactivator that associates with the VDR-RXR heterodimer in association with other NR coactivators, and the interaction with each coactivator class is required for VDR-mediated transactivation. To test this hypothesis, we propose three Specific Aims that will: (1) establish the functional relevance or biological significance of NCoA62; (2) characterize functional domains of NCoA62; and (3) probe the molecular mechanisms of multi-coactivator assemblies with the VDR-RXR heterodimer. These studies are designed to improve our understanding of the fundamental mechanisms involved in vitamin D- and estrogen-dependent transcription in bone, and they will define important transcriptional factors involved in this complex process. Identifying novel components that are essential to the basic transcriptional mechanisms of VDR and ER in bone cells may eventually provide molecular targets to develop novel drugs that protect against osteoporotic bone loss, or the more general skeletal abnormalities observed in the elderly.
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