The bioactive form of vitamin D, calcitriol [1a,25-dihydroxyvitamin D3; VitD] is primarily a calcemic hormone. However, it not only stimulates intestinal calcium absorption, bone calcium resorption and renal calcium reabsorption, but also regulates growth and differentiation of many cell types, and displays immunoregulatory and anti-inflammatory activities. The latter properties provide a basis to treat many diseases (e.g. osteoporosis, autoimmune diseases, neurodegenerative diseases and various types of cancer). Unfortunately, as supra-physiological doses of VitD that are required to exhibit therapeutic activities induce hypercalcemia, causing mineralization of the kidneys, heart, blood vessels and cutaneous tissues that can lead to organ failure and death, this hormone cannot be used as such as a treatment. Calcitriol mediates pleiotropic effects through activation of the Vitamin D Receptor (VDR), a member of the nuclear receptor superfamily which heterodimerizes with retinoid X receptors (RXRs), to control target gene transcription. Rare genetic mutations in human, termed type II Hereditary Vitamin D Resistant Ricket (HVDRR), have confirmed the importance of VDR in the skeletal system. The discovery that VDR is expressed in many cell types/tissues highlights a central role of VitD signaling in many processes. Besides genomic and rapid non-genomic actions of calcitriol, VDR has also ligand-independent functions. Many companies and academic laboratories have synthesized VitD analogs to potentiate beneficial properties of VitD agonists. These analogs were designed before the crystal structure of liganded VDR was elucidated, by introducing chemical modifications of the calcitriol skeleton, and subsequent in vivo screening for specific activities, as their calcemic activity could not be predicted. However, up to now, all compounds identified with decreased calcemic activities have only a low therapeutic index, thus limiting their clinical use. Therefore, to facilitate the development of VitD analogs with wider therapeutic windows, it is crucial to unveil the molecular mechanisms that underlie cell and/or gene selectivity of VitD. To unravel these mechanisms, we propose a novel approach combining structure-function studies and mouse models, based on recent contributions made by the applicants that open up novel questions to be addressed. Taking advantage of VDR-null mice and VDRgem mice, expressing a mutant VDR the activity of which is selectively induced by Gemini VitD analogs, recently established in collaboration between the two partners involved in this project, combined with structural-functional studies, we intend to: - demonstrate that VDR mediates VitD-induced calcemic effects, - identify direct VDR target genes in various mouse tissues, - determine structural modifications of VitD analogs that promote cell and/or promoter specific actions via VDR, - and identify the key structural modifications of VDR responsible for such specificities. The results generated through this challenging project should uncover the cellular and molecular mechanisms underlying tissue/cell specific actions of VitD. The use of this novel knowledge will allow to design in silico VDR agonists and establish cell-based screens, to identify potent VitD analogs with reduced side effects, for the treatment of various diseases.