The vitamin D binding protein (DBP) transports vitamin D3, an important factor in proliferation of monocytes and osteoblasts, to its target tissues throughout the body. Our laboratory's study of the molecular biology of the vitamin D binding protein has led to the cloning, characterization and chromosomal mapping of the human DBP gene. The DBP cDNA probe identified DBP mRNA transcripts in cultured bone cells (osteoblast-like cells) and in bone-forming tissue of humans and rodents. Synthesis of DBP protein by osteoblast-like cell lines was confirmed immunologically. Since no person with a significant deficiency of DBP has been found after screening human populations throughout the world for 35 years, it has been suggested that an inherited deletion of the DBP gene is incompatible with fetal life. Increased plasma DBP levels are induced in chickens and in humans by estrogen administration. Temporal regulation of the DBP gene expression occurs during development and aging in mice. Because of its tissue-specific expression in liver and bone and because its expression is subject to both hormonal and developmental regulation, the DBP gene is an excellent model for obtaining information on the regulation of gene expression. Characterization of the genomic sequence of the 5' promoter region of the DBP gene has recently been completed and contains conserved DNA sequences identical or homologous to specific cis- regulatory elements in other genes that respond to environmental and developmental signals. Recombinant DNA technology will be utilized for the introduction of normal and chemically altered DBP minigenes into transfected cells capable of expressing DBP in vitro and into transgenic mice capable of expressing DBP in vitro and into transgenic mice capable of expressing DBP in vivo. The goals of this proposal are to (1) quantitate transcription of DBP mRNA in cultured human hepatoma and osteoblast-like cell lines modulated by hormonal and inflammatory agents and (2) to identify regulatory elements in the promoter region of the DBP gene that respond to these agents in vitro and in vivo, by correlating DBP expression with minigene constructs containing altered or deleted DNA sequences. The study proposed here will furnish a molecular explanation of gene regulation of the vitamin D binding protein in liver and bone-forming cells. The results may provide information important to age-related bone abnormalities such as osteoporosis.
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