Vitamin D must undergo hydroxylation to 25-hydroxyvitamin D [25(OH)D] by the enzyme vitamin D-25-hydroxylase (CYP25) to become biologically active. Porcine CYP25 cDNA was cloned and characterized. The gene encodes a cytochrome P450 microsomal enzyme expressed in liver and kidney that catalyzes the conversion of vitamins D2 and D3 to 25(OH)D3, respectively. Patients with isolated 25(OH)D deficiency and rickets who respond to pharmacologic doses of vitamin D and physiologic doses of 25(OH)D3, 1alpha(OH)D3, or related drugs are presumed to have CYP25 deficiency.
The specific aims of this proposal are to use portions of porcine CYP25 cDNA to isolate and characterize human CYP25 cDNA, to clone and characterize the human CYP25 gene, to determine chromosome locus of CYP25 DNA and tissue expression of CYP25 mRNA, to use gene mapping, linkage analysis and positional cloning, to characterize family pedigrees and determine molecular mechanisms of patients with isolated 25(OH)D deficiency and rickets, to characterize CYP25 gene polymorphisms in affected populations, and to correlate genotype with phenotype. The fact that porcine CYP25 cDNA has 83 percent homology with human CYP2D6 cDNA, a cytochrome P450 microsomal enzyme that plays a role in drug metabolism, has interfered with cloning of human CYP25 cDNA. Pig liver extracts are reported to have no CYP2DK activity. Using a radiation hybrid cell panel (ImpRH), we found that the porcine CYP25 gene maps to pig chromosome 5p14-15 close to the ACO2 gene, a region syntenic ot human chromosome 22q12-13 where human CYP2D6 was mapped. We used haplotype analysis of family pedigrees of patients with isolated CYP25 deficiency and rickets and found that the disease does not map to either the CYP2D6 gene locus or chromosome 22. The results of the proposed studies should provide important new information about the molecular structure and chromosomal locus of the CYP25 gene and the molecular causes of isolated 25(OH)D and rickets.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Research Project (R01)
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General Medicine B Study Section (GMB)
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Malozowski, Saul N
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Medical University of South Carolina
Internal Medicine/Medicine
Schools of Medicine
United States
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