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ARS Home » Midwest Area » Ames, Iowa » National Animal Disease Center » Ruminant Diseases and Immunology Research » Research » Publications at this Location » Publication #119168
Title: NOVEL MUTATIONS IN THE 25-HYDROXYVITAMIN D3 1-ALPHA-HYDROXYLASE GENE IN PATIENTS WITH VITAMIN D-DEPENDENT RICKETS TYPE 1

Author
item HUANG, D - MGGILL UNIV., MONTREAL
item HORST, RONALD
item RHIM, J - UNIFORM SRV, ROCKVILLE MD
item KREMER, R - MCGILL UNIV, MONTREAL

Submitted to: American Society for Bone and Mineral Research
Publication Type: Abstract Only
Publication Acceptance Date: 9/22/2000
Publication Date: N/A
Citation: N/A

Interpretive Summary:

Technical Abstract: It was recently established that Vitamin D-Dependent Rickets Type 1 (VDDR- 1) is caused by inactivating mutations in the 25-hydroxyvitamin D3 1- alpha-hydroxylase (1a-hydroxylase) gene. In this study, we cloned the 25- hydroxyvitamin D3 1a-hydroxylase gene from two French-Canadian patients with VDDR-1. Nucleotide sequence analysis revealed that both patients were compound heterozygous for the 1a-hydroxylase gene. Patient 1 had frameshift mutations on exon 1 at codon E63G and on exon 9 at codon 1501T. Patient 2 had frameshift mutations on exon 1 at codon S32T and on exon 9 at colon P482S. In order to analyze the effect of these mutations on 1a- hydroxylase activity, we then performed site directed mutagenesis on a human 25-hydroxyvitamin D3 1a-hydroxylase cDNA cloned from normal human keratinocytes into the pcDNA3 vector. In this vector cloned wild-type and mutants 1a-hydroxylase cDNAs are driven by a strong CMV promoter. All four cDNA mutants were then transiently transfected in an established normal human renal proximal tubule epithelial cell line incubated with cold 25-hydroxyvitamin D3 and compared to wild-type 1a-hydroxylase cDNA, 1a-hydroxylase activity measured in the conditioned media indicated a strong, although variable, reduction in activity depending on the location of the mutation. Our results, therefore, indicate that 1a-hydroxylase inactivating mutations are expressed with variable phenotypes.