Novel gene mutations in patients with 1alpha-hydroxylase deficiency that confer partial enzyme activity in vitro.

Abstract

The rate-limiting, hormonally regulated step in the biological activation of vitamin D is its 1alpha-hydroxylation to 1,25-dihydroxyvitamin D [1,25-(OH)(2)D] in the kidney, catalyzed by the mitochondrial cytochrome P450 enzyme, P450c1alpha. We previously cloned the human P450c1alpha cDNA and gene, and identified 14 different mutations, including 7 missense, in 19 patients with 1alpha-hydroxylase deficiency, also known as vitamin D-dependent rickets type 1. None of the missense mutations encoded a protein with detectable enzymatic activity in vitro. Although there is phenotypic variation among such patients, the molecular basis of this variation is unknown. We analyzed 6 additional patients with clinical and radiographic features of rickets; in 4 patients the laboratory abnormalities were typical of 1alpha-hydroxylase deficiency, but in 2 they were unusually mild [mild hypocalcemia and normal serum 1,25-(OH)(2)D concentration]. Direct sequencing revealed that all patients had P450c1alpha mutations on both alleles. Five new and 2 known mutations were identified. The new mutations included a 5-bp deletion with a 6-bp novel insertion causing a frameshift in exon 2, and a G to A change at +1 of intron 2; a minigene experiment proved that this intronic mutation prevented proper splicing. Three new missense mutations were found and tested by expressing the mutant cDNA in mouse Leydig MA-10 cells. The R389G mutant was totally inactive, but mutant L343F retained 2.3% of wild-type activity, and mutant E189G retained 22% of wild-type activity. The two mutations that confer partial enzyme activity in vitro were found in the 2 patents with mild laboratory abnormalities, suggesting that such mutations contribute to the phenotypic variation observed in patients with 1alpha-hydroxylase deficiency.

• Authors: Xuemei Wang, Martin Y H Zhang, Walter L Miller, Anthony A Portale
• Journal: The Journal of clinical endocrinology and metabolism (J Clin Endocrinol Metab) Vol. 87 Issue 6 Pg. 2424-30 (Jun 2002) ISSN: 0021-972X [Print] United States
• PMID: 12050193 (Publication Type: Case Reports, Journal Article, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, P.H.S.)

Chemical References

• DNA, Complementary
• 25-Hydroxyvitamin D3 1-alpha-Hydroxylase

Topics

• 25-Hydroxyvitamin D3 1-alpha-Hydroxylase (genetics, metabolism)
• Animals
• Base Sequence (genetics)
• Cell Line
• DNA, Complementary (genetics)
• Female
• Gene Expression
• Humans
• Mice
• Molecular Sequence Data
• Mutation (physiology)
• Mutation, Missense (physiology)
• Vitamin D Deficiency (genetics)