Finding the mutation responsible for inherited rickets in Corriedale sheep

Abstract

Inherited rickets of Corriedale sheep is of simple autosomal recessive inheritance. The gross and histological lesions include segmental thickening of physes, persistence of hypertrophic chondrocytes at sites of endochondral ossification, excessive osteoclastic resorption, microfractures and wide unmineralised osteoid seams lining trabeculae and filling secondary osteons. Initial testing revealed concentrations of 25-hydroxyvitamin D3 were normal, but those of 1,25-dihydroxyvitamin D3 were increased in serum of affected sheep, suggesting a defect in the vitamin D receptor. However, in vitro studies on cultured skin fibroblasts revealed normal binding and function of the vitamin D receptor. A genome-wide association study was conducted using the Illumina OvineSNP50 BeadChip. A homozygous region of 199 consecutive single-nucleotide polymorphism loci was identified in affected sheep, covering a region of 10 megabase pairs on ovine chromosome 6. Of the 91 candidate genes in this region, dentin matrix protein 1 gene (dmp1) was considered the most likely as it is known to play critical roles in osteocyte maturation and mineral metabolism. Sequencing of DNA from carriers showed a nonsense mutation 253T/C in dmp1. This T/C transition introduced a stop codon (R145X) that could truncate C-terminal amino acids. Using PCR restriction fragment length polymorphism for this mutation, genotyping showed that all affected sheep were “TT” genotypes, and phenotypically normal sheep were either “CT” or “CC”. These findings suggest that a mutation in dmp1 is responsible for inherited rickets of Corriedale sheep. A simple diagnostic test is currently being designed to identify carriers with the defective “T” alleles.