A bovine model of rhizomelic chondrodysplasia punctata caused by a deep intronic splicing variant in the GNPAT gene.

BACKGROUND: Genetic defects that occur naturally in livestock species provide valuable models for investigating the molecular mechanisms underlying rare human diseases. Livestock breeds are subject to the regular emergence of recessive genetic defects due to genetic drift and recent inbreeding. At the same time, their large population sizes provide easy access to case and control individuals and to massive amounts of pedigree, genomic and phenotypic information recorded for management and selection purposes. In this study, we investigated a lethal form of recessive chondrodysplasia observed in 21 stillborn calves of the Aubrac beef cattle breed. RESULTS: Detailed examinations of three affected calves revealed proximal limb shortening, epiphyseal calcific deposits, and other pathological signs consistent with human rhizomelic chondrodysplasia punctata, a rare peroxisomal disorder caused by recessive variants in one of five genes (AGPS, FAR1, GNPAT, PEX5, and PEX7). Using homozygosity mapping, whole genome sequencing of two affected individuals, and filtering for variants found in 1867 control genomes, we reduced the list of candidate variants to a single deep intronic substitution in GNPAT (NC_037355.1:g.4039268G > A on chromosome 28 of the ARS-UCD1.2 bovine genome assembly). For verification, we performed large-scale genotyping of this variant using a custom SNP array and found a perfect genotype-phenotype correlation in 21 cases and 26 of their parents, and a complete absence of homozygotes in 1195 unaffected Aubrac controls. The g.4039268A allele segregated at a frequency of 2.6% in this population and was absent in 375,535 additional individuals from 17 breeds. Then, using in vivo and in vitro analyses, we demonstrated that the derived allele activates cryptic splice sites within intron 11 resulting in abnormal transcripts. Finally, by mining the wealth of records available in the French bovine database, we also reported suggestive effects on juvenile mortality (and not just stillbirth) in homozygotes and on muscle development in heterozygotes, which merit further investigation. CONCLUSIONS: We report the first spontaneous large animal model of rhizomelic chondrodysplasia punctata and provide a diagnostic test to select against this defect in cattle. Our work also brings interesting insights into the molecular consequences of complete or partial GNPAT insufficiency in mammals.