A Novel Nonsense Mutation in the <em>DMP1</em> Gene Identified by a Genome-Wide Association Study Is Responsible for Inherited Rickets in Corriedale Sheep

Inherited rickets of Corriedale sheep is characterized by decreased growth rate, thoracic lordosis and angular limb deformities. Previous outcross and backcross studies implicate inheritance as a simple autosomal recessive disorder. A genome wide association study was conducted using the Illumina OvineSNP50 BeadChip on 20 related sheep comprising 17 affected and 3 carriers. A homozygous region of 125 consecutive single-nucleotide polymorphism (SNP) loci was identified in all affected sheep, covering a region of 6 Mb on ovine chromosome 6. Among 35 candidate genes in this region, the dentin matrix protein 1 gene (DMP1) was sequenced to reveal a nonsense mutation 250C/T on exon 6. This mutation introduced a stop codon (R145X) and could truncate C-terminal amino acids. Genotyping by PCR-RFLP for this mutation showed all 17 affected sheep were “T T” genotypes; the 3 carriers were “C T”; 24 phenotypically normal related sheep were either “C T” or “C C”; and 46 unrelated normal control sheep from other breeds were all “C C”. The other SNPs in DMP1 were not concordant with the disease and can all be ruled out as candidates. Previous research has shown that mutations in the DMP1 gene are responsible for autosomal recessive hypophosphatemic rickets in humans. Dmp1_knockout mice exhibit rickets phenotypes. We believe the R145X mutation to be responsible for the inherited rickets found in Corriedale sheep. A simple diagnostic test can be designed to identify carriers with the defective “T” allele. Affected sheep could be used as animal models for this form of human rickets, and for further investigation of the role of DMP1 in phosphate homeostasis.考力代绵羊的遗传性佝偻病以生长速率降低、胸椎前凸及肢端成角畸形为特征。先前的异交与回交研究表明，该疾病属于简单的常染色体隐性遗传病。本研究使用Illumina OvineSNP50基因芯片，对20只相关绵羊开展全基因组关联研究，其中包含17只患病个体与3只携带者。在所有患病绵羊中，均鉴定出一段包含125个连续单核苷酸多态性（single-nucleotide polymorphism, SNP）位点的纯合区域，该区域位于绵羊6号染色体上，跨度6 Mb。该区域内共有35个候选基因，对牙本质基质蛋白1基因（dentin matrix protein 1 gene, DMP1）进行测序后，发现其第6外显子存在一处250C>T的无义突变。该突变引入了终止密码子R145X，可导致C端氨基酸序列截短。通过聚合酶链反应-限制性片段长度多态性（PCR-RFLP）对该突变进行基因分型的结果显示：17只患病绵羊均为T/T纯合基因型；3只携带者均为C/T杂合基因型；24只表型正常的相关绵羊为C/T或C/C基因型；46只来自其他品种的无亲缘关系正常对照绵羊均为C/C基因型。DMP1基因内的其他SNP位点与疾病表型不共分离，均可排除作为致病候选位点。既往研究表明，人类DMP1基因突变可导致常染色体隐性低血磷性佝偻病；Dmp1基因敲除小鼠可表现出佝偻病表型。本研究认为，R145X突变是考力代绵羊遗传性佝偻病的致病原因。可设计一种简便的诊断检测方法，以鉴定携带致病T等位基因的携带者。患病绵羊可作为该型人类佝偻病的动物模型，也可用于进一步研究DMP1在磷酸盐稳态调控中的作用。