Identification of the bovine Arachnomelia mutation by massively parallel sequencing implicates sulfite oxidase (SUOX) in bone development

Abstract

Arachnomelia is a monogenic recessive defect of skeletal development in cattle. The causative mutation was previously mapped to a ∼7 Mb interval on chromosome 5. Here we show that array-based sequence capture and massively parallel sequencing technology, combined with the typical family structure in livestock populations, facilitates the identification of the causative mutation. We re-sequenced the entire critical interval in a healthy partially inbred cow carrying one copy of the critical chromosome segment in its ancestral state and one copy of the same segment with the arachnomelia mutation, and we detected a single heterozygous position. The genetic makeup of several partially inbred cattle provides extremely strong support for the causality of this mutation. The mutation represents a single base insertion leading to a premature stop codon in the coding sequence of the SUOX gene and is perfectly associated with the arachnomelia phenotype. Our findings suggest an important role for sulfite oxidase in bone development.

Figure 1. Phenotype of bovine arachnomelia in Brown Swiss cattle.

(A,B) Stillborn affected calves; note the abnormal length of all legs (dolichostenomelia), the angular deformities in the distal part of the hind legs with arthrogryposis of the distal joints, and a muscular atrophy of the legs. (C,D) Typical facial deformities of affected calves; note a concave rounding of the dorsal profile of the maxilla and brachygnathia inferior. (E) Radiography of the hind limbs of the affected calf from (C); note that the joint ends (epiphyses) of the long bones are of normal size but with marked thinning of the shafts (diaphyses) showing increased fragility. (F) Radiography of the left hind leg of a non-affected control calf.

Figure 2. Pedigree of selected inbred Brown Swiss cattle.

The special structure of livestock families facilitates the mutation identification: The bull Lilason (arrow) is the founder animal for arachnomelia. Thus the arachnomelia mutation (indicated by an A) must have occurred either in the germline of Larry or during the early embryonic development of Lilason. Beneath the animals the two copies of BTA 5 are indicated. Black symbols represent the ancestral chromosome, on which the mutation occurred. Gray symbols represent any other wild-type copy of BTA 5. Due to the inbreeding loop, the cow Valbella has inherited two copies of the same chromosome from her ancestor Larry. Her paternal copy of the critical 7.19 Mb interval carries the arachnomelia mutation, while her maternal copy is still in its ancestral wild-type state. The BTA 5 haplotypes were confirmed by microsatellite genotyping of available samples from Beautician, Leon, Amaranto, Elegant, Collection, and Valbella .

Figure 3. Mutation identification in the bovine SUOX gene.

Electropherograms of a wild-type control animal (A), a carrier cow (B), and an arachnomelia affected calf (C) are shown. A single G insertion, which was homozygous in all analyzed cases, is marked by an arrow. The protein translation of the wild-type and mutant sequence is shown above the electropherograms.