A newly described bovine type 2 scurs syndrome segregates with a frame-shift mutation in TWIST1

Abstract

The developmental pathways involved in horn development are complex and still poorly understood. Here we report the description of a new dominant inherited syndrome in the bovine Charolais breed that we have named type 2 scurs. Clinical examination revealed that, despite a strong phenotypic variability, all affected individuals show both horn abnormalities similar to classical scurs phenotype and skull interfrontal suture synostosis. Based on a genome-wide linkage analysis using Illumina BovineSNP50 BeadChip genotyping data from 57 half-sib and full-sib progeny, this locus was mapped to a 1.7 Mb interval on bovine chromosome 4. Within this region, the TWIST1 gene encoding a transcription factor was considered as a strong candidate gene since its haploinsufficiency is responsible for the human Saethre-Chotzen syndrome, characterized by skull coronal suture synostosis. Sequencing of the TWIST1 gene identified a c.148_157dup (p.A56RfsX87) frame-shift mutation predicted to completely inactivate this gene. Genotyping 17 scurred and 20 horned founders of our pedigree as well as 48 unrelated horned controls revealed a perfect association between this mutation and the type 2 scurs phenotype. Subsequent genotyping of 32 individuals born from heterozygous parents showed that homozygous mutated progeny are completely absent, which is consistent with the embryonic lethality reported in Drosophila and mouse suffering from TWIST1 complete insufficiency. Finally, data from previous studies on model species and a fine description of type 2 scurs symptoms allowed us to propose different mechanisms to explain the features of this syndrome. In conclusion, this first report on the identification of a potential causal mutation affecting horn development in cattle offers a unique opportunity to better understand horn ontogenesis.

Figure 1. Phenotypic description of type 2 scurs syndrome.

Ten-(A), two-(B) and four-(C) year old females affected by type 2 scurs syndrome. Note the marked phenotypic variability and the negative correlation between the size of scurs and the importance of acrocephaly and ridge-shaped extra bone deposition along the interfrontal suture. (D). Four-year old horned female.

Figure 2. Details of the type 2 scurs keratin sheath.

(A). Scaly patches. (B). Small scurs (∼2 cm) with irregular keratin sheath. (C and D). Long scurs (∼15 cm) with an irregular keratin sheath at their end (see arrows). (E). End of a normal horn (∼25 cm) with a regular keratin sheath.

Figure 3. Frontal radiographs of type 2 scurred and horned skulls.

(A). Control skull of a five-year old horned female. (B). Skull of a four-year old female affected by the type 2 scurs and carrying long scurs. Note: (1) the particular shape of the poll, (2) the denser calcification of the interfrontal suture, (3) the absence of fusion between the frontal bone and the horn bony core, (4) the absence of frontal bone drawing up and (5) the non-pneumatization of the bony core in affected vs. horned skulls.

Figure 4. Genome-wide multipoint linkage analysis of type 2 scurs syndrome.

The genome-wide scan reveals a significant linkage (maximum LOD score = 7.2) with clusters of markers located on chromosome 4. The 95% confidence interval spans 1.7 Mb encompassing six different genes.

Figure 5. Characterization of the TWIST1 c.148_157dup mutation.

(A) TWIST1 gene's organization scheme; (B) DNA sequencing chromatograms showing the c.148_157dup mutation of type 2 scurs affected animals and the wild type allele; (C) putative p.A56RfsX87 mutated protein and multispecies alignment of the TWIST1 protein sequence using CLUSTALW . Cattle (Bta), human (Hsa), mouse (Mmu), chicken (Gga), anolis (Aca), xenopus (Xtr) and zebrafish (Dre) sequences accession numbers are respectively DAA30767, NP_000465, AAH33434, NP_990070, DAA06059, AAH74558, ABC73066 in Genbank. The frame-shift change in the bovine p.A56RfsX87 mutated protein (Mut.) is underlined. The basic, helix-loop-helix and tryptophan–arginine domains are respectively highlighted with dashed, solid and mixed dashed boxes .