Identification of the keratin-associated protein 13-3 (KAP13-3) gene in sheep

Abstract

Keratin-associated proteins (KAPs) are a major stru- ctural component of hair and wool fibres, and play a critical role in determining the properties of the fibre. To date, forty functional high sulphur KAP genes from fourteen families have been identified in hu- mans, but only seven functional high sulphur KAP genes have been identified in sheep. This led us to search for the ovine KAP13-3 gene, a gene encoding a high sulphur KAP. In this study, the notional KAP13- 3 gene (KRTAP13-3) was amplified using primers designed based on a reported bovine KRTAP13-3 se- quence. PCR-single stranded conformational poly- morphism (PCR-SSCP) analysis was used to screen amplicons derived from the gene in one hundred and forty seven New Zealand Romney cross-bred sheep. Five unique banding patterns were revealed. Either one PCR-SSCP pattern (homozygous) or a combina- tion of two patterns (heterozygous) was observed for each sheep. Sequencing of PCR amplicons represent- tative of different SSCP patterns revealed five dif- ferent DNA sequences. The sequences derived from the amplicons showed a low homology to other known ovine KRTAPs, but had a high homology with previous reported KRTAP13-n sequences from hu- man and cattle, with the closest homology being with bovine KRTAP13-3, suggesting the sequences repre- sent the ovine KRTAP13-3 locus. Among the five allele sequences, four nucleotide substitutions were identi- fied within the coding region. Of these substitutions, three were non-synonymous and would result in amino acid changes (p.Arg79Cys, p.Arg81Gln and p.Tyr130His). This variation in the KAP13-3 gene may affect gene expression, the structure and assem- bly of the protein, and consequently influence wool traits, if KAP13-3 is of importance to wool fibre structure.