Ovine KRTAP36-2: A New Keratin-Associated Protein Gene Related to Variation in Wool Yield.

Keratin-associated proteins (KAPs) are structural components of wool fibres. High-glycine/tyrosine (HGT)-KAPs are a subset of the KAP family, and their abundance in fibres varies. In this study, we report the discovery of an ovine HGT-KAP gene to which we assigned the name KRTAP36-2. Polymerase chain reaction and single-strand conformation polymorphism (PCR-SSCP) analyses revealed four variants of this gene in a screening population of 170 sheep from a variety of breeds. The DNA sequencing of the variants revealed four single-nucleotide polymorphisms (SNPs) and a dinucleotide deletion. Three of these SNPs were in the coding region, and one of these was non-synonymous and potentially led to the amino acid substitution p.Cys27Gly near the middle of the protein. The remaining SNP was located near the putative TATA box, and the di-nucleotide deletion was near the putative transcription initiation site. The effect of this variation in KRTAP36-2 was investigated in 274 Southdown × Merino lambs that were the progeny of five sires. Variation was only found to be associated with wool yield, that is, the proportion of the greasy fleece that remained as clean fleece upon scouring (expressed as a percentage). This may have some value in increasing wool production.

Variation in Ovine DGAT1 and Its Association with Carcass Muscle Traits in Southdown Sheep.

Diacylglycerol O-acyltransferase 1 (DGAT1) is a microsomal enzyme that plays a key role in the synthesis of triglycerides. Its gene (DGAT1) is regarded as a candidate gene for variation in milk and meat traits in cattle. The objective of this study was to use a PCR single-strand conformation polymorphism approach to explore sequence variation in two regions of ovine DGAT1 and to assess its effect on meat traits in New Zealand Southdown sheep. Three variant nucleotide sequences were identified in each region, with two single nucleotide polymorphisms (SNPs) and one nucleotide deletion being detected in intron 1 and two SNPs being found in exon 17. The effect of the exon 17 variation was not investigated due to one variant being predominant and the other two variants occurring at low frequencies. In intron 1, one variant (B1) was found to be associated with increase loin meat yield, suggesting that this may have value as a gene marker for improving meat traits.

A Preliminary Investigation of Myostatin Gene (MSTN) Variation in Red Deer (Cervus elaphus) and Its Implications for Venison Production in New Zealand.

Myostatin (MSTN), also known as growth differentiation factor 8 (GDF-8), is a negative regulator of lean muscle tissue growth. Variation in the gene has been studied in many domesticated species, because of its potential to dramatically increase muscle mass. It has, however, not been investigated in red deer (Cervus elaphus). In this study, variation in MSTN intron 1 was investigated in 211 male New Zealand red deer, for which phenotypic measurements of M. Longissimus dorsi (eye muscle) (width, depth, and area, together with 12-month weight) were recorded. Two sequence variants (named A and B) differing by one nucleotide (c.373 + 224) were identified in the intron 1 region of the gene resulting in three genotypes (AA, AB, and BB; frequencies of 63.5%, 30.8%, and 5.7%, respectively), but no association between this variation and any of the quantitative measurements was detected. These results suggest that the deer MSTN is less variable than for other livestock species and that its activity may be controlled to maintain a size-growth equilibrium.

Ovine Toll-like Receptor 9 (TLR9) Gene Variation and Its Association with Flystrike Susceptibility.

Toll-like receptors (TLRs) are a family of proteins that play a role in innate immune responses by recognising pathogen-associated molecular patterns derived from various microbes. Of these receptors, TLR9 recognises bacterial and viral DNA containing unmethylated cytosine-phosphate-guanine (CpG) motifs, and variation in TLR9 has been associated with resistance to various infectious diseases. Flystrike is a problem affecting the sheep industry globally and the immune response of the sheep has been suggested as one factor that influences the response to the disease. In this study, variation in ovine TLR9 from 178 sheep with flystrike and 134 sheep without flystrike was investigated using a polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) approach. These sheep were collected from both commercial and stud farms throughout New Zealand and they were of 13 different breeds, cross-breds and composites. Four alleles of TLR9 were detected, including three previously identified alleles (*01, *02 and *03) and a new allele (*04). In total six single nucleotide polymorphisms (SNPs) were found. Of the three common alleles in the sheep studied, the presence of *03 was found to be associated with a reduced likelihood of flystrike being present (OR = 0.499, p = 0.024). This suggests that variation in ovine TLR9 may affect a sheep's response to flystrike, and thus the gene may have value as a genetic marker for improving resistance to the disease.

The Complexity of the Ovine and Caprine Keratin-Associated Protein Genes.

Sheep (Ovis aries) and goats (Capra hircus) have, for more than a millennia, been a source of fibres for human use, be it for use in clothing and furnishings, for insulation, for decorative and ceremonial purposes, or for combinations thereof. While use of these natural fibres has in some respects been superseded by the use of synthetic and plant-based fibres, increased accounting for the carbon and water footprint of these fibres is creating a re-emergence of interest in fibres derived from sheep and goats. The keratin-associated proteins (KAPs) are structural components of wool and hair fibres, where they form a matrix that cross-links with the keratin intermediate filaments (KIFs), the other main structural component of the fibres. Since the first report of a complete KAP protein sequence in the late 1960s, considerable effort has been made to identify the KAP proteins and their genes in mammals, and to ascertain how these genes and proteins control fibre growth and characteristics. This effort is ongoing, with more and more being understood about the structure and function of the genes. This review consolidates that knowledge and suggests future directions for research to further our understanding.

Sequence Variation in the Bovine Lipin-1 Gene (LPIN1) and Its Association with Milk Fat and Protein Contents in New Zealand Holstein-Friesian × Jersey (HF × J)-cross Dairy Cows.

Lipin-1 is known to play a regulatory role in tissues that function in lipid metabolism. In dairy cows, the lipin-1 gene (LPIN1) is highly expressed in the mammary gland, but its function in milk production is less understood. In this study, we used PCR-single strand conformation polymorphism analysis to investigate sequence variation in three regions of bovine LPIN1 in New Zealand Holstein-Friesian × Jersey (HF × J)-cross dairy cows, including part of the 5' non-coding region, the region containing the LPIN1ß-spliced exon, and the sixth coding exon that encodes the putative transcriptional activating domain of the protein. No variation was found in the LPIN1ß-spliced exon, but two sequence variants containing one single nucleotide polymorphism (SNP) were identified in the 5' non-coding region and four sequence variants containing four non-synonymous SNPs were identified in the sixth coding exon. Among the three common variants of the sixth coding exon, variant C was found to be associated with an increase in milk fat percentage (presence 4.96 ± 0.034% vs. absence 4.81 ± 0.050%; p = 0.006) and milk protein percentage (presence 4.09 ± 0.017% vs. absence 3.99 ± 0.025%; p = 0.001), but no associations (p > 0.01) were detected for milk yield. These results suggest that variation in LPIN1 affect the synthesis of fat and proteins in milk and has potential as a gene-marker to improve milk production traits.

Identification of the association between FABP4 gene polymorphisms and milk production traits in Sfakia sheep.

The aim of this study was to estimate the effect of variation in the fatty acid binding protein 4 gene (FABP4) on milk production traits in Greek Sfakia sheep. Polymerase chain reaction - single-stranded conformational polymorphism (PCR-SSCP) analysis was used to genotype a total of 374 Sfakia ewes for two regions of FABP4 located around exon 2-intron 2 (Region 1) and exon 3-intron 3 (Region 2). Each month, for a period of 6 months, milk samples were collected from the ewes to measure total milk yield, fat content, protein content, lactose content, non-fat solid content, pH, and somatic cell count (SCC). A general linear model was used to test the association between the variation observed in FABP4 and milk production traits. Four gene variants (A1-A4) were found in Region 1 and two variants (C1-C2) were found in Region 2. In the first region, the FABP4 genotype significantly affected ( P < 0.05 ) non-fat solid levels, fat content, and SCC. The presence of the A2 variant was significantly associated ( P < 0.05 ) with decreased SCC, while the presence of A4 was significantly associated with decreased milk yield ( P < 0.01 ), increased non-fat solid content ( P < 0.05 ), decreased fat content ( P < 0.01 ), increased lactose content ( P < 0.05 ), and increased pH ( P < 0.05 ). In the second region, FABP4 genotype had an effect ( P < 0.05 ) on protein content and the presence of the C2 variant was associated ( P < 0.05 ) with increased protein content, decreased SCC, and lower pH. The results suggest an association between variation in ovine FABP4 and milk production traits in Greek Sfakia sheep. Nevertheless, further analyses in independent sheep populations of increased size will strengthen these findings.

Characterisation of an Ovine Keratin Associated Protein (KAP) Gene, Which Would Produce a Protein Rich in Glycine and Tyrosine, but Lacking in Cysteine.

The keratin-associated proteins (KAPs) are structural components of hair/wool fibres. All of the KAPs identified to date contain cysteine, which is thought to form disulphide bonds cross-linking the keratin intermediate filaments. Here, we report the identification of a KAP gene in sheep that would produce a protein that contains a high proportion (63.2 mol%) of glycine and tyrosine, but would not contain any cysteine. This suggests that other forms of intra- and inter-strand interaction may occur with this KAP, such as interactions via ring-stacking and hydrogen-bonding. The gene was dissimilar to any previously reported KAP gene, and was therefore assigned to a new family, and named KRTAP36-1. The KRTAP36-1 genome sequence was almost identical to some EST sequences from sheep and goat skin follicles, suggesting that it is present and expressed in sheep and goats. A BLAST search of the human genome assembly sequence did not reveal any human homologue. Three variant sequences (named A to C) of ovine KRTAP36-1 were identified and four single nucleotide polymorphisms (SNPs) were detected. One SNP was located 32 bp upstream of the coding region, and all of the others were in the coding region and were nonsynonymous. After correcting for potential linkage to the proximal KRTAP20-1, variant B of KRTAP36-1 was found to be associated with increased prickle factor (PF) in wool, suggesting that variation in the gene may have the potential to be used as gene marker for breeding sheep with lower PF.

An updated nomenclature for keratin-associated proteins (KAPs).

Most protein in hair and wool is of two broad types: keratin intermediate filament-forming proteins (commonly known as keratins) and keratin-associated proteins (KAPs). Keratin nomenclature was reviewed in 2006, but the KAP nomenclature has not been revised since 1993. Recently there has been an increase in the number of KAP genes (KRTAPs) identified in humans and other species, and increasingly reports of variation in these genes. We therefore propose that an updated naming system is needed to accommodate the complexity of the KAPs. It is proposed that the system is founded in the previous nomenclature, but with the abbreviation sp-KAPm-nL*x for KAP proteins and sp-KRTAPm-n(p/L)*x for KAP genes. In this system "sp" is a unique letter-based code for different species as described by the protein knowledge-based UniProt. "m" is a number identifying the gene or protein family, "n" is a constituent member of that family, "p" signifies a pseudogene if present, "L" if present signifies "like" and refers to a temporary "place-holder" until the family is confirmed and "x" signifies a genetic variant or allele. We support the use of non-italicised text for the proteins and italicised text for the genes. This nomenclature is not that different to the existing system, but it includes species information and also describes genetic variation if identified, and hence is more informative. For example, GenBank sequence JN091630 would historically have been named KRTAP7-1 for the gene and KAP7-1 for the protein, but with the proposed nomenclature would be SHEEP-KRTAP7-1*A and SHEEP-KAP7-1*A for the gene and protein respectively. This nomenclature will facilitate more efficient storage and retrieval of data and define a common language for the KAP proteins and genes from all mammalian species.