Genome-wide association study identifies loci linked to serum electrolyte traits in Chinese Holstein cattle.

We aimed to identify QTL for serum electrolyte traits by performing a GWAS of calcium, chloride, sodium, potassium and magnesium ion concentrations in Chinese Holstein cattle. We detected five SNPs that had significant associations with calcium ion concentrations and identified GATA2 from Bos taurus chromosome (BTA)22 as having the highest significance. Among the genes with significant results, we speculate that TMEM123 might be related to calcium channel proteins according to the functions of the TMEM family.

Investigation of TG gene variants and their effects on growth, carcass composition, and meat quality traits in Chinese steers.

Thyroid hormones play an important role in regulating metabolism and can affect metabolism-related traits such as fat deposition. The thyroglobulin (TG) gene produces the precursor of thyroid hormones and has been proposed as a candidate gene for a quantitative trait locus with an effect on fat deposition. In this study, we identified 4 novel single nucleotide polymorphisms (SNPs) in the 5' flanking region of the TG gene using a DNA sequencing method. The SNP marker association analysis indicated that the T1355C SNPs were significantly associated with meat percentage (P < 0.05). A significant association between the G1356A polymorphism and live weight and loin muscle area was also detected (P < 0.05). However, no significant association was found between 4 SNPs and the other growth, carcass composition, and meat quality traits including intramuscular fat. The results of this study suggest that TG gene-specific SNPs may be a useful marker for growth traits in marker-assisted selection programs in beef cattle.

Polymorphism of MyoD1 and Myf6 genes and associations with carcass and meat quality traits in beef cattle.

Myogenic determination factor 1 (MyoD1) and myogenic factor 6 (Myf6) genes belong to the myogenic differentiation (MyoD) gene family, which play key roles in growth and muscle development. The study aimed to investigate the effects of variants in cattle MyoD1 and Myf6 on carcass and meat traits. We screened single nucleotide polymorphisms (SNPs) of both genes in 8 cattle populations, including Simmental, Angus, Hereford, Charolais, Limousin, Qinchuan, Luxi, and Jinnan by sequencing. The G782A locus was identified in exon 1 of MyoD1 (MyoD1-BglI) as well as the T186C locus in exon 1 of Myf6 (Myf6-ApaLI). For the two SNPs, the A allele was significantly more frequent than the B allele in the populations tested. The χ(2) test showed that the MyoD1-BglI locus conformed to Hardy-Weinberg equilibrium in the 8 populations, as did the Myf6-ApaLI locus, with the exception of the Simmental population (P > 0.05). Association analysis revealed that the MyoD1-BglI locus was significantly associated with loin muscle area (LMA) (P < 0.05), and the Myf6-ApaLI locus was significantly associated with carcass length (CL) (P < 0.05). Animals with BB and AB genotypes for the MyoD1-BglI locus had larger LMAs compared to animals with AA genotype. Individuals with BB genotype had longer CLs compared to those with AA and AB genotypes. We conclude that the two SNPs might provide useful genetic markers, opening up new possibilities for cattle breeding and improvements in gene-assisted selection.

Detecting a deletion in the coding region of the bovine bone morphogenetic protein 15 gene (BMP15).

The aim of this study was to detect polymorphism in the bovine bone morphogenetic protein 15 (BMP15) gene. On the basis of PCR-SSCP and DNA sequencing, a 4-bp deletion was identified in the coding region of the gene. Sequence analysis revealed that the deletion altered the reading frame and introduced a stop codon at position 264. Eight breeds (Luxi, Qinchuan, Nanyang, Jinnan, Bohai Black, Menggolian, Holstein, and Simmental) were genotyped by PCR-SSCP. No cows homozygous for this mutation were observed in these breeds. Heterozygous cows were detected in Luxi, Qinchuan, Nanyang, Jinnan and Bohai Black cattle. Fecundity was not increased in heterozygous individuals.

Toll-like receptor 2 gene polymorphism and its relationship with SCS in dairy cattle.

Toll-like receptor 2 (TLR2) plays an important role in the innate immune response to a variety of pathogens. In this study, bovine TLR2 gene was taken as a candidate gene for mastitis resistance. Through PCR-SSCP analysis and sequencing, three missense mutations at T385 G, G398A, and G1884A were detected in the coding region that encoded extracellular domain. Altogether 240 dairy cattle of three breeds (Holstein, Simmental, and Sanhe cattle) were genotyped and allele frequencies were determined. The effects of TLR2 polymorphisms on somatic cell score (SCS) were analyzed and significant association was found between T385 G and SCS. The mean of genotype GG was significantly lower than those of genotype TT and TG. No significant associations were found with SCS for G398A and G1884A. Information provided in this research will be useful in further studies to determine the role of TLR2 gene in the mastitis resistance.

Identification of a specific methionine in mammalian 15-lipoxygenase which is oxygenated by the enzyme product 13-HPODE: dissociation of sulfoxide formation from self-inactivation.

Mammalian 15-lipoxygenases undergo a characteristic self-inactivation. The oxygenation of a single methionine to methionine sulfoxide, by 13(S)-hydroperoxyoctadecadienoic acid (13-HPODE), was previously suggested as the cause of the inactivation of rabbit reticulocyte lipoxygenase. The site of oxygenation is potentially near the enzyme's active site; however, the specific location of the modified amino acid residue has not been identified. To determine which of the methionine residues is oxygenated, we inactivated both human and rabbit 15-lipoxygenases with 13-HPODE and sequentially denatured, reduced, carboxymethylated, and digested the enzymes with trypsin. The digested mixtures were analyzed by reverse-phase HPLC chromatography. Mass spectrometric analysis of each of the methionine-containing fractions enabled us to locate the peptide segments containing the oxidized methionine in both enzymes separately. Tandem electrospray mass spectrometry identified the oxidized methionine residues to be amino acid 590 in the human enzyme and 591 in the rabbit enzyme. To investigate the significance of this oxygenation, Met590 in human 15-lipoxygenase was substituted with leucine by site-directed mutagenesis. The mutant protein was inactivated by 13-HPODE, yet no oxygenated peptide or other modified peptide could be identified by HPLC-MS analysis. We also found that human 15-lipoxygenase was inactivated during arachidonate oxidation and by the reaction product 15(S)-hydroperoxyeicosatetraenoic acid (15-HPETE), and no modified peptide was detected. Thus, methionine oxygenation is not essential for the inactivation of human 15-lipoxygenase. We suggest, however, that Met590 is an amino acid in the substrate binding pocket of human 15-lipoxygenase and interacts with the enzyme product 13-HPODE.

Defining the arachidonic acid binding site of human 15-lipoxygenase. Molecular modeling and mutagenesis.

Mammalian lipoxygenases have been implicated in the pathogenesis of several inflammatory disorders and are, therefore, important targets for drug discovery. Both plant and mammalian lipoxygenases catalyze the dioxygenation of polyunsaturated fatty acids, which contain one or more 1,4-cis,cis-pentadiene units to yield hydroperoxide products. At the time this study was initiated, soybean lipoxygenase-1 was the only lipoxygenase for which an atomic resolution structure had been determined. No structure of lipoxygenase with substrate or inhibitor bound is currently available. A model of arachidonic acid docked into the proposed substrate binding site in the soybean structure is presented here. Analysis of this model suggested two residues, an aromatic residue and a positively charged residue, could be critical for substrate binding. Validation of this model is provided by site-directed mutagenesis of human 15-lipoxygenase, despite the low amino acid sequence identity between the soybean and mammalian enzymes. Both a positively charged amino acid residue (Arg402) and an aromatic amino acid residue (Phe414) are identified as critical for the binding of fatty acid substrates in human 15-lipoxygenase. Thus, binding determinants shown to be characteristic of non-enzymatic fatty acid-binding proteins are now implicated in the substrate binding pocket of lipoxygenases.