Yaran Zhang

Novel polymorphisms of SIX4 gene and their association with body measurement traits in Qinchuan cattle.

Sine oculis homeobox homolog 4 (SIX4) gene belongs to the sine oculis/SIX gene family, which includes six members in vertebrates. SIX4 gene plays a crucial role in skeletal myogenesis, and its genetic variations or deficiency may cause hypopituitarism, suggesting that SIX4 gene is a potential candidate gene affecting body measurement traits (BMTs) in animals. Herein, the objectives of this study were to identify genetic polymorphisms of bovine SIX4 gene and to analyze potential association between single nucleotide polymorphisms (SNPs) and body measurement traits in Qinchuan cattle. In the present study, we investigated polymorphisms of SIX4 gene in 426 Qinchuan cattle using DNA sequencing and polymerase chain reaction-restriction fragment length polymorphisms. Three novel SNPs were identified within bovine SIX4 gene. Associations between body measurement traits and SIX4 gene polymorphisms were investigated, and significant statistical associations were found between polymorphisms of these three SNPs and body measurement traits (P<0.05). Hence, based on results obtained from this study, we conjectured that SIX4 gene may have potential effects on body measurement traits in Qinchuan cattle population and could be used for marker-assisted selection.

Characterization of the promoter region of the bovine long-chain acyl-CoA synthetase 1 gene: Roles of E2F1, Sp1, KLF15, and E2F4

The nutritional value and eating qualities of beef are enhanced when the unsaturated fatty acid content of fat is increased. Long-chain acyl-CoA synthetase 1 (ACSL1) plays key roles in fatty acid transport and degradation, as well as lipid synthesis. It has been identified as a plausible functional and positional candidate gene for manipulations of fatty acid composition in bovine skeletal muscle. In the present study, we determined that bovine ACSL1was highly expressed in subcutaneous adipose tissue and longissimus thoracis. To elucidate the molecular mechanisms involved in bovine ACSL1 regulation, we cloned and characterized the promoter region of ACSL1. Applying 5′-rapid amplification of cDNA end analysis (RACE), we identified multiple transcriptional start sites (TSSs) in its promoter region. Using a series of 5′ deletion promoter plasmids in luciferase reporter assays, we found that the proximal minimal promoter of ACSL1 was located within the region −325/−141 relative to the TSS and it was also located in the predicted CpG island. Mutational analysis and electrophoretic mobility shift assays demonstrated that E2F1, Sp1, KLF15 and E2F4 binding to the promoter region drives ACSL1 transcription. Together these interactions integrate and frame a key functional role for ACSL1 in mediating the lipid composition of beef.

Molecular Characterization of Bovine SMO Gene and Effects of Its Genetic Variations on Body Size Traits in Qinchuan Cattle (Bos taurus).

Smoothened (Smo)-mediated Hedgehog (Hh) signaling pathway governs the patterning, morphogenesis and growth of many different regions within animal body plans. This study evaluated the effects of genetic variations of the bovine SMO gene on economically important body size traits in Chinese Qinchuan cattle. Altogether, eight single nucleotide polymorphisms (SNPs: 1–8) were identified and genotyped via direct sequencing covering most of the coding region and 3ʹUTR of the bovine SMO gene. Both the p.698Ser.>Ser. synonymous mutation resulted from SNP1 and the p.700Ser.>Pro. non-synonymous mutation caused by SNP2 mapped to the intracellular C-terminal tail of bovine Smo protein; the other six SNPs were non-coding variants located in the 3ʹUTR. The linkage disequilibrium was analyzed, and five haplotypes were discovered in 520 Qinchuan cattle. Association analyses showed that SNP2, SNP3/5, SNP4 and SNP6/7 were significantly associated with some body size traits (p < 0.05) except SNP1/8 (p > 0.05). Meanwhile, cattle with wild-type combined haplotype Hap1/Hap1 had significantly (p < 0.05) greater body length than those with Hap2/Hap2. Our results indicate that variations in the SMO gene could affect body size traits of Qinchuan cattle, and the wild-type haplotype Hap1 together with the wild-type alleles of these detected SNPs in the SMO gene could be used to breed cattle with superior body size traits. Therefore, our results could be helpful for marker-assisted selection in beef cattle breeding programs.

Molecular Characterization and Transcriptional Regulation Analysis of the Bovine PDHB Gene

The pyruvate dehydrogenase beta subunit (PDHB) is a subunit of pyruvate dehydrogenase (E1), which catalyzes pyruvate into acetyl-CoA and provides a linkage between the tricarboxylic acid cycle (TCA) and the glycolysis pathway. Previous studies demonstrated PDHB to be positively related to the intramuscular fat (IMF) content. However, the transcriptional regulation of PDHB remains unclear. In our present study, the cDNA of bovine PDHB was cloned and the genomic structure was analyzed. The phylogenetic tree showed bovine PDHB to be closely related to goat and sheep, and least related to chicken. Spatial expression pattern analysis revealed the products of bovine PDHB to be widely expressed with the highest level in the fat of testis. To understand the transcriptional regulation of bovine PDHB, 1899 base pairs (bp) of the 5’-regulatory region was cloned. Sequence analysis neither found consensus TATA-box nor CCAAT-box in the 5’-flanking region of bovine PDHB. However, a CpG island was predicted from nucleotides -284 to +117. Serial deletion constructs of the 5’-flanking region, evaluated in dual-luciferase reporter assay, revealed the core promoter to be located 490bp upstream from the transcription initiation site (+1). Electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation assay (ChIP) in combination with asite-directed mutation experiment indicated both myogenin (MYOG) and the CCAAT/enhancer-binding protein beta (C/EBPß) to be important transcription factors for bovine PDHB in skeletal muscle cells and adipocytes. Our results provide an important basis for further investigation of the bovine PDHB function and regulation in cattle.

Profile of muscle tissue gene expression specific to water buffalo: Comparison with domestic cattle by genome array

In contrast with the past, the water buffalo is now not only a draft animal, but also an important food source of milk and meat. It is increasingly apparent that the water buffalo have huge potential for meat production, but its breeding needs to be investigated. Regarding the molecular mechanisms involved in the meat quality difference between the buffalo (Bubalus bulabis) and yellow cattle (Bos taurus), 12 chemical-physical characteristics related to the meat quality of longissimus thoracis muscles (LTM) have been compared at the age of 36 months. Intramuscular lipid and b* (yellowness) were greater in cattle than the buffalo, whereas a* (redness) was greater in the buffalo. Gene expression profiles were constructed by bovine genome array. A total of 8884 and 10,960 probes were detected in buffalo and cattle, respectively, with 1580 genes being differentially expressed. Over 400 probes were upregulated and nearly 1200 were downregulated in LTM of the buffalo, most being involved in ribosomal RNA (rRNA) processing, cholesterol homeostasis, regulation of transcription, response to hypoxia, and glycolysis. Quantitative real-time PCR was used to validate the microarray data. Enriched GO analyses of highly expressed genes in LTM showed that protein biosynthesis, striated muscle contraction, iron homeostasis, iron transport, glycolysis and glucose metabolism were similar between the buffalo and cattle. High protein content, low fat content and deep meat color of buffalo LTM may be closely associated with the increased expression of genes involved in cholesterol and iron homeostasis, while also reducing the expression of genes involved in ubiquitin-mediated proteolysis and protein oxidative phosphorylation. These results establish the groundwork for further studies on buffalo meat quality and will be beneficial in improving water buffalo breeding by molecular biotechnology.