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Background: To identify differentially expressed genes (DEGs) between muscle and adipose in cattle, we analyzed the data from the RNA sequencing of three Angus × Qinchuan crossbred cattle. Results: Searched the Gene Expression Omnibus (GEO) for a microarray dataset of Yan yellow cattle, GSE49992. After the DEGs were identified, we used STRING and Cytoscape to construct a protein–protein interaction (PPI) network, subsequently analyzing the major modules of key genes. In total, 340 DEGs were discovered, including 21 hub genes, which were mainly enriched in muscle contraction, skeletal muscle contraction, troponin complex, lipid particle, Z disc, tropomyosin binding, and actin filament binding. Conclusions: In summary, these genes can be regarded as candidate biomarkers for the regulation of muscle and adipose development. Normal 0 false false false EN-US X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Tabla normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin-top:0cm; mso-para-margin-right:0cm; mso-para-margin-bottom:8.0pt; mso-para-margin-left:0cm; line-height:107%; mso-pagination:widow-orphan; font-size:11.0pt; font-family:"Calibri",sans-serif; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin; mso-bidi-font-family:"Times New Roman"; mso-bidi-theme-font:minor-bidi;}

This study investigated the effect of ELOVL6 (elongation of very long chain fatty acids protein 6) and its underlying mechanism on lipid metabolism in bovine adipocytes. The ELOVL6 gene was overexpressed in bovine adipocytes by adenoviruses, and RNA sequencing was performed. Overexpression of ELOVL6 showed reduced proportions of C14:0 (Myristic) and C16:0 (palmitate) fatty acids and increased proportions of C18.0 (stearate) and C20:4n6 (arachidonic) fatty acids in adipocytes. In addition, a total of 2170 differentially expressed genes (DEGs) were found, containing 1802 up-regulated and 368 down-regulated genes. KEGG pathway analysis revealed that the down-regulated genes were linked with the regulation of lipolysis and the Wnt signaling pathway. The up-regulated genes were mainly involved in the FoxO signaling pathway; the PI3K-Akt signaling pathway; and the cAMP signaling pathway. In conclusion, our results suggest that ELOVL6 could affect the fatty acid composition in bovine adipocytes. We identified numerous related DEGs and pathways, which may provide a basis for studying the function and molecular mechanism of the ELOVL6 gene in regulating lipid metabolism.

SIRT5 (sirtuin 5) is located in the mitochondria and plays an important role in biological processes such as maintaining the balance of lipid metabolism and promoting fatty acid oxidation mobilization. In this study, the bovine preadipocyte differentiation and obese mouse models were constructed; combined with transcriptome sequencing (RNA-seq) and morphological identification, the regulatory and molecular mechanisms underlying the effects of SIRT5 on bovine preadipocyte differentiation and lipid metabolism were studied. The results reveal that during the differentiation of preadipocytes, SIRT5 inhibited the expression of key genes that promote lipid formation and differentiation in fatty acid biosynthesis and PPAR pathways. SIRT5 significantly activated the adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK) pathway and repressed the mitogen-activated protein kinase (MAPK) pathway. Eventually, SIRT5 significantly inhibited the differentiation of bovine preadipocytes and simultaneously inhibited lipid synthesis and lipid deposition in adipocytes. The verification experiments performed using obese mice also yielded consistent results in vivo.

Copy number variation (CNV) refers to a kind of structural variation, having functional and evolutionary effects on phenotypes. Thus far, further elucidation of the CNVs in different Chinese indigenous cattle breeds by whole genome sequencing have yet not been done. In this study, a comprehensive genomic analysis was performed on 75 cattle individuals including six Chinese indigenous cattle breeds and two non-native specialized beef cattle breeds. Based on the 11,486 CNVRs discovered, population analysis was performed, showed that all the cattle breeds clustered in to three clades, consistent with their lineages Bos taurus, Bos taurus × Bos indicus and Bos indicus. Importantly, a set of CNVRs related genes were found to be associated with the traits of interest, which include meat production or quality (CAST, ACTC1, etc.), adaption (BLA-DQB, EGLN2, etc.) and coat color (KIT, MITF, etc.). These results provide valuable full genome variation resources for Chinese bovine genome research and would be helpful for cattle breeding and selection programs in the future.

The BBS2 gene plays a vital role in human obesity and fat deposition. However, little is known about it in beef cattle. Therefore, this study investigates the function of BBS2 in the fat deposition of beef cattle and screens the effective SNPs marker for meat quality traits in cattle breeding. The expression of BBS2 is negatively correlated with marbling ratios of beef cattle. Moreover, the knockdown of BBS2 promoted adipogenesis and lipid accumulation of bovine preadipocytes by stimulating PPARγ, FABP4, and FASN expression (P 0.01). Four novel SNPs in the exons of BBS2 in Chinese Qinchuan cattle were identified and of which the g.24226239C T (Q527), g.24223562G A (V441I), and g.24227851A G (Q627R) were significantly associated with the meat quality of Qinchuan cattle (P 0.01, P 0.05). The findings suggested that BBS2 could be used as a candidate gene for meat quality improvement in Qinchuan cattle. Furthermore, these genotypes can be exploited as molecular markers in future beef breeding projects.

Mongolian cattle (MG, Bos taurus) and Minnan cattle (MN, Bos indicus) are two different breeds of Chinese indigenous cattle, representing North type and South type, respectively. However, their value and potential have not yet been discovered at the genomic level. In this study, 26 individuals of MN and MG were sequenced for the first time at an average of 13.9- and 12.8-fold, respectively. Large numbers of different variations were identified. In addition, the analyses of phylogenetic and population structure showed that these two cattle breeds are distinct from each other, and results of linkage disequilibrium analysis revealed that these two cattle breeds have undergone various degrees of intense natural or artificial selection. Subsequently, 496 and 306 potential selected genes (PSRs) were obtained in MN and MG, containing 1,096 and 529 potential selected genes (PSGs), respectively. These PSGs, together with the analyzed copy number variation (CNV)-related genes, showed potential relations with their phenotypic characteristics, including environmental adaptability (e.g., DVL2, HSPA4, CDHR4), feed efficiency (e.g., R3HDM1, PLAG1, XKR4), and meat/milk production (e.g., PDHB, LEMD3, APOF). The results of this study help to gain new insights into the genetic characteristics of two distinct cattle breeds and will contribute to future cattle breeding., Graphical Abstract, Mongolian cattle (Bos taurus) and Minnan cattle (Bos indicus) are two different breeds of Chinese indigenous cattle, representing North type and South type, respectively. Authors carried out comprehensive genome-wide analysis of them, and results revealed that these two cattle breeds have undergone various degrees of intense natural or artificial selection.

E. coli is the main causative agent of mastitis in dairy cows, but the mechanism of molecular regulation underlying the occurrence and development of mastitis has not yet been fully elucidated. In this study, an E. coli-induced mastitis model was created and RNASeq technology was used to measure the miRNA expression profiles at different times post-infection (0, 1, 3, 5, 7 dpi), as well as to screen for differentially expressed miRNA. The results show detection of 2416 miRNAs, including 628 known miRNAs and 1788 newly discovered miRNAs. A total of 200 differentially expressed miRNAs were found at different time points. Bioinformatics analysis showed that these differentially expressed miRNAs may regulate the occurrence and development of mastitis in dairy cows through seven signal transduction pathways, namely cytokine-cytokine receptor interaction, MAPK signaling pathway, chemokine signaling pathway, leukocyte transendothelial migration, T cell receptor signaling pathway, Toll-like receptor signaling pathway, and cell adhesion molecules. In addition, bta-miR-200a, bta-miR-205, bta-miR-122, bta-miR-182 and the newly discovered conservative_15_7229 might be involved in immune process in late stage of E. coli-induced mastitis. The results of this study lay the foundation for molecular network analysis of mastitis and molecular breeding of dairy cows.

Fatty acid synthase (FASN) is an enzyme involved with fat deposition and fatty acid composition in cattle. This study was conducted to detect single nucleotide polymorphisms (SNPs) of the FASN gene and explore their relationships with ultrasound carcass traits in order to assess the potential use of the FASN gene for the breeding selection of Qinchuan cattle for desirable carcass traits. The frequencies of SNP g.12740C>T, g.13192T>C and g.13232C>T were identified in 525 individual Qinchuan cattle which were also assessed for backfat depth, eye muscle area and intramuscular fat by ultrasound. According to the PIC values, g.13192T>C possessed an intermediate polymorphism (0.25 T, g.12740C>T possessed low polymorphism (PIC C were in Hardy-Weinberg disequilibrium (c2 C was associated with a greater eye muscle area and the TT genotype at g.13232C>T was associated with greater intramuscular fat. When these genotypes were combined there was no difference in eye muscle area and intramuscular fat between the diplotypes. The H2H2 diplotype was associated with carcass traits that are likely to provide economic advantage in Qinchuan cattle. Variations in the FASN genes and their corresponding genotypes may be considered as molecular markers for economic traits in cattle breeding.

MicroRNAs (miRNAs) play crucial roles in regulating innate and adaptive immunity in humans and animals. Infection with E. coli or S. aureus can cause inflammation of the mammary glands, which results in significant economic losses in dairy cattle. However, the regulatory mechanisms of miRNAs in response to E. coli or S. aureus infection in bovine mammary glands have not been thoroughly explored. To discover the differential expression of miRNA in bovine mammary gland challenged with E. coli or S. aureus, we performed miRNA sequencing on tissue samples. A total of 1838 miRNAs were identified, including 580 known-miRNAs (included in the miRbase database) and 1258 predicted novel miRNAs. The miRNA expression patterns indicated that, compared with control samples, 279 miRNAs and 305 miRNAs were differentially expressed miRNAs (DIE-miRNA) in S. aureus and E. coli infected tissues, respectively. Moreover, the results of comparison the DIE-miRNAs between the E. coli and S. aureus infected groups showed that 197 DIE-miRNAs are identical, 108 DIE-miRNAs are specific to the E. coli group, and 82 DIE-miRNAs are specific to the S. aureus group. Many DIE-miRNAs, such as bta-miR-144, bta-miR-451 and bta-miR-7863, might be the useful biomarkers of mastitis caused by E. coli and S. aureus. In addition, target genes of the DIE-miRNAs were predicted. The Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis indicated that these DIE-miRNAs are likely involved in many immune signaling pathways, including the Toll-like receptor signaling pathways, MAPK signaling pathway, cell adhesion molecules, TGF-β signaling pathway, leukocyte trans endothelial migration, cytokine-cytokine receptor interaction, and chemokine signaling pathways. This study has provided supportive evidence that miRNAs may serve as diagnostic biomarkers of mastitis in dairy cows, and suggests potentially of effective strategies to combat mastitis.

The bovine genetic resources in China are diverse, but their value and potential are yet to be discovered. To determine the genetic diversity and population structure of Chinese cattle, we analyzed the whole genomes of 46 cattle from six phenotypically and geographically representative Chinese cattle breeds, together with 18 Red Angus cattle genomes, 11 Japanese black cattle genomes and taurine and indicine genomes available from previous studies. Our results showed that Chinese cattle originated from hybridization between Bos taurus and Bos indicus. Moreover, we found that the level of genetic variation in Chinese cattle depends upon the degree of indicine content. We also discovered many potential selective sweep regions associated with domestication related to breed-specific characteristics, with selective sweep regions including genes associated with coat color (ERCC2, MC1R, ZBTB17, and MAP2K1), dairy traits (NCAPG, MAPK7, FST, ITFG1, SETMAR, PAG1, CSN3, and RPL37A), and meat production/quality traits (such as BBS2, R3HDM1, IGFBP2, IGFBP5, MYH9, MYH4, and MC5R). These findings substantially expand the catalogue of genetic variants in cattle and reveal new insights into the evolutionary history and domestication traits of Chinese cattle.

Adipose tissue is the largest metabolic organ because of adipogenesis controlled by numerous miRNAs. MiR-145 is classified into the same cluster with famous miR-143. However, few studies have investigated the role of miR-145 in adipogenesis. In the current study, we observed that the expression of miR-145 was downregulated during bovine adipogenesis in vivo and in vitro. The results of RNA-Seq analysis showed that miR-145 mainly disturb the PI3K/Akt and MAPK signaling pathways in bovine preadipocytes. MiR-145 inhibited bovine preadipocyte differentiation and downregulated phosphorylation level of Akt and ERK1/2 proteins. Furthermore, insulin, as a powerful inducer initiating adipogenesis and an activator of the PI3K/Akt and MAPK signaling pathways, was able to rescue the downregulation of Akt and ERK1/2 phosphorylation levels caused by miR-145. Taken together, our findings suggest that miR-145 is a potent inhibitor of adipogenesis that may function by reducing the activity of PI3K/Akt and MAPK signaling pathways.

Chinese indigenous pigs exhibit considerable phenotypic diversity, but their population structure and the genetic basis of agriculturally important traits need further exploration. Here, we sequenced the whole genomes of 24 individual pigs representing 22 breeds distributed throughout China. For comparison with European and commercial breeds (one pig per breed), we included seven published pig genomes with our new genomes for analyses. Our results showed that breeds grouped together based on morphological classifications are not necessarily more genetically similar to each other than to breeds from other groups. We found that genetic material from European pigs likely introgressed into five Chinese breeds. We have identified two new subpopulations of domestic pigs that encompass morphology-based criteria in China. The Southern Chinese subpopulation comprises the classical South Chinese Type and part of the Central China Type. In contrast, the Northern Chinese subpopulation comprises the North China Type, the Lower Yangtze River Basin Type, the Southwest Type, the Plateau Type, and the remainder of the Central China Type. Eight haplotypes and two recombination sites were identified within a conserved 40.09 Mb linkage-disequilibrium (LD) block on the X chromosome. Potential candidate genes (LEPR, FANCC, COL1A1, and PCCA) influencing body size were identified. Our findings provide insights into the phylogeny of Chinese indigenous pig breeds and benefit gene mining efforts to improve major economic traits.

This review considers the unique characteristics of Chinese cattle and intramuscular fat content (IMF) as factors influencing meat quality, including tenderness, flavor, and juiciness of meat. Due to its nutritional qualities, meat contributes to a healthy and balanced diet. The intramuscular fat content and eating quality of beef are influenced by many factors, which can generally be divided into on-farm and pre-slaughter factors (breed, sex of cattle, age at slaughter, housing system, diet, and pre-slaughter handling) and postmortem factors (post-slaughter processing, chilling temperature, and packaging). Meat quality traits can also be influenced by the individual genetic background of the animal. Worldwide, the function of genes and genetic polymorphisms that have potential effects on fattening of cattle and beef quality have been investigated. The use of DNA markers is recognized as a powerful and efficient approach to achieve genetic gain for desirable phenotypic characteristics, which is helpful for economic growth. The polymorphisms of the SIRT4, SIRT6, SIRT7, CRTC3, ABHD5, KLF6, H-FABP, and ELOVL6 genes for body and growth characteristics of cattle, and also for beef quality, are considered with the aim of highlighting the significance of beef intramuscular fat content, and that growth, body, and meat quality characteristics are polygenically regulated.

The Src homology 2 B 2 (SH2B2) gene regulate energy balance and body weight at least partially by enhancing Janus kinase-2 (JAK2)-mediated cytokine signaling, including leptin and/or GH signaling. Leptin is an adipose hormone that controls body weight. The objective of the present study is to evaluate the association between body measurement traits and SH2B2 gene polymorphisms as responsible mutations. For this purpose, we selected four single-nucleotide polymorphisms (SNPs) in SH2B2 gene, including two in intron 5 (g.20545A>G, and g.20570G>A, one synonymous SNP g.20693T>C, in exon 6 and one in intron 8 (g.24070C>A, and genotyped them in Qinchuan cattle. SNPs in sample populations were in medium polymorphism level (0.250A, g.20693T>C, and g.24070C>A, significantly (P < 0.05) associated with body length (BL) and chest circumference (CC) in Qinchuan cattle. In addition, H4H3 and H5H5 diplotype had highly significantly (P < 0.01) greater body length (BL), rump length (RL), and chest circumference (CC) than H4H2. Our investigation will not only extend the spectrum of genetic variation of bovine SH2B2 gene, but also provide useful information for the marker assisted selection in beef cattle breeding program.

Background Chinese indigenous pigs exhibit considerable phenotypic diversity, but their population structure and the genetic basis of agriculturally important traits have not been explored. Results Here, we sequenced the whole genomes of 24 individual pigs representing 22 breeds distributed throughout China. For comparison with European and commercial breeds (one pig per breed), we integrated seven published pig genomes with our new genomes. Our results showed that pig domestication occurred at three places in Southeastern Asia, namely the Mekong region, the middle to downstream regions of the Yangtze River, and Tibetan highlands. Moreover, we demonstrated that classic morphological characteristics such as coat color are not consistent with genetic data. We found that genetic material from European pigs likely introgressed into five Chinese breeds. Two new subpopulations of domestic pigs have been identified in South and North China that encompass morphology-based criteria. The Southern Chinese subpopulation comprises the classical Southern China Type and part of the Central China Type, whereas the Northern Chinese subpopulation comprises the North China Type, the Lower Yangtze River Basin Type, the Southwest Type, the Plateau Type, and the remainder of the Central China Type. Eight haplotypes and two recombination sites were identified within a conserved 40.09 Mb linkage-disequilibrium block on the X chromosome. Potential selection and domestication signatures were identified, mainly influencing body size, along with adaptations to cold and hot temperature environments. Conclusions Our findings provide insights into the phylogeny of Chinese indigenous pig breeds, and will be of enormous benefit in identifying beneficial genes to develop superior pig breeds.

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.

Kruppel-like factor 6 (KLF6) genes plays a significant role in the regulation of cell differentiation, proliferation and muscle development. The aim of this study is to investigate the genetic variation and the haplotype combination of the KLF6 gene in Qinchuan cattle and verify its contribution to bovine carcass traits and body measurements. The data were analyzed by real-time quantitative PCR (qPCR) to detect the expression profile of the KLF6 gene in the various tissues of Qinchuan cattle. PCR amplicons sequencing explored three novel SNPs at loci 3332C > G; 3413C > T and 3521G > A in the 2nd exon region of the KLF6 gene. The expression of KLF6 in the liver, kidney and lung was greater than that of other tissues. Allelic and genotypic frequencies of these SNPs were found to be in Hardy Weinberg equilibrium (P < 0.05). In SNP1, genotype CC, in SNP2, genotype CT and in SNP3 genotype GG were associated (P < 0.05) with larger body and carcass measurements. Association analysis results indicated that individuals with the Hap1/4 diplotype had a longer body and rump, were taller at the withers, and were wider at the hip than the other combinations. In terms of ultrasound carcass measures, Hap1/4 was associated with a larger muscle area and more intramuscular fat than other combinations. The bioinformatics study of the KLF6 protein showed a high degree of conservation in different mammalian species. The above results suggest that the KLF6 gene can used as potential candidate markers gene for the beef breed improvement through marker assisted selection of Qinchuan cattle.

Sirtuins, NAD+-dependent deacylases and ADP-ribosyltransferases, are critical regulators of metabolism involved in many biological processes, and are involved in mediating adaptive responses to the cellular environment. SIRT4 is a mitochondrial sirtuin and has been shown to play a critical role in maintaining insulin secretion and glucose homeostasis. As a regulator of lipid homeostasis, SIRT4 can repress fatty acid oxidation and promote lipid anabolism in nutrient-replete conditions. Using real-time quantitative PCR (qPCR) to explore the molecular mechanisms of transcriptional regulation of bovine SIRT4 during adipocyte differentiation, we found that bovine SIRT4 is expressed at high levels in bovine subcutaneous adipose tissue. SIRT4 knockdown led to decreased expression of adipogenic differentiation marker genes during adipocyte differentiation. The core promoter of bovine SIRT4 was identified in the −402/−60 bp region of the cloned 2-kb fragment containing the 5′-regulatory region. Binding sites were identified in this region for E2F transcription factor-1 (E2F1), CCAAT/enhancer-binding protein β (CEBPβ), homeobox A5 (HOXA5), interferon regulatory factor 4 (IRF4), paired box 4 (PAX4), and cAMP responsive element-binding protein 1 (CREB1) by using Electrophoretic mobility shift assay (EMSA) and luciferase reporter gene assay. We also found that E2F1, CEBPβ, and HOXA5 transcriptionally activate SIRT4 expression, whereas, IRF4, PAX4, and CREB1 transcriptionally repress SIRT4 expression. We further verified that SIRT4 knockdown could affect the ability of these transcription factors (TFs) to regulate the differentiation of bovine adipocytes. In conclusion, our results shed light on the mechanisms underlying the transcriptional regulation of SIRT4 expression in bovine adipocytes.

A new strain of Qinchuan cattle (QNS) has been obtained after more than forty years of selective breeding, and it shows good performance and production traits. To characterize the genetic changes that have resulted from breeding, we sequenced 10 QNS and 10 of the original breed Qinchuan cattle (QCC) for the first time, with average of 12.5-fold depth. A total of 31,242,284 and 29,612,517 SNPs were identified in the QCC and QNS genomes, 47.81% and 44.36% of which were found to be novel, respectively. Furthermore, population structure analysis revealed the selection that these cattle had experienced. Then, 332 and 571 potential selected genes were obtained, associated with enhanced immunity and acclimatization in QCC (CD5, SMARCA2, CATHL2, etc.) and production or meat quality traits in QNS (PLCD3, MB, PPARGC1A, etc.). These results revealed the efforts of selective breeding for Chinese Qinchuan cattle, and will be helpful for future cattle breeding.

Sirtuin 5 (SIRT5) belongs to the mitochondrial sirtuin family, which constitutes a highly conserved family of nicotinamide adenine dinucleotide NAD+-dependent deacetylases and ADP-ribosyltransferases that play important regulatory roles in stress resistance and metabolic homeostasis. SIRT5 was shown to have deacetylase, desuccinylase, and demalonylase activities. However, the mechanisms regulating SIRT5 transcription remain unclear. To explore the molecular regulation of bovine SIRT5 expression, we obtained a 500-base pair fragment of the 5'-regulatory region of bovine SIRT5 by molecular cloning, which contained a region with 3 CpG islands. Electrophoretic mobility shift assays and luciferase reporter assays revealed the E2F transcription factor 4 (E2F4) and Kruppel-like factor 6 (KLF6) binding sites as transcriptional activators or repressors in the promoter region of SIRT5. We further verified that E2F4 and KLF6 bind to the SIRT5 promoter by chromatin immunoprecipitation assays. Additionally, methylation and luciferase report assays showed that SIRT5 promoter activity was enhanced by demethylation, and transcriptional activation by E2F4 and transcriptional inhibition by KLF6 of SIRT5 expression was strengthened by demethylation during adipocytes differentiation. This study focused on the mechanism underlying the methylation and transcriptional regulation of SIRT5 expression in bovine adipocytes.

Previous studies have shown that the cell death-inducing DFF45-like effector-C (CIDEC) gene is involved in lipid storage and energy metabolism, suggesting that it is a potential candidate gene that affects body measurement traits (BMTs) and meat quality traits (MQTs). The aim of this study was to identify polymorphisms of the bovine CIDEC gene and analyze their possible associations with BMTs and MQTs in 531 randomly selected Qinchuan cattle aged between 18 and 24 months. DNA sequencing and polymerase chain reaction-restriction fragment length polymorphism were employed to detect CIDEC single nucleotide polymorphisms (SNPs). We found five SNPs: two in exon 5 (SNP1, g.9815G>A and SNP2, g.9924C>T) and three in the 3'-untranslated region (SNP3, g.13281C>T; SNP4, g.13297A>G; and SNP5, g.13307G>A). SNP1 was a missense mutation that resulted in an arginine to glutamine amino acid change, and exhibited two genotypes (GG and AG). SNP2 was a synonymous mutation that exhibited three genotypes (CC, CT, and TT). SNP3, 4, and 5 were completely linked, and only exhibited two genotypes (CC-AA-GG and CT-AG-GA). We found significant associations between these polymorphisms and BMTs and MQTs (P < 0.05); GG, CT, and CT-AG-GA appeared to be the most beneficial genotypes. Therefore, CIDEC may affect BMTs and MQTs in Qinchuan cattle, and could be used in marker-assisted selection.

The SIX1 gene belongs to the family of six homeodomain transcription factors (TFs), that regulates the extracellular signal-regulated kinase 1/2 (ERK1/2) pathway and mediate skeletal muscle growth and regeneration. Previous studies have demonstrated that SIX1 is positively correlated with body measurement traits (BMTs). However, the transcriptional regulation of SIX1 remains unclear. In the present study, we determined that bovine SIX1 was highly expressed in the longissimus thoracis. To elucidate the molecular mechanisms involved in bovine SIX1 regulation, 2-kb of the 5′ regulatory region were obtained. Sequence analysis identified neither a consensus TATA box nor a CCAAT box in the 5′ flanking region of bovine SIX1. However, a CpG island was predicted in the region −235 to +658 relative to the transcriptional start site (TSS). An electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) assay in combination with serial deletion constructs of the 5′ flanking region, site-directed mutation and siRNA interference demonstrated that MyoD, PAX7 and CREB binding occur in region −689/−40 and play important roles in bovine SIX1 transcription. In addition, MyoG drives SIX1 transcription indirectly via the MEF3 motif. Taken together these interactions suggest a key functional role for SIX1 in mediating skeletal muscle growth in cattle.

Understanding the molecular mechanisms of skeletal myoblast differentiation is essential for studying muscle developmental biology. In our previous study, we reported that knockdown of myocyte enhancer factor 2A (MEF2A) inhibited myoblast differentiation. Here in this study, we further identified that MEF2A controlled this process through regulating the maternally expressed 3 (MEG3)&mdash, iodothyronine deiodinase 3 (DIO3) miRNA mega cluster and protein phosphatase 2A (PP2A) signaling. MEF2A was sufficient to induce MEG3 expression in bovine skeletal myoblasts. A subset of miRNAs in the MEG3-DIO3 miRNA cluster was predicted to target PP2A subunit genes. Consistent with these observations, MEF2A regulated PP2A signaling through its subunit gene protein phosphatase 2 regulatory subunit B, gamma (PPP2R2C) during bovine myoblast differentiation. MiR-758 and miR-543 in the MEG3-DIO3 miRNA cluster were down-regulated in MEF2A-depleted myocytes. Expression of miR-758 and miR-543 promoted myoblast differentiation and repressed PPP2R2C expression. Luciferase activity assay showed that PPP2R2C was post-transcriptionally targeted by miR-758 and miR-543. Taken together, these results reveal that the MEG3-DIO3 miRNAs function at downstream of MEF2A to modulate PP2A signaling in bovine myoblast differentiation.

Gayal (Bos frontalis) is a semi-wild and endangered bovine species that differs from domestic cattle (Bos taurus and Bos indicus) and its genetic background remains unclear. Here, we performed whole-genome sequencing of one Gayal for the first time, with one Red Angus cattle and one Japanese Black cattle as controls. In total, 97.8 Gb of sequencing reads were generated with an average 11.78-fold depth and >98.44% coverage of the reference sequence (UMD3.1). Numerous different variations were identified, 62.24% of the total single nucleotide polymorphisms (SNPs) detected in Gayal were novel and 16,901 breed-specific nonsynonymous SNPs (BS-nsSNPs) that might be associated with traits of interest in Gayal were further investigated. Moreover, the demographic history of bovine species was first analyzed and two population expansions and two population bottlenecks were identified. The obvious differences among their population sizes supported that Gayal was not B. taurus. The phylogenic analysis suggested that Gayal was a hybrid descendant from crossing of male wild gaur and female domestic cattle. These discoveries will provide valuable genomic information regarding potential genomic markers that could predict traits of interest for breeding programs of these cattle breeds and may assist relevant departments with future conservation and utilization of Gayal.

DNA methylation is a major epigenetic modification of the genome and has an essential role in muscle development. The SIX1 gene is thought to play a principal role in mediating skeletal muscle development. In the present study, we determined that bovine SIX1 expression levels were significantly higher in the fetal bovine group (FB) and in undifferentiated Qinchuan cattle muscle cells (QCMCs) than in the adult bovine group (AB) and in differentiated QCMCs. Moreover, a bisulfite sequencing polymerase chain reaction (BSP) analysis of DNA methylation levels showed that three CpG sites in the core promoter region (−216/−28) of the bovine SIX1 gene exhibited significantly higher DNA methylation levels in the AB and differentiated QCMCs groups. In addition, we found that DNA methylation of SIX1 core promoter in vitro obviously influences the promoter activities. An electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) assay, in combination with site-directed mutation and siRNA interference, demonstrated that histone H4 and E2F2 bind to the −216/−28 region and play important roles in SIX1 methylation regulation during development. The results of this study provide the foundation for a better understanding of the regulation of bovine SIX1 expression via methylation and muscle developmental in beef cattle.

The CDIPT is crucial to the fatty acid metabolic pathway, intracellular signal transduction and energy metabolism in eukaryotic cells. We detected three SNPs at 3'-untranslated regions (UTR), named 3'-UTR_108 A > G, 3'-UTR_448 G > A and 3'-UTR_477 C > G, of the CDIPT gene in 618 Qinchuan cattle using PCR-RFLP and DNA sequencing methods. At each of the three SNPs, we found three genotypes named as follows: AA, AB, BB (3'-UTR_108 A > G), CC, CD, DD (3'-UTR_448 G > A) and EE, EF, FF (3'-UTR_477 C > G.). Based on association analysis of these SNPs with ultrasound measurement traits, individuals of genotype BB had a significantly larger loin muscle area than genotype AA. Individuals of genotype CC had significantly thicker back fat than individuals of genotype DD. Individuals of genotype EE also had significantly thicker back fat than did individuals of genotype FF. We conclude that these SNPs of the CDIPT gene could be used as molecular markers for selecting and breeding beef cattle with superior body traits, depending on breeding goals.