Transcription factors and miRNA act as contrary regulators of gene expression in the testis and epididymis of Bos indicus animals.

Spermatogenesis is highly conserved among mammalians, but its gene expression and regulatory profile are not entirely known. As transcription factors (TFs) and miRNAs are crucial for gene expression regulation, identifying genes negatively regulated by miRNAs and positively regulated by TFs in the testis and epididymis can provide a deeper understanding of gene expression and regulatory patterns. To do this, we used expression data coming from RNA-Seq and miRNA-Seq experiments made with biopsies from testicular parenchyma, head of the epididymis, and tail of the epididymis of four Brahman bulls. We identified miRNA differentially expressed (DE) by comparing the three distinct tissues. A co-expression analysis combined with a regulatory impact factor approach identified miRNAs and TFs with regulatory impact over gene expression regulation in the Bos indicus tissues studied. We identified 116 DE miRNAs, 206 miRNAs and 237 TFs with a significant regulatory impact on mRNA patterns in the tissues' comparisons. bta-miR-196b was the only DE miRNA for all tissue comparisons and it may be a regulator of spermatogenesis through its links with adipogenesis and insulin biosynthesis. DE genes and TFs involved in contrary regulations between the epididymis head and testis parenchyma were associated with spermatogenesis, sexual reproduction, and sperm motility. Our results provide possible mechanisms, governed by the contrary effect of miRNA and TF, leading to the differential expression between the studied tissues. We have demonstrated that our predictions of miRNAs and TFs co-regulations over target DE genes can retrieve known regulatory mechanisms and predict new ones that merit further validation.

Repressive epigenetic mechanisms, such as the H3K27me3 histone modification, were predicted to affect muscle gene expression and its mineral content in Nelore cattle.

Epigenetic repression has been linked to the regulation of different cell states. In this study, we focus on the influence of this repression, mainly by H3K27me3, over gene expression in muscle cells, which may affect mineral content, a phenotype that is relevant to muscle function and beef quality. Based on the inverse relationship between H3K27me3 and gene expression (i.e., epigenetic repression) and on contrasting sample groups, we computationally predicted regulatory genes that affect muscle mineral content. To this end, we applied the TRIAGE predictive method followed by a rank product analysis. This methodology can predict regulatory genes that might be affected by repressive epigenetic regulation related to mineral concentration. Annotation of orthologous genes, between human and bovine, enabled our investigation of gene expression in the Longissimus thoracis muscle of Bos indicus cattle. The animals under study had a contrasting mineral content in their muscle cells. We identified candidate regulatory genes influenced by repressive epigenetic mechanisms, linking histone modification to mineral content in beef samples. The discovered candidate genes take part in multiple biological pathways, i.e., impulse transmission, cell signalling, immunological, and developmental pathways. Some of these genes were previously associated with mineral content or regulatory mechanisms. Our findings indicate that epigenetic repression can partially explain the gene expression profiles observed in muscle samples with contrasting mineral content through the candidate regulators here identified.

Bivariate GWAS reveals pleiotropic regions among feed efficiency and beef quality-related traits in Nelore cattle.

Feed-efficient cattle selection is among the most leading solutions to reduce cost for beef cattle production. However, technical difficulties in measuring feed efficiency traits had limited the application in livestock. Here, we performed a Bivariate Genome-Wide Association Study (Bi-GWAS) and presented candidate biological mechanisms underlying the association between feed efficiency and meat quality traits in a half-sibling design with 353 Nelore steers derived from 34 unrelated sires. A total of 13 Quantitative Trait Loci (QTL) were found explaining part of the phenotypic variations. An important transcription factor of adipogenesis in cattle, the TAL1 (rs133408775) gene located on BTA3 was associated with intramuscular fat and average daily gain (IMF-ADG), and a region located on BTA20, close to CD180 and MAST4 genes, both related to fat accumulation. We observed a low positive genetic correlation between IMF-ADG (r = 0.30 ± 0.0686), indicating that it may respond to selection in the same direction. Our findings contributed to clarifying the pleiotropic modulation of the complex traits, indicating new QTLs for bovine genetic improvement.

Prune homolog 2 with BCH domain (PRUNE2) gene expression is associated with feed efficiency-related traits in Nelore steers.

Animal feeding is a critical factor in increasing producer profitability. Improving feed efficiency can help reduce feeding costs and reduce the environmental impact of beef production. Candidate genes previously identified for this trait in differential gene expression studies (e.g., case-control studies) have not examined continuous gene-phenotype variation, which is a limitation. The aim of this study was to investigate the association between the expression of five candidate genes in the liver, measured by quantitative real-time PCR and feed-related traits. We adopted a linear mixed model to associate liver gene expression from 52 Nelore steers with the following production traits: average daily gain (ADG), body weight (BW), dry matter intake (DMI), feed conversion ratio (FCR), feed efficiency (FE), Kleiber index (KI), metabolic body weight (MBW), residual feed intake (RFI), and relative growth ratio (RGR). The total expression of the prune homolog 2 (PRUNE2) gene was significantly associated with DMI, FCR, FE, and RFI (P < 0.05). Furthermore, we have identified a new transcript of PRUNE2 (TCONS_00027692, GenBank MZ041267) that was inversely correlated with FCR and FE (P < 0.05), in contrast to the originally identified PRUNE2 transcript. The cytochrome P450 subfamily 2B (CYP2B6), early growth response protein 1 (EGR1), collagen type I alpha 1 chain (COL1A1), and connective tissue growth factor (CTGF) genes were not associated with any feed efficiency-related traits (P > 0.05). The findings reported herein suggest that PRUNE2 expression levels affects feed efficiency-related traits variation in Nelore steers.

Single nucleotide polymorphisms affect miRNA target prediction in bovine.

Single nucleotide polymorphisms (SNPs) can have significant effects on phenotypic characteristics in cattle. MicroRNAs (miRNAs) are small, non-coding RNAs that act as post-transcriptional regulators by binding them to target mRNAs. In the present study, we scanned ~56 million SNPs against 1,064 bovine miRNA sequences and analyzed, in silico, their possible effects on target binding prediction, primary miRNA formation, association with QTL regions and the evolutionary conservation for each SNP locus. Following target prediction, we show that 71.6% of miRNA predicted targets were altered as a consequence of SNPs located within the seed region of the mature miRNAs. Next, we identified variations in the Minimum Free Energy (MFE), which represents the capacity to alter molecule stability and, consequently, miRNA maturation. A total of 48.6% of the sequences analyzed showed values within those previously reported as sufficient to alter miRNA maturation. We have also found 131 SNPs in 46 miRNAs, with altered target prediction, occurring in QTL regions. Lastly, analysis of evolutionary conservation scores for each SNP locus suggested that they have a conserved biological function through the evolutionary process. Our results suggest that SNPs in microRNAs have the potential to affect bovine phenotypes and could be of great value for genetic improvement studies, as well as production.

Genetic regulators of mineral amount in Nelore cattle muscle predicted by a new co-expression and regulatory impact factor approach.

Mineral contents in bovine muscle can affect meat quality, growth, health, and reproductive traits. To better understand the genetic basis of this phenotype in Nelore (Bos indicus) cattle, we analysed genome-wide mRNA and miRNA expression data from 114 muscle samples. The analysis implemented a new application for two complementary algorithms: the partial correlation and information theory (PCIT) and the regulatory impact factor (RIF), in which we included the estimated genomic breeding values (GEBVs) for the phenotypes additionally to the expression levels, originally proposed for these methods. We used PCIT to determine putative regulatory relationships based on significant associations between gene expression and GEBVs for each mineral amount. Then, RIF was adopted to determine the regulatory impact of genes and miRNAs expression over the GEBVs for the mineral amounts. We also investigated over-represented pathways, as well as pieces of evidences from previous studies carried in the same population and in the literature, to determine regulatory genes for the mineral amounts. For example, NOX1 expression level was positively correlated to Zinc and has been described as Zinc-regulated in humans. Based on our approach, we were able to identify genes, miRNAs and pathways not yet described as underlying mineral amount. The results support the hypothesis that extracellular matrix interactions are the core regulator of mineral amount in muscle cells. Putative regulators described here add information to this hypothesis, expanding the knowledge on molecular relationships between gene expression and minerals.

Muscle transcriptome analysis reveals genes and metabolic pathways related to mineral concentration in Bos indicus.

Mineral content affects the biological processes underlying beef quality. Muscle mineral concentration depends not only on intake-outtake balance and muscle type, but also on age, environment, breed, and genetic factors. To unveil the genetic factors involved in muscle mineral concentration, we applied a pairwise differential gene expression analysis in groups of Nelore steers genetically divergent for nine different mineral concentrations. Here, based on significant expression differences between contrasting groups, we presented candidate genes for the genetic regulation of mineral concentration in muscle. Functional enrichment and protein-protein interaction network analyses were carried out to search for gene regulatory processes concerning each mineral. The core genetic regulation for all minerals studied, except Zn, seems to rest on interactions between components of the extracellular matrix. Regulation of adipogenesis-related pathways was also significant in our results. Antagonistic patterns of gene expression for fatty acid metabolism-related genes may explain the Cu and Zn antagonistic effect on fatty acid accumulation. Our results shed light on the role of these minerals on cell function.

Linkage disequilibrium and haplotype block structure in a composite beef cattle breed.

BACKGROUND: The development of linkage disequilibrium (LD) maps and the characterization of haplotype block structure at the population level are useful parameters for guiding genome wide association (GWA) studies, and for understanding the nature of non-linear association between phenotypes and genes. The elucidation of haplotype block structure can reduce the information of several single nucleotide polymorphisms (SNP) into the information of a haplotype block, reducing the number of SNPs in a coherent way for consideration in GWA and genomic selection studies. RESULTS: The maximum average LD, measured by r2 varied between 0.33 to 0.40 at a distance of < 2.5 kb, and the minimum average values of r2 varied between 0.05 to 0.07 at distances ranging from 400 to 500 kb, clearly showing that the average r2 reduced with the increase in SNP pair distances. The persistence of LD phase showed higher values at shorter genomic distances, decreasing with the increase in physical distance, varying from 0.96 at a distance of < 2.5 kb to 0.66 at a distance from 400 to 500 kb. A total of 78% of all SNPs were clustered into haplotype blocks, covering 1,57 Mb of the total autosomal genome size. CONCLUSIONS: This study presented the first high density linkage disequilibrium map and haplotype block structure for a composite beef cattle population, and indicates that the high density SNP panel over 700 k can be used for genomic selection implementation and GWA studies for Canchim beef cattle.