An across breed, diet and tissue analysis reveals the transcription factor NR1H3 as a key mediator of residual feed intake in beef cattle.

BACKGROUND: Provision of feed is a major determinant of overall profitability in beef production systems, accounting for up to 75% of the variable costs. Thus, improving cattle feed efficiency, by way of determining the underlying genomic control and subsequently selecting for feed efficient cattle, provides a method through which feed input costs may be reduced. The objective of this study was to undertake gene co-expression network analysis using RNA-Sequence data generated from Longissimus dorsi and liver tissue samples collected from steers of two contrasting breeds (Charolais and Holstein-Friesian) divergent for residual feed intake (RFI), across two consecutive distinct dietary phases (zero-grazed grass and high-concentrate). Categories including differentially expressed genes (DEGs) based on the contrasts of RFI phenotype, breed and dietary source, as well as key transcription factors and proteins secreted in plasma were utilised as nodes of the gene co-expression network. RESULTS: Of the 2,929 DEGs within the network analysis, 1,604 were reported to have statistically significant correlations (≥ 0.80), resulting in a total of 43,876 significant connections between genes. Pathway analysis of clusters of co-expressed genes revealed enrichment of processes related to lipid metabolism (fatty acid biosynthesis, fatty acid ß-oxidation, cholesterol biosynthesis), immune function, (complement cascade, coagulation system, acute phase response signalling), and energy production (oxidative phosphorylation, mitochondrial L-carnitine shuttle pathway) based on genes related to RFI, breed and dietary source contrasts. CONCLUSIONS: Although similar biological processes were evident across the three factors examined, no one gene node was evident across RFI, breed and diet contrasts in both liver and muscle tissues. However within the liver tissue, the IRX4, NR1H3, HOXA13 and ZNF648 gene nodes, which all encode transcription factors displayed significant connections across the RFI, diet and breed comparisons, indicating a role for these transcription factors towards the RFI phenotype irrespective of diet and breed. Moreover, the NR1H3 gene encodes a protein secreted into plasma from the hepatocytes of the liver, highlighting the potential for this gene to be explored as a robust biomarker for the RFI trait in beef cattle.

Effect of short term diet restriction on gene expression in the bovine hypothalamus using next generation RNA sequencing technology.

BACKGROUND: Negative energy balance (NEB) is an imbalance between energy intake and energy requirements for lactation and body maintenance affecting high-yielding dairy cows and is of considerable economic importance due to its negative impact on fertility and health in dairy herds. It is anticipated that the cow hypothalamus experiences extensive biochemical changes during the early post partum period in an effort to re-establish metabolic homeostasis. However, there is variation in the tolerance to NEB between individual cows. In order to understand the genomic regulation of ovulation in hypothalamic tissue during NEB, mRNA transcriptional patterns between tolerant and sensitive animals were examined. A short term dietary restriction heifer model was developed which induced abrupt onset of anoestrus in some animals (Restricted Anovulatory; RA) while others maintained oestrous cyclicity (Restricted Ovulatory; RO). A third control group (C) received a higher level of normal feeding. RESULTS: A total of 15,295 genes were expressed in hypothalamic tissue. Between RA and C groups 137 genes were differentially expressed, whereas between RO and C, 32 genes were differentially expressed. Differentially expressed genes were involved in the immune response and cellular motility in RA and RO groups, respectively, compared to C group. The largest difference between groups was observed in the comparison between RA and RO heifers, with 1094 genes shown to be significantly differentially expressed (SDE). Pathway analysis showed that these SDE genes were associated with 6 canonical pathways (P < 0.01), of which neuroactive ligand-receptor interaction was the most significant. Within the comparisons the main over-represented pathway functions were immune response including neuroprotection (CXCL10, Q1KLR3, IFIH1, IL1 and IL8; RA v C and RA v RO); energy homeostasis (AgRP and NPY; RA v RO); cell motility (CADH1, DSP and TSP4; RO v C) and prevention of GnRH release (NTSR1 IL1α, IL1ß, NPY and PACA; RA v RO). CONCLUSIONS: This information will assist in understanding the genomic factors regulating the influence of diet restriction on fertility and may assist in optimising nutritional and management systems for the improvement in reproductive performance.

Effect of dietary restriction and subsequent re-alimentation on the transcriptional profile of hepatic tissue in cattle.

BACKGROUND: Compensatory growth (CG) is an accelerated growth phenomenon observed in animals upon re-alimentation following a period of dietary restriction. It is typically utilised in livestock systems to reduce feed costs during periods of reduced feed availability. The biochemical mechanisms controlling this phenomenon, however, are yet to be elucidated. This study aimed to uncover the molecular mechanisms regulating the hepatic expression of CG in cattle, utilising RNAseq. RNAseq was performed on hepatic tissue of bulls following 125 days of dietary restriction (RES) and again following 55 days of subsequent re-alimentation during which the animals exhibited significant CG. The data were compared with those of control animals offered the same diet on an ad libitum basis throughout (ADLIB). Elucidation of the molecular control of CG may yield critical information on genes and pathways which could be targeted as putative molecular biomarkers for the selection of animals with improved CG potential. RESULTS: Following a period of differential feeding, body-weight and liver weight were 161 and 4 kg higher, respectively, for ADLIB compared with RES animals. At this time RNAseq analysis of liver tissue revealed 1352 significantly differentially expressed genes (DEG) between the two treatments. DEGs indicated down-regulation of processes including nutrient transport, cell division and proliferation in RES. In addition, protein synthesis genes were up-regulated in RES following a period of restricted feeding. The subsequent 55 days of ad libitum feeding for both groups resulted in the body-weight difference reduced to 84 kg, with no difference in liver weight between treatment groups. At the end of 55 days of unrestricted feeding, 49 genes were differentially expressed between animals undergoing CG and their continuously fed counterparts. In particular, hepatic expression of cell proliferation and growth genes were greater in animals undergoing CG. CONCLUSIONS: Greater expression of cell cycle and cell proliferation genes during CG was associated with a 100 % recovery of liver weight during re-alimentation. Additionally, an apparent up-regulation in capacity for cellular protein synthesis during restricted feeding may contribute to and sustain CG during re-alimentation. DEGs identified are potential candidate genes for the identification of biomarkers for CG, which may be incorporated into future breeding programmes.

Insulin secretion and signaling in response to dietary restriction and subsequent re-alimentation in cattle.

The objectives of this study were to examine systemic insulin response to a glucose tolerance test (GTT) and transcript abundance of genes of the insulin signaling pathway in skeletal muscle, during both dietary restriction and re-alimentation-induced compensatory growth. Holstein Friesian bulls were blocked to one of two groups: 1) restricted feed allowance for 125 days (period 1) (RES, n = 15) followed by ad libitum feeding for 55 days (period 2) or 2) ad libitum access to feed throughout (periods 1 and 2) (ADLIB, n = 15). On days 90 and 36 of periods 1 and 2, respectively, a GTT was performed. M. longissimus dorsi biopsies were harvested from all bulls on days 120 and 15 of periods 1 and 2, respectively, and RNA-Seq analysis was performed. RES displayed a lower growth rate during period 1 (RES: 0.6 kg/day, ADLIB: 1.9 kg/day; P < 0.001), subsequently gaining more during re-alimentation (RES: 2.5 kg/day, ADLIB: 1.4 kg/day; P < 0.001). Systemic insulin response to glucose administration was lower in RES in period 1 (P < 0.001) with no difference observed during period 2. The insulin signaling pathway in M. longissimus dorsi was enriched (P < 0.05) in response to dietary restriction but not during re-alimentation (P > 0.05). Genes differentially expressed in the insulin signaling pathway suggested a greater sensitivity to insulin in skeletal muscle, with pleiotropic effects of insulin signaling interrupted during dietary restriction. Collectively, these results indicate increased sensitivity to glucose clearance and skeletal muscle insulin signaling during dietary restriction; however, no overall role for insulin was apparent in expressing compensatory growth.

Effect of feed restriction and subsequent re-alimentation on hormones and genes of the somatotropic axis in cattle.

The objective of this study was to characterize the effect of feed restriction and compensatory growth during re-alimentation on the functionality of the somatotropic axis. We blocked 60 bulls into one of two groups: 1) restricted feed allowance for 125 days (period 1) (RES, n = 30) followed by ad libitum feeding for 55 days (period 2) or 2) ad libitum access to feed throughout (ADLIB, n = 30). A growth hormone releasing hormone (GHRH) challenge was performed during each period. At the end of each period, 15 animals from each treatment were slaughtered and hepatic tissue collected. Hepatic expression of 13 genes of the somatotropic axis was measured by qRT-PCR. RES displayed a lower growth rate during period 1 (0.6 vs. 1.9 kg/day; P < 0.001), subsequently gaining more than ADLIB animals during period 2 (2.5 vs. 1.4 kg/day; P < 0.001). Growth hormone response to GHRH was not different between treatments at either time-point (P > 0.05); however, resultant plasma IGF-1 was lower in period 1 and greater in period 2 in RES animals (P < 0.05). Expression of IGFBP2 was higher (P < 0.01) and IGF1 (P < 0.001) and GHRIA (P < 0.05) lower in RES compared with ADLIB during period 1, with no difference evident in period 2 (P > 0.05). Collectively, the results of this study are consistent with uncoupling of the somatotropic axis following feed restriction. However, there is no evidence from this study that the somatotropic axis per se is a significant contributor to compensatory growth.

Transcriptional response to an alternative diet on liver, muscle, and rumen of beef cattle.

Feed cost represents a major economic determinant within cattle production, amounting to an estimated 75% of the total variable costs. Consequently, comprehensive approaches such as optimizing feed utilization through alternative feed sources, alongside the selection of feed-efficient animals, are of great significance. Here, we investigate the effect of two diets, traditional corn-grain fed and alternative by-product based, on 14 phenotypes related to feed, methane emission and production efficiency and on multi-tissue transcriptomics data from liver, muscle, and rumen wall, derived from 52 Nellore bulls, 26 on each diet. To this end, diets were contrasted at the level of phenotype, gene expression, and gene-phenotype network connectivity. As regards the phenotypic level, at a P value < 0.05, significant differences were found in favour of the alternative diet for average daily weight gain at finishing, dry matter intake at finishing, methane emission, carcass yield and subcutaneous fat thickness at the rib-eye muscle area. In terms of the transcriptional level of the 14,776 genes expressed across the examined tissues, we found 487, 484, and 499 genes differentially expressed due to diet in liver, muscle, and rumen, respectively (P value < 0.01). To explore differentially connected phenotypes across both diet-based networks, we focused on the phenotypes with the largest change in average number of connections within diets and tissues, namely methane emission and carcass yield, highlighting, in particular, gene expression changes involving SREBF2, and revealing the largest differential connectivity in rumen and muscle, respectively. Similarly, from examination of differentially connected genes across diets, the top-ranked most differentially connected regulators within each tissue were MEOX1, PTTG1, and BASP1 in liver, muscle, and rumen, respectively. Changes in gene co-expression patterns suggest activation or suppression of specific biological processes and pathways in response to dietary interventions, consequently impacting the phenotype. The identification of genes that respond differently to diets and their associated phenotypic effects serves as a crucial stepping stone for further investigations, aiming to build upon our discoveries. Ultimately, such advancements hold the promise of improving animal welfare, productivity, and sustainability in livestock farming.

Early life nutrition affects the molecular ontogeny of testicular development in the young bull calf.

Enhanced early life nutrition accelerates sexual development in the bull calf through neuroendocrine-signalling mediated via the hypothalamic-pituitary-testicular axis. Our aim was to assess the impact of contrasting feeding regimes in bull calves during the first 12 weeks of life on the testes transcriptome and proteome. Holstein-Friesian bull calves were offered either a high (HI) or moderate (MOD) plane of nutrition, designed to support target growth rates of 1.0 and 0.5 kg/day, respectively. At 12 weeks of age all calves were euthanized, testicular parenchyma sampled, and global transcriptome (miRNAseq and mRNAseq) and proteome analyses undertaken. Bioinformatic analyses revealed 7 differentially expressed (DE) miRNA and 20 DE mRNA. There were no differentially abundant proteins between the two dietary groups. Integration of omics results highlighted a potential role for the cadherin gene, CDH13, in earlier reproductive development. Furthermore, co-regulatory network analysis of the proteomic data revealed CDH13 as a hub protein within a network enriched for processes related to insulin, IGF-1, androgen and Sertoli cell junction signalling pathways as well as cholesterol biosynthesis. Overall, results highlight a potential role for CDH13 in mediating earlier reproductive development as a consequence of enhanced early life nutrition in the bull calf.

Genome-wide association study reveals candidate markers related to field fertility and semen quality traits in Holstein-Friesian bulls.

In vitro assessment of bull semen quality is routinely used in bull semen processing centres in order to ensure that semen destined to be used in the field has passed minimum standards. Despite these stringent quality control checks, individual bulls that pass the quality control checks can still vary in field fertility by up to 25%. A genome-wide association study was undertaken to determine genetic markers associated with prefreeze and post-thaw bull sperm quality traits as well as field fertility. Genome-wide association analysis was performed using a single nucleotide polymorphism (SNP) regression mixed linear model in WOMBAT. Genes within a 250 Kb span of a suggestive (P ≤ 1 × 10-5) SNP were considered as candidate genes. One SNP was associated with adjusted pregnancy rate, and 21 SNPs were associated across the seven semen quality traits (P ≤ 1 × 10-5). Functional candidate genes include SIPA1L2 which was associated with adjusted pregnancy rate. This encodes a Rap GTPase-activating protein involved in Rap1 signalling pathway and was previously found to play a role in the process of sperm differentiation. Gene ontology (GO) analysis also identified significantly enriched biological processes involved protein tyrosine kinase activity including genes such as DYRK1A, TEC and TXK that were associated with sperm motility prior to freezing. Another candidate gene associated with post-thaw sperm motility was FHDC1 which coordinates actin filament and microtubule dynamics. The induced 11 GO terms in the ejaculates rejected after freezing trait were related to ATPase, phosphatase and hydrolase activity. These results reveal novel specific genomic regions and candidate genes associated with economically important phenotypes such as field fertility and semen quality traits.

Effect of breed and diet on the M. longissimus thoracis et lumborum transcriptome of steers divergent for residual feed intake.

Improving cattle feed efficiency through selection of residual feed intake (RFI) is a widely accepted approach to sustainable beef production. A greater understanding of the molecular control of RFI in various breeds offered contrasting diets is necessary for the accurate identification of feed efficient animals and will underpin accelerated genetic improvement of the trait. The aim of this study was to determine genes and biological processes contributing to RFI across varying breed type and dietary sources in skeletal muscle tissue. Residual feed intake was calculated in Charolais and Holstein-Friesian steers across multiple dietary phases (phase-1: high concentrate (growing-phase); phase-2: zero-grazed grass (growing-phase); phase-3: high concentrate (finishing-phase). Steers divergent for RFI within each breed and dietary phase were selected for muscle biopsy collection, and muscle samples subsequently subjected to RNAseq analysis. No gene was consistently differentially expressed across the breed and diet types examined. However, pathway analysis revealed commonality across breeds and diets for biological processes including fatty acid metabolism, immune function, energy production and muscle growth. Overall, the lack of commonality of individual genes towards variation in RFI both within the current study and compared to the published literature, suggests other genomic features warrant further evaluation in relation to RFI.

Metabolic influence of core ciliates within the rumen microbiome.

Protozoa comprise a major fraction of the microbial biomass in the rumen microbiome, of which the entodiniomorphs (order: Entodiniomorphida) and holotrichs (order: Vestibuliferida) are consistently observed to be dominant across a diverse genetic and geographical range of ruminant hosts. Despite the apparent core role that protozoal species exert, their major biological and metabolic contributions to rumen function remain largely undescribed in vivo. Here, we have leveraged (meta)genome-centric metaproteomes from rumen fluid samples originating from both cattle and goats fed diets with varying inclusion levels of lipids and starch, to detail the specific metabolic niches that protozoa occupy in the context of their microbial co-habitants. Initial proteome estimations via total protein counts and label-free quantification highlight that entodiniomorph species Entodinium and Epidinium as well as the holotrichs Dasytricha and Isotricha comprise an extensive fraction of the total rumen metaproteome. Proteomic detection of protozoal metabolism such as hydrogenases (Dasytricha, Isotricha, Epidinium, Enoploplastron), carbohydrate-active enzymes (Epidinium, Diplodinium, Enoploplastron, Polyplastron), microbial predation (Entodinium) and volatile fatty acid production (Entodinium and Epidinium) was observed at increased levels in high methane-emitting animals. Despite certain protozoal species having well-established reputations for digesting starch, they were unexpectedly less detectable in low methane emitting-animals fed high starch diets, which were instead dominated by propionate/succinate-producing bacterial populations suspected of being resistant to predation irrespective of host. Finally, we reaffirmed our abovementioned observations in geographically independent datasets, thus illuminating the substantial metabolic influence that under-explored eukaryotic populations have in the rumen, with greater implications for both digestion and methane metabolism.

Identification of differentially expressed mRNAs and miRNAs in spermatozoa of bulls of varying fertility.

Bulls used in artificial insemination, with apparently normal semen quality, can vary significantly in their field fertility. This study aimed to characterize the transcriptome of spermatozoa from high (HF) and low (LF) fertility bulls at the mRNA and miRNA level in order to identify potential novel markers of fertility. Holstein-Friesian bulls were assigned to either the HF or LF group (n = 10 per group) based on an adjusted national fertility index from a minimum of 500 inseminations. Total RNA was extracted from a pool of frozen-thawed spermatozoa from three different ejaculates per bull, following which mRNA-seq and miRNA-seq were performed. Six mRNAs and 13 miRNAs were found differentially expressed (P < 0.05, FC > 1.5) between HF and LF bulls. Of particular interest, the gene pathways targeted by the 13 differentially expressed miRNAs were related to embryonic development and gene expression regulation. Previous studies reported that disruptions to protamine 1 mRNA (PRM1) had deleterious consequences for sperm chromatin structure and fertilizing ability. Notably, PRM1 exhibited a higher expression in spermatozoa from LF than HF bulls. In contrast, Western Blot analysis revealed a decrease in PRM1 protein abundance for spermatozoa from LF bulls; this was not associated with increased protamine deficiency (measured by the degree of chromatin compaction) or DNA fragmentation, as assessed by flow cytometry analyses. However, protamine deficiency was positively and moderately correlated with the percentage of spermatozoa with DNA fragmentation, irrespective of fertility group. This study has identified potential biomarkers that could be used for improving semen quality assessments of bull fertility.

Effect of plane of nutrition in early life on the transcriptome of visceral adipose tissue in Angus heifer calves.

Adipose tissue represents not only an important energy storage tissue but also a major endocrine organ within the body, influencing many biochemical systems including metabolic status, immune function and energy homeostasis. The objective of this study was to evaluate the effect of an enhanced dietary intake during the early calfhood period on the transcriptome of visceral adipose tissue. Artificially reared Angus × Holstein-Friesian heifer calves were offered either a high (HI, n = 15) or moderate (MOD, n = 15) plane of nutrition from 3 to 21 weeks of life. At 21 weeks of age all calves were euthanized, visceral adipose harvested and samples subsequently subjected to mRNA sequencing. Plane of nutrition resulted in the differential expression of 1214 genes within visceral adipose tissue (adj. p < 0.05; fold change > 1.5). Differentially expressed genes were involved in processes related to metabolism and energy production. Biochemical pathways including Sirtuin signalling (adj. p < 0.0001) and the adipogenesis pathways (adj. p = 0.009) were also significantly enriched, indicating greater metabolic processing and adipogenesis in the calves on the high plane of nutrition. Results from this study identify novel genes regulating the molecular response of visceral adipose tissue to an improved plane of nutrition during early calfhood.

An examination of skeletal muscle and hepatic tissue transcriptomes from beef cattle divergent for residual feed intake.

The selection of cattle with enhanced feed efficiency is of importance with regard to reducing feed costs in the beef industry. Global transcriptome profiling was undertaken on liver and skeletal muscle biopsies from Simmental heifers and bulls divergent for residual feed intake (RFI), a widely acknowledged feed efficiency phenotype, in order to identify genes that may be associated with this trait. We identified 5 genes (adj. p < 0.1) to be differentially expressed in skeletal muscle between high and low RFI heifers with all transcripts involved in oxidative phosphorylation and mitochondrial homeostasis. A total of 11 genes (adj. p < 0. 1) were differentially expressed in liver tissue between high and low RFI bulls with differentially expressed genes related to amino and nucleotide metabolism as well as endoplasmic reticulum protein processing. No genes were identified as differentially expressed in either heifer liver or bull muscle analyses. Results from this study show that the molecular control of RFI in young cattle is modified according to gender, which may be attributable to differences in physiological maturity between heifers and bulls of the same age. Despite this we have highlighted a number of genes that may hold potential as molecular biomarkers for RFI cattle.

Transcriptome assisted label free proteomics of hepatic tissue in response to both dietary restriction and compensatory growth in cattle.

Compensatory growth (CG) is a naturally occurring phenomenon where, following a period of under nutrition, an animal exhibits accelerated growth upon re-alimentation. The objective was to identify and quantify hepatic proteins involved in the regulation of CG in cattle. Forty Holstein Friesian bulls were equally assigned to one of four groups. Groups; A1 and A2 had ad libitum access to feed for 125 days, groups R1 and R2 were feed restricted. Following this, R1 and A1 animals were slaughtered. Remaining animals (R2 and A2) were slaughtered following ad libitum feeding for a successive 55 days. At slaughter hepatic tissue samples were collected and label-free quantitative proteomics undertaken with spectra searched against a custom built transcriptome database specific to the animals in this study. 24 differentially abundant proteins were identified during CG (R2 vs. R1) including; PSPH, ASNS and GSTM1, which are involved in nutrient metabolism, immune response and cellular growth. Proteins involved in biochemical pathways related to nutrient metabolism were down-regulated during CG, indicating a possible adaptive response by the liver to a period of fluctuating nutrient availability. The livers ability to regulate its metabolic activity may have profound effects on the efficiency of whole body energy utilization during CG. SIGNIFICANCE: This study is the first to unravel the effect of compensatory growth on the hepatic proteome of cattle using transcriptome-assisted shot gun proteomics. Proteins identified as being affected by dietary restriction and subsequent expression of compensatory growth in this study may, following appropriate validation, contribute to the identification of functional genetic variants. Such information could be harnessed within the context of genomic selection in cattle breeding programs to identify animals with a greater genetic potential to undergo compensatory growth, thus increasing the profitability of the beef sector and accelerating genetic gain.

A high plane of nutrition during early life alters the hypothalamic transcriptome of heifer calves.

The aim was to examine the effect of rapid body weight gain during early calfhood consistent with earlier sexual development on the transcriptional profile of the hypothalamus. Angus X Holstein-Friesian heifer calves (19 ± 5 days of age) were offered a high (HI, n = 14) or moderate (MOD, n = 15) plane of nutrition from 3 to 21 weeks of age to achieve a growth rate of 1.2 kg/d and 0.5 kg/d, respectively. Following euthanasia at 21 weeks, the arcuate nucleus (ARC) region was separated from the remainder of the hypothalamus and both were subjected to RNA-Seq. HI calves exhibited altered expression of 80 and 39 transcripts in the ARC and the remaining hypothalamus, respectively (P < 0.05) including downregulation of AGRP and NPY and upregulation of POMC, previously implicated in precocious sexual development. Stress-signaling pathways were amongst the most highly dysregulated. Organ morphology, reproductive system development and function, and developmental disorder were amongst the networks derived from differentially expressed genes (DEGs) in the ARC. Gene co-expression analysis revealed DEGs within the ARC (POMC, CBLN2, CHGA) and hypothalamus (PENK) as hub genes. In conclusion, enhanced nutrition during early calfhood alters the biochemical regulation of the hypothalamus consistent with advanced sexual development in the prepubertal heifer.

Effect of plane of nutrition during the first 12 weeks of life on growth, metabolic and reproductive hormone concentrations, and testicular relative mRNA abundance in preweaned Holstein Friesian bull calves.

The objective of this study was to examine the effect of nutrition during the first 12 wk of life on aspects of the physiological and transcriptional regulation of testicular and overall sexual development in the bull calf. Holstein Friesian bull calves with a mean (SD) age and bodyweight of 17.5 (2.85) d and 48.8 (5.30) kg, respectively, were assigned to either a high (HI; n = 15) or moderate (MOD; n = 15) plane of nutrition and were individually fed milk replacer and concentrate to achieve overall target growth rates of at least 1.0 and 0.5 kg/d, respectively. Throughout the trial, animal growth performance, feed intake, and systemic concentrations of metabolites, metabolic hormones, and reproductive hormones were assessed. Additionally, pulsatility of reproductive hormones (luteinizing hormone, follicle-stimulating hormone, and testosterone) was recorded at 15-min intervals during a 10-h period at 10 wk of age. At 87 ± 2.14 d of age, all calves were euthanized, testes were weighed, and testicular tissue was harvested. Differential expression of messenger ribonucleic acid (mRNA) candidate genes involved in testicular development was examined using quantitative polymerase chain reaction assays. All data were analyzed using the MIXED procedure in Statistical Analysis Software using terms for treatment as well as time for repeated measures. Blood metabolites and metabolic hormones generally reflected the improved metabolic status of the calves on the HI plane of nutrition though the concentrations of reproductive hormones were not affected by diet. Calves on the HI diet had greater mean (SED) slaughter weight (112.4 vs. 87.70 [2.98] kg; P < 0.0001) and testicular tissue weight (29.2 vs. 20.1 [2.21] g; P = 0.0003) than those on the MOD diet. Relative mRNA abundance data indicated advanced testicular development through upregulation of genes involved in cellular metabolism (SIRT1; P = 0.0282), cholesterol biosynthesis (EBP; P = 0.007), testicular function (INSL3; P = 0.0077), and Sertoli cell development (CLDN11; P = 0.0054) in HI compared with MOD calves. In conclusion, results demonstrate that offering dairy-bred male calves a high plane of nutrition during the first 3 mo of life not only improves growth performance and metabolic status but also advances testicular development consistent with more precocious sexual maturation.

Effects of dietary n-3-PUFA supplementation, post-insemination plane of nutrition and pregnancy status on the endometrial transcriptome of beef heifers.

Supplementation of cattle diets with n-3-polyunsaturated fatty acids (PUFA) can improve reproductive efficiency. Conversely, short-term fluctuations in feed supply can impact pregnancy establishment. The objectives of this study were to examine the effects of (1) dietary supplementation with n-3-PUFA and (2) post-insemination plane of nutrition on the endometrial transcriptome. Beef crossbred heifers were offered concentrate based diets fortified with n-3-PUFA (PUFA; n = 32) or not (CONT; n = 28) for 30 days prior to breeding at a synchronised oestrous. Following artificial insemination, heifers were allocated within treatment to either a high or low plane of nutrition. Heifers were maintained on these diets for 16 days following which endometrial tissue was harvested at slaughter for subsequent RNAseq analysis. The influence of pregnancy status on the endomentrial transcriptome, within each dietary treatment group, was also examined. Post-insemination diet affected (P < 0.05) the endometrial transcriptome. Specifically, within n-3-PUFA-supplemented heifers, genes involved in embryonic development and mTOR signalling pathways, important in pregnancy establishment, were identified as differentially expressed. Results indicate that dietary supplementation of cattle diets with n-3-PUFA may have a positive effect on the expression of key fertility-related genes and pathways, during the critical window of maternal recognition of pregnancy, particularly where animals are underfed.

Label-free quantitative proteomic analysis of M. longissimus dorsi from cattle during dietary restriction and subsequent compensatory growth.

Compensatory growth (CG) is a naturally occurring physiological process whereby an animal has the ability to undergo enhanced growth following a period of restricted feeding. This studies objective was to identify key proteins involved in the expression of CG. Forty Holstein Friesian bulls were equally assigned to one of four groups. R1 and R2 groups were subjected to restricted feed allowance for 125 days (Period 1). A1 and A2 animals had ad libitum access to feed in Period 1. Following Period 1, all animals from R1 and A1 were slaughtered. Remaining animals (R2 and A2) were slaughtered following ad libitum access to feed for successive 55 days (Period 2). M. longissimus dorsi samples were collected at slaughter from all animals. Proteins were isolated from samples and subjected to label-free mass spectrometry proteomic quantification. Proteins which were differentially abundant during CG (n = 39) were involved in cellular binding processes, oxidative phosphorylation and mitochondrial function. There was also evidence for up regulation of three pathways involved in nucleotide biosynthesis. Genetic variants in or regulating genes pertaining to proteins identified in this study may hold potential for use as DNA based biomarkers for genomic selection of animals with a greater ability to undergo CG.

Gene co-expression networks contributing to the expression of compensatory growth in metabolically active tissues in cattle.

Compensatory growth (CG) is an accelerated growth phenomenon which occurs in animals upon re-alimentation following a period of dietary restriction. The objective of this study was to perform gene co-expression analysis on metabolic tissues of animals undergoing CG, in order to elucidate the molecular control governing this phenomenon. Thirty Holstein Friesian bulls were fed a restricted diet for 125 days, after which they received feed ad libitum. Following 55 days of ad libitum feeding all animals were slaughtered. RNAseq and gene co-expression analyses were performed on tissue samples collected at slaughter including liver, rumen papillae and jejunum epithelium tissues. A period of CG resulted in 15 networks of co-expressed genes. One network of genes, involved in proteasome core complex, signal transduction and protein synthesis was found to be similar across liver and jejunum tissue datasets (r = 0.68, P = 0.04). Results from this study also showed that a large portion of co-expressed genes had not previously been implicated in the expression of CG, thus this study identifies novel genes involved in controlling CG across tissues, with hub genes holding potential for use as biomarkers for the selection of animals with a greater propensity to display CG.

Liver transcriptome profiling of beef steers with divergent growth rate, feed intake, or metabolic body weight phenotypes1.

Average daily gain (ADG) and daily dry matter intake (DMI) are key determinants of beef industry profitability. These traits together with metabolic body weight (MWT) are combined as component traits to calculate residual feed intake (RFI), a common measure of feed efficiency in beef cattle. Recently, there have been significant efforts towards molecular genetic characterization of RFI through transcriptomic studies in different breeds and tissues. However, molecular mechanisms of RFI component traits still remain predominately unexplored. Therefore, in the current study, we investigated the hepatic transcriptomic profiles and their associations with ADG, DMI, and MWT in Angus, Charolais, and Kinsella Composite (KC) populations through global RNAseq analyses. In each population and for each trait, 12 steers with extreme phenotypes (n = 6 low and n = 6 high) were analyzed for differential gene expression. These animals were from 20 beef steers of each Angus, Charolais, and KC breed population that were initially selected for a transcriptome study of RFI. At a false discovery rate <0.05 and fold change >1.5, we identified 123, 102, and 78 differentially expressed (DE) genes between high- and low-ADG animals of Angus, Charolais, and KC populations, respectively. For DMI, 108, 180, and 156 DE genes were identified between high- and low-DMI from Angus, Charolais, and KC populations, respectively, while for MWT, 80, 82, and 84 genes were differentially expressed between high- and low-MWT animals in Angus, Charolais, and KC populations, respectively. The identified DE genes were largely breed specific (81.7% for ADG, 82.7% for DMI, and 83% for MWT), but were largely involved in the same biological functions across the breeds. Among the most enriched biological functions included metabolism of major nutrients (lipids, carbohydrates, amino acids, vitamins, and minerals), small molecule biochemistry, cellular movement, cell morphology, and cell-to-cell signaling and interaction. Notably, we identified multiple DE genes that are involved in cholesterol biosynthesis, and immune response pathways for the 3 studied traits. Thus, our findings present potential molecular genetic mechanisms and candidate genes that influence feed intake, growth, and MWT of beef cattle.