Longissimus dorsi muscle transcriptome in pure and crossbred Iberian pigs at birth and growing stages
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ABSTRACT: Iberian ham production includes both purebred (IB) and Duroc-crossbred (IBxDU) Iberian pigs, which show important differences in meat quality and production traits, such as muscle growth and fatness. This experiment was conducted to investigate gene expression differences that could be associated with the observed phenotypic differences between IB and IBxDU pigs. Nine IB and 10 IBxDU pigs were slaughtered at birth, and seven IB and 10 IBxDU at four months of age (growing period). Muscle transcriptome was analyzed on LD samples with RNA-Seq technology. We detected 261 differentially expressed genes between IB and IBxDU neonates and 113 at growth (p < 0.01 and Fold-Change > 1. 5). Newborns transcriptome reflected a highly proliferative developmental stage, while older pigs showed upregulation of catabolic and muscle functioning processes. Regarding the genetic type effect, IBxDU newborns showed enrichment of gene pathways involved in muscle growth, in agreement with the higher prenatal growth observed in these pigs. However, IB growing pigs showed enrichment of pathways involved in protein deposition and cellular growth, supporting the catch-up growth experienced by IB pigs during this period. Moreover, newborn and growing IB pigs showed more active glucose and lipid metabolism than IBxDU pigs.
Project description:Iberian ham production includes both purebred (IB) and Duroc-crossbred (IBxDU) Iberian pigs, which show important differences in meat quality and production traits, such as muscle growth and fatness. This experiment was conducted to investigate gene expression differences, transcriptional regulation and genetic polymorphisms that could be associated with the observed phenotypic differences between IB and IBxDU pigs. Nine IB and 10 IBxDU pigs were slaughtered at birth. Morphometric measures and blood samples were obtained and samples from Biceps femoris muscle were employed for compositional and transcriptome analysis by RNA-Seq technology. Phenotypic differences were evident at this early age, including greater body size and weight in IBxDU and greater Biceps femoris intramuscular fat and plasma cholesterol content in IB newborns. We detected 150 differentially expressed genes between IB and IBxDU neonates (p < 0.01 and Fold-Change > 1. 5). Several were related to adipose and muscle tissues development (DLK1, FGF21 or UBC). The functional interpretation of the transcriptomic differences revealed enrichment of functions and pathways related to lipid metabolism in IB and to cellular and muscle growth in IBxDU. Protein catabolism, cholesterol biosynthesis and immune system were functions enriched in both genotypes. We identified transcription factors potentially affecting the observed gene expression differences. Some of them have known functions on adipogenesis (CEBPA, EGRs), lipid metabolism (PPARGC1B) and myogenesis (FOXOs, MEF2D, MYOD1), which suggest a key role in the meat quality differences existing between IB and IBxDU hams. We also identified several polymorphisms showing differential segregation between IB and IBxDU pigs. Among them, non-synonymous variants were detected in several transcription factors as PPARGC1B gene, which could be associated to altered gene function. Taken together, these results provide information about candidate genes, metabolic pathways and genetic polymorphisms potentially involved in phenotypic differences between IB and IBxDU associated to meat quality and production traits.
Project description:We studied the influence of genetic type (pure Iberian pigs vs crossbred with Duroc) on l.dorsi transcriptome Background: The two main genetic types in Iberian pig production show important phenotypic differences in growth, fattening and tissue composition since early developmental stages. The objective of this work was the evaluation of muscle transcriptome profile in young animals of both genetic types, in order to identify genes, pathways and transcription factors responsible of their phenotypic differences. Contemporary families coming from pure Iberian pigs (IB) or from crossing with Duroc boars (DUxIB) were generated. Piglets (14 from each genetic type) were sacrificed at weaning (28 days) and longissimus dorsi muscle was sampled for composition and gene expression studies. RNA was obtained and hybridized to Affymetrix Porcine Genechip expression arrays. Results: Loin muscle composition showed significant differences between genetic types in intramuscular fat content (6% vs. 4.2% in IB and DUxIB animals, respectively, P=0.009) and in saturated and monounsaturated fatty acid contents (P=0.019 and P=0.044, respectively). Statistical analysis of gene expression data allowed the identification of 256 differentially expressed (DE) genes between genetic types (FDR<0.10), 102 upregulated in IB and 154 upregulated in DUxIB. The size of the effects ranged between 1.2x and 7.8x. Transcript differences were validated for a subset of DE genes by qPCR. Some candidate genes with known effects on muscle growth were found among the DE genes upregulated in DUxIB (IGF2, collagens, integrins). Genes related to lipid metabolism and proteolysis were found among the genes upregulated in IB (ME1, ELOVL6, caspases, catepsins, ubiquitination proteins). We observed alteration in biological functions related to extracellular matrix organization, cellular adhesion, muscle growth and lipid metabolism. Transcription factors (TF) potentially involved in the expression differences found were identified by calculating the regulatory impact factors (RIF). Sixteen TF were found, some of them with known relationship with muscle development (MSTN, SIX4), adipogenesis (CEBPD, PPARGC1B), or extracellular matrix processes (MAX, MXI1). Correlation among the expression of these TF and DE genes show relevant differences between genetic types. Conclusion: The results contribute valuable information about molecular mechanisms determining the phenotypic differences of growth and meat quality between the genetic types studied, mainly related to the development and function of the extracellular matrix and also to some metabolic processes as proteolysis and lipid metabolism. 28 male piglets were employed. 14 animals coming for pure Iberian parents and 14 coming from the crossing of Duroc boars with Iberian sows. Animals were sacrificed at 28 d of age. At the sacrifice, longissimus dorsi muscle tissue was sampled from each animal.
Project description:We studied the influence of genetic type (pure Iberian pigs vs crossbred with Duroc) on l.dorsi transcriptome Background: The two main genetic types in Iberian pig production show important phenotypic differences in growth, fattening and tissue composition since early developmental stages. The objective of this work was the evaluation of muscle transcriptome profile in young animals of both genetic types, in order to identify genes, pathways and transcription factors responsible of their phenotypic differences. Contemporary families coming from pure Iberian pigs (IB) or from crossing with Duroc boars (DUxIB) were generated. Piglets (14 from each genetic type) were sacrificed at weaning (28 days) and longissimus dorsi muscle was sampled for composition and gene expression studies. RNA was obtained and hybridized to Affymetrix Porcine Genechip expression arrays. Results: Loin muscle composition showed significant differences between genetic types in intramuscular fat content (6% vs. 4.2% in IB and DUxIB animals, respectively, P=0.009) and in saturated and monounsaturated fatty acid contents (P=0.019 and P=0.044, respectively). Statistical analysis of gene expression data allowed the identification of 256 differentially expressed (DE) genes between genetic types (FDR<0.10), 102 upregulated in IB and 154 upregulated in DUxIB. The size of the effects ranged between 1.2x and 7.8x. Transcript differences were validated for a subset of DE genes by qPCR. Some candidate genes with known effects on muscle growth were found among the DE genes upregulated in DUxIB (IGF2, collagens, integrins). Genes related to lipid metabolism and proteolysis were found among the genes upregulated in IB (ME1, ELOVL6, caspases, catepsins, ubiquitination proteins). We observed alteration in biological functions related to extracellular matrix organization, cellular adhesion, muscle growth and lipid metabolism. Transcription factors (TF) potentially involved in the expression differences found were identified by calculating the regulatory impact factors (RIF). Sixteen TF were found, some of them with known relationship with muscle development (MSTN, SIX4), adipogenesis (CEBPD, PPARGC1B), or extracellular matrix processes (MAX, MXI1). Correlation among the expression of these TF and DE genes show relevant differences between genetic types. Conclusion: The results contribute valuable information about molecular mechanisms determining the phenotypic differences of growth and meat quality between the genetic types studied, mainly related to the development and function of the extracellular matrix and also to some metabolic processes as proteolysis and lipid metabolism.
Project description:Low protein diets supplied during the growing period of pigs can diminish their growth rate and increase the intramuscular fat (IMF) content which affects the sensorial and technological characteristics of the traits. In the present study, the effects of a low protein diet supplied during the growing diet of Duroc x Iberian crossbred pigs on several phenotypic traits and liver and longissimus dorsi transcriptome, were analysed 20 days after the differential treatment was started (EARLY) and at the end of it (FINAL). A total of 20 crossbred pigs were assigned to two different dietary treatments during the growing period: a control diet (C) and a low protein diet (LP) with the same energy and lower levels of raw protein (11%) and lysine (0.60%). The transcriptomes of liver and longissimus dorsi were quantified through RNAseq. A total of 134 differentially expressed annotated genes and new isoforms (DEGs) between C and LP diets in liver of EARLY animals; 480 DEGs in liver of LATE animals, and 128 DEGs and 68 DEGs in longissimus dorsi of EARLY and LATE animals were detected. The functional analyses revealed that low protein diet diminishes the expression in liver of genes codifying for proteins involved in immune system both in EARLY and LATE animals, affects the expression of genes involved in cholesterol homeostasis in liver and in the energy process and growth in longissimus dorsi. Pigs fed with LP diet had not higher IMF content than C ones, although some lipogenesis genes such as FASN, SCD or SREBF1 were higher expressed on their liver. A low protein diet supplied during growing period affects multiple biological process that could compromise the immune and energy state of the Duroc x Iberian crossbred pigs. These results point out that we should be very cautious before implementing this type of regime in Duroc x Iberian pigs.
Project description:Background: Transcriptome variability is due to genetic and environmental causes, much like any other complex phenotype. Ascertaining the transcriptome differences between individuals is an important step to understand how selection and genetic drift may affect gene expression. To that end, extant divergent livestock breeds offer an ideal genetic material. Results: We have analyzed with microarrays five tissues from the endocrine axis (hypothalamus, adenohypophysis, thyroid gland, gonads and fat tissue) of 16 pigs from both sexes pertaining to four extreme breeds (Duroc, Large White, Iberian and a cross with SinoEuropean hybrid line). Using a Bayesian linear model approach, we observed that the largest breed variability corresponded to the male gonads, and was larger than at the remaining tissues, including ovaries. Measurement of sex hormones in peripheral blood at slaughter did not detect any breed-related differences. Not unexpectedly, the gonads were the tissue with the largest number of sex biased genes. There was a strong correlation between sex and breed bias expression, although the most breed biased genes were not the most sex biased genes. A combined analysis of connectivity and differential expression suggested three biological processes as being primarily different between breeds: spermatogenesis, muscle differentiation and several metabolic processes. Conclusion: These results suggest that differences across breeds in gene expression of the male gonads are larger than in other endocrine tissues in the pig. Nevertheless, the strong presence of breed biased genes in the male gonads cannot be explained solely by changes in spermatogenesis nor by differences in the reproductive tract development. Experiment Overall Design: Sixteen animals, four from each of four breeds, Large White (LW), Duroc (DU), Youli (YL) and Iberian (IB) piglets were sampled. These breeds represent a wide genetic variability in current pig breeding schemes. There were two males and two females per breed except in Youli, represented by three males and one female. Animals were bought from three breeding companies and transferred to the University experimental farms at weaning, i.e., aged one month approximately. Pigs were housed simultaneously, fed the same diets during the fattening period, that lasted two months, and were weighed at weekly intervals. At the time of slaughter, the average ages were 87, 83, 80 and 89 days for Large White, Duroc, Youli and Iberian pigs, respectively. Their mean live weights at that time were 27.2 (LW), 23.1 (DU), 18.9 (YL) and 17.4 kg (IB). Experiment Overall Design: Animals were euthanized, after 24h fasting, by an overdose of intravenous sodium thiobarbital. At necropsy, tissue samples were collected, snap frozen in liquid nitrogen and stored at -80 ºC. The average time gap between euthanasia and tissue collection was ~ 15 minutes, maximum time was 25 minutes. The tissues collected were hypothalamus (HYPO), adenohypophysis (AHYP), which was separated from the neurohypophysis, thyroid gland (THYG), gonads (GONA) from both sexes, and back fat tissue (FATB). The hypothalamus included the mamillary body and grey tubercle but excluded the chiasma opticum. Throughout this work, each sample was identified by the acronym of the tissue followed by the animal id, e.g., FATBLWF1 refers to back fat tissue from female 1 Large White.
Project description:Meat color is the first perceived sensory feature and one of the most important quality traits. Myoglobin is the main pigment in meat, giving its characteristic cherry-red color which is highly appreciated by consumers. In the current study, we used the RNA-seq technique to characterize the longissimus dorsi muscle transcriptome in two groups of Iberian pigs with divergent breeding values for myoglobin content. As a result, we identified 57 differentially expressed genes and transcripts (DEG). Moreover, we have validated the RNA-seq expression of a set of genes by qPCR. Functional analyses revealed an enrichment of DEGs in biological process related to oxidation (HBA1), lipid metabolism (ECH1, PLA2G10, PLD2), inflammation (CHST1, CD209, PLA2G10), and immune system (CD209, MX2, LGALS3, LGALS9). The upstream analysis showed a total of five transcription regulatory factors and eight master regulators that could regulate the expression of some DEGs, highlighting SPI1 and MAPK1, since they regulate the expression of DEGs involved in immune defence and inflammatory processes. Iberian pigs with high myoglobin content also showed higher expression of the HBA1 gene and it has been described that both molecules have a protective effect against oxidative and inflammatory processes. Therefore, the HBA1 gene is a very promising candidate gene to harbour polymorphisms underlying the myoglobin content, whereby, further studies should be carried out for its potential use in an Iberian pig selection program.
Project description:Obese and lean pig breeds showed obvious phenotypic variations and physiological differences in skeletal muscle growth. Prenatal muscle development programs postnatal performance. In this study we initially conducted full transcriptional profiling of prenatal skeletal muscle from Tongcheng pigs (obese) and Landrace pigs (lean) at 33, 65 and 90 days post coitus (dpc), using long serial analysis of gene expression (LongSAGE). We subsequently sequenced 317,115 LongSAGE tags and identified 1400 and 1201 differentially expressed transcripts that showed eight expression patterns for Tongcheng and Landrace, respectively. These two breeds had more significant differences in their gene expression profiles at 65 than that at 33 and 90 dpc. We also identified 532, 653 and 459 transcripts that were differentially expressed at 33, 65 and 90 dpc between the two breeds, respectively. The cellular function of the differentially expressed transcripts that matched annotated genes revealed that each stage had a uniquely altered profile between the two breeds in various functional categories including muscle fiber constitute and contraction, apoptosis, protein synthesis, and signalling transduction. Our results suggest that skeletal muscle development potentially has a greater lag in its growth rate at 33-65 dpc in Tongcheng pigs when compared with their Landrace counterparts. Our analyses therefore not only will provide valuable resources in the further identification of candidate genes for meat production traits, but also assist in the elucidation of the development of prenatal skeletal muscle in pigs and other vertebrates. Keywords: comparative transcriptome analysis
Project description:Growth is dependent on genotype and diet, even at early developmental stages. In this study, we investigated the effects of genotype, sex, and body weight on the fetal muscle transcriptome of purebred Iberian and crossbred Iberian x Large White pigs sharing the same uterine environment. RNA sequencing was performed on 16 purebred and crossbred fetuses with high body weight (340±14g and 415±14g, respectively) and 16 with low body weight (246±14g and 311±14g, respectively), on gestational day 77. Genotype had the greatest effect on gene expression, with 645 genes identified as differentially expressed (DE) between purebred and crossbred animals. Functional analysis showed differential regulation of pathways involved in energy and lipid metabolism, muscle development, and tissue disorders. In purebred animals, fetal body weight was associated with 35 DE genes involved in development, lipid metabolism and adipogenesis. In crossbred animals, fetal body weight was associated with 60 DE genes involved in muscle development, viability, and immunity. Interestingly, the results suggested an interaction genotype*weight for some DE genes. Fetal sex had only a modest effect on gene expression. This study allowed the identification of genes, metabolic pathways, biological functions and regulators related to fetal genotype, weight and sex, in animals sharing the same uterine environment. Our findings contribute to a better understanding of the molecular events that influence prenatal muscle development and highlight the complex interactions affecting transcriptional regulation during development.
Project description:This study was performed to determine the effects of dietary fat sources, i.e., beef tallow, soybean oil, olive oil and coconut oil (each 3% in feed), on the growth performance, meat quality and gene expression in growing-finishing pigs. The results of this study indicate that the type of dietary fat affects fatty acid composition and insulin signaling-related gene expression in the longissimus dorsi muscle of pigs.