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Liver transcriptome profile in pigs with extreme phenotypes of intramuscular fatty acid composition

ABSTRACT: The liver transcriptomes of two female groups (High and Low) with phenotypically extreme intramuscular fatty acid composition were sequenced using RNA-Seq [accn: SRA053452, subid: 86092, Bioproject: PRJNA168072]. A total of 146 and 180 unannotated protein-coding genes were identified in intergenic regions for the L and H groups, respectively. In addition, a range of 5.8 to 7.3% of repetitive elements was found, with SINEs being the most abundant elements. The expression in liver of 186 (L) and 270 (H) lncRNAs was also detected. The higher reproducibility of the RNA-Seq data was validated by RT-qPCR and porcine expression microarrays, therefore showing a strong correlation between RT-qPCR and RNA-Seq data (ranking from 0.79 to 0.96), as well as between microarrays and RNA-Seq (r=0.72). A differential expression analysis between H and L animals identified 55 genes differentially-expressed between groups. Pathways analysis revealed that these genes belong to biological functions, canonical pathways and three gene networks related to lipid and fatty acid metabolism. In concordance with the phenotypic classification, the pathways analysis inferred that linolenic and arachidonic acids metabolism was altered between extreme individuals. In addition, a connection was observed among the top three networks, hence suggesting that these genes are interconnected and play an important role in lipid and fatty acid metabolism.

ORGANISM(S): Sus scrofa

PROVIDER: GSE38588 | GEO | 2013/07/16

SECONDARY ACCESSION(S): PRJNA168198

REPOSITORIES: GEO

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Project description:In pigs, adipose tissue is one of the principal organs involved in the regulation of lipid metabolism. It is particulary involved in the overall fatty acid synthesis with consequences in other lipid-target organs such as muscles and the liver. With this in mind, we have used massive, parallel high-throughput sequencing technologies to characterize the porcine adipose tissue transcriptome architecture in six Iberian x Landrace crossbred pigs showing extreme phenotypes for intramuscular fatty acid composition (three per group). High-throughput RNA sequencing was used to generate a whole characterization of adipose tissue (backfat) transcriptome. A total of 4,130 putative unannotated protein-coding sequences were identified in the 20% of reads which mapped in intergenic regions. Furthermore, 36% of the unmapped reads were represented by interspersed repeats, SINEs being the most abundant elements. Differential expression analyses identified 396 candidate genes among divergent animals for intramuscular fatty acid composition. Sixty-two percent of these genes (247/396) presented higher expression in the group of pigs with higher content of intramuscular SFA and MUFA, while the remaining 149 showed higher expression in the group with higher content of PUFA. Pathway analysis related these genes to biological functions and canonical pathways controlling lipid and fatty acid metabolisms. In concordance with the phenotypic classification of animals, the major metabolic pathway differentially modulated between groups was de novo lipogenesis, the group with more PUFA being the one that showed lower expression of lipogenic genes. These results will help in the identification of genetic variants at loci that affect fatty acid composition traits. The implications of these results range from the improvement of porcine meat quality traits to the application of the pig as an animal model of human metabolic diseases. The supplementary files contains the five arrays reported in the paper. Backfat from five animals were assayed with high-density oligonucleotide microarray chips (GeneChipM-BM-. Porcine) from Affymetrix, in order to validate RNA-Seq data.

Project description:Purpose: The aim of present research was to identify miRNAs potentially related with fattness traits in pigs Methods: miRNA-seq analysis was performed on subcutaneous fat samples collected from 22 pigs representing two sire-line breeds – Pietrain and Hampshire and one dam-line – Large White. Based on dissection data, pigs were selected from a larger population in terms of fatness traits to obtain the most extreme groups in each breed. The cDNA libraries were constructed form 300 ng of total RNA with the use NEBNext Multiplex Small RNA Library Prep Set for Illumina (New England Biolabs, Ipswich, MA, United States) according to the protocol. The quantification of obtained libraries was performed on Qubit 2.0 (Invitrogen, Life Technologies) and TapeStation 2200 (D1000 ScreenTape; Agilent). The RNA-seq was performed in 36 single-end cycles on HiScanSQ platform (Illumina) with the use of ruSeq SR Cluster Kit v3- CBOT-HS and TruSeq SBS Kit v 3 - HS (Illumina). Each library was sequences in 4 technical replications. The DEGs were detected using DESEq2 software and validation was performed by qPCR. Results: RNA-seq approach allowed to identify miRNAs differentially expressed between pigs with various fatness traits in each analyzed breeds: 64 for Pietrain pigs; 41 for Hampshire and 46 for Large White pigs (pvalue <0.05; fold change≤1.5). The comparison of obtained miRNAs sets showed 13 miRNAs identified in all three breeds. The Gene Ontology analysis (mirPath v.3.0 DIANA Tools web with DIANA—TarBase v7.0 as a reference) confirmed that detected miRNAs were involved in fatty acid biosynthesis (pvalue <1e-325); fatty acid metabolism (pvalue 6.661338e-16); ECM-receptor interaction (pvalue <1e-325). The most important targeted genes regulated by identified miRNAs were: fatty acid synthase – FASN; Acetyl-CoA carboxylase 1 – ACACA; Malonyl CoA-acyl carrier protein transacylase – MCAT and acetyl-CoA acyltransferase 1 and 2 (ACAA1 and ACAA2 genes). Conclusions: Obtained results allow to propose the panel of miRNAs, which can be related with fatness in pig. Such analysis can be the basis of future research in terms of identification of molecular mechanisms related with adipogenesis and fatness traits in pigs.

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Liver transcriptome profile in pigs with extreme phenotypes of intramuscular fatty acid composition

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