Gene Expression Essential for Myostatin Signaling and Skeletal Muscle Development Is Associated With Divergent Feed Efficiency in Pedigree Male Broilers

Background: Feed efficiency (FE, gain to feed) is an important genetic trait as 70% of the cost of raising animals is due to feed costs. The objective of this study was to determine mRNA expression of genes involved in muscle development and hypertrophy, and the insulin receptor-signaling pathway in breast muscle associated with the phenotypic expression of FE.Methods: Breast muscle samples were obtained from Pedigree Male (PedM) broilers (8 to 10 week old) that had been individually phenotyped for FE between 6 and 7 week of age. The high FE group gained more weight but consumed the same amount of feed compared to the low FE group. Total RNA was extracted from breast muscle (n = 6 per group) and mRNA expression of target genes was determined by real-time quantitative PCR.Results: Targeted gene expression analysis in breast muscle of the high FE phenotype revealed that muscle development may be fostered in the high FE PedM phenotype by down-regulation several components of the myostatin signaling pathway genes combined with upregulation of genes that enhance muscle formation and growth. There was also evidence of genetic architecture that would foster muscle protein synthesis in the high FE phenotype. A clear indication of differences in insulin signaling between high and low FE phenotypes was not apparent in this study.Conclusion: These findings indicate that a gene expression architecture is present in breast muscle of PedM broilers exhibiting high FE that would support enhanced muscle development-differentiation as well as protein synthesis compared to PedM broilers exhibiting low FE

Enrichment of Autophagy and Proteosome Pathways in Breast Muscle of Feed Efficient Pedigree Male Broilers

Background: Feed efficiency (FE) is an important genetic trait in poultry and livestock. Autophagy (self-eating) and proteosomes are cellular processes that remove damaged cell components (e.g., proteins, organelles). As evidence of extensive protein oxidation was observed in Pedigree Male (PedM) broilers exhibiting a low FE (LFE) phenotype compared to a high FE (HFE) phenotype, the main goal of this study was to assess gene and protein expression of the autophagy and proteosome pathways in breast muscle obtained in PedM broilers exhibiting HFE and LFE phenotypes.Methods: Feed efficiency was calculated as weight gain divided by feed intake gain in individual PedM broilers that were measured between 6 and 7 weeks of age. Targeted gene expression was conducted on breast muscle using quantitative real-time polymerase chain reaction (qPCR) to determine mRNA expression of genes associated with the autophagy pathway; AMP-activated protein kinase alpha 1 (AMPKα1), mammalian target of rapamycin (mTOR), Beclin 1, and autophagy genes (Atg) 3, Atg7, and Atg16L1. Binomial distribution analysis was conducted on transcriptomic and data obtained by RNAseq and shotgun proteomics, respectively on the same set of tissues for genes associated with autophagy, vacuole formation, and proteosome expression.Results: Greater efficiency was attained in the HFE PedM broilers by greater weight gain on the same amount of feed consumed resulting in FEs of 0.65 ± 0.01 and 0.46 ± 0.01 in the HFE and LFE phenotypes, respectively. Targeted mRNA expression analysis revealed significant (P < 0.05) elevations in AMPKa1, mTOR, Atg16L1, and Atg7 and a marginal (P = 0.07) elevation in Beclin1. Binomial distribution analysis transcriptomic and proteomic data revealed significant skews favoring autophagy-, vacuole-, and proteosome-related genes in the HFE phenotype. These results indicate that the autophagy and proteosome expression is enhanced in the HFE compared to the LFE pedigree male broiler phenotype suggesting that protein and organelle quality control may be enhanced in high feed efficiency

Correction: Proteomics of Breast Muscle Tissue Associated with the Phenotypic Expression of Feed Efficiency within a Pedigree Male Broiler Line: I. Highlight on Mitochondria.

[This corrects the article DOI: 10.1371/journal.pone.0155679.]

Proteogenomics reveals enriched ribosome assembly and protein translation in Pectoralis major of high feed efficiency pedigree broiler males

Background: In production animal agriculture, the cost of feed represents 60-70% of the total cost of raising an animal to market weight. Thus, development of viable biomarkers for feed efficiency (FE, g gain/g feed) to assist in genetic selection of breeding stock remains an important goal in commercial breeding programs. Methods: Global gene (cDNA microarray, RNAseq) and protein expression (shotgun proteomics) analyses have been conducted on breast muscle samples obtained from pedigree broiler males (PedM) exhibiting high and low FE phenotypes. Using the entire datasets (i.e., no cutoffs for significance or fold difference in expression) the number of genes or proteins that were expressed numerically higher or lower in the high FE compared to the low FE phenotype for key terms or functions, e.g., ribosomal, mitochondrial ribosomal, tRNA, RNA binding motif, RNA polymerase, small nuclear ribonucleoprotein, and protein tyrosine phosphatase, were determined. Bionomial distribution analysis (exact) was then conducted on these datasets to determine significance between numerically up or down expression. Results: Processes associated with mitochondrial proteome expression (e.g., mitochondrial ribosomal proteins, mitochondrial transcription, mitochondrial tRNA, and translation) were enriched in breast muscle from the high FE compared to the low FE pedigree male broiler phenotype. Furthermore, the high FE phenotype exhibited enrichment of ribosome assembly (e.g., RNA polymerase, mitochondrial and cytosolic ribosomes, small, and heterogeneous nuclear ribonucleoproteins), as well as nuclear transport and protein translation processes compared to the low FE phenotype. Quality control processes (proteosomes and autophagy) were also enriched in the high FE phenotype. In contrast, the low FE phenotype exhibited enrichment of cytoskeletal proteins, protein tyrosine phosphatases, and tyrosine kinases compared to the high FE phenotype. These results suggest that processes of mitochondrial and cytosolic ribosomal construction, activity, and protein translation would be enhanced in high FE breast muscle, and that phosphorylation of tyrosine moieties of proteins could be prolonged in the high compared to low FE phenotype. The results indicate the presence of a proteogenomic architecture that could enhance ribosome construction, protein translation, and quality control processes and contribute to the phenotypic expression of feed efficiency in this PedM broiler model

Proteomics of breast muscle tissue associated with the phenotypic expression of feed efficiency within a pedigree male broiler line: I. Highlight on mitochondria

The fifth author's name is spelled incorrectly. The correct name is: Antonio Reverter. The correct citation is: Kong B-W, Lassiter K, Piekarski-Welsher A, Dridi S, Reverter A, Hudson NJ, et al. (2016) Proteomics of Breast Muscle Tissue Associated with the Phenotypic Expression of Feed Efficiency within a Pedigree Male Broiler Line: I. Highlight on Mitochondria. PLoS ONE 11(5): e0155679. doi:10.1371/journal.pone.0155679

A hierarchically clustered heat map of the 40 most differentially expressed proteins as identified by PIF.

<p>Each cell contains the normalized expression of the protein at the individual bird level. Red denotes high expression, green denotes low expression. All 8 birds discriminate into their correct treatment group of origin (HFE or LFE) and relatedness within a group is also apparent. Proteins are clustered by patterns of co-expression across the 8 birds. Strong co-expression is observed for functionally related proteins e.g. ARF1 with ARF4 and KRT3 with KRT5 and KRT14 and the muscle structural proteins, namely TPM1, TPM2 and CAPZB. Different fragment of the same protein are highly co-expressed. Note: ‘B’ are birds with a low FE phenotype whereas ‘G’ are birds with a high FE phenotype. <b>Abbreviations:</b> (RTN4), reticulon 4; (NDUFV2), NADH dehydrogenase (ubiquinone) Fe-S protein 7; (PHKG1), phosphorylase kinase gamma 1; (IDE), insulin degrading enzyme; (CACNA15), calcium channel voltage dependent 1 L type, alpha 15; (ARF), ADP ribosylation factor 4 and 1; (SAR1B), SAR1-ADP ribosylation factor, (SPEG), striated muscle preferentially expressed protein; (Gga), golgi associated gamma adaptin; (RYR3) ryanodine receptor 3; (SLC26A4), adenine nucleotide translocase 1; (MYO18A), myosin 18A; (KRT), keratin; (TPM), tropomyosin; (BAT2), large proline rich protein; (CAPZB), F-actin capping protein subunit beta; (DCTN2), dynactin 2; (ACTA1), alpha actin 1; (GSN), gelsolin; (SMYD1), myosin interacting protein; (NEB), lambda protein phosphatase; (OIH) ovoinhibitor; (UBQLN), ubituilin family proteins</p

Abbreviations and names of proteins not found in Table 1 or figure legends.

<p>Abbreviations and names of proteins not found in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0155679#pone.0155679.t001" target="_blank">Table 1</a> or figure legends.</p

Top canonical pathways.

<p>The top 6 canonical pathways (based on p value) generated by Ingenuity Pathway Analysis based on differentially expressed proteins in breast muscle obtained from broiler breeder males.</p

Upstream regulators predicted to be activated or inhibited in the proteomic dataset.

<p>Upstream regulators predicted to be activated or inhibited in the proteomic dataset.</p