Ranran Liu

Identification of differentially expressed genes and pathways for intramuscular fat deposition in pectoralis major tissues of fast-and slow-growing chickens

BackgroundIntramuscular fat (IMF) is one of the important factors influencing meat quality, however, for chickens, the molecular regulatory mechanisms underlying this trait have not yet been determined. In this study, a systematic identification of candidate genes and new pathways related to IMF deposition in chicken breast tissue has been made using gene expression profiles of two distinct breeds: Beijing-you (BJY), a slow-growing Chinese breed possessing high meat quality and Arbor Acres (AA), a commercial fast-growing broiler line.ResultsAgilent cDNA microarray analyses were conducted to determine gene expression profiles of breast muscle sampled at different developmental stages of BJY and AA chickens. Relative to d 1 when there is no detectable IMF, breast muscle at d 21, d 42, d 90 and d 120 (only for BJY) contained 1310 differentially expressed genes (DEGs) in BJY and 1080 DEGs in AA. Of these, 34–70 DEGs related to lipid metabolism or muscle development processes were examined further in each breed based on Gene Ontology (GO) analysis. The expression of several DEGs was correlated, positively or negatively, with the changing patterns of lipid content or breast weight across the ages sampled, indicating that those genes may play key roles in these developmental processes. In addition, based on KEGG pathway analysis of DEGs in both BJY and AA chickens, it was found that in addition to pathways affecting lipid metabolism (pathways for MAPK & PPAR signaling), cell junction-related pathways (tight junction, ECM-receptor interaction, focal adhesion, regulation of actin cytoskeleton), which play a prominent role in maintaining the integrity of tissues, could contribute to the IMF deposition.ConclusionThe results of this study identified potential candidate genes associated with chicken IMF deposition and imply that IMF deposition in chicken breast muscle is regulated and mediated not only by genes and pathways related to lipid metabolism and muscle development, but also by others involved in cell junctions. These findings establish the groundwork and provide new clues for deciphering the molecular mechanisms underlying IMF deposition in poultry. Further studies at the translational and posttranslational level are now required to validate the genes and pathways identified here.

Genome-wide association study identifies Loci and candidate genes for body composition and meat quality traits in Beijing-You chickens.

Body composition and meat quality traits are important economic traits of chickens. The development of high-throughput genotyping platforms and relevant statistical methods have enabled genome-wide association studies in chickens. In order to identify molecular markers and candidate genes associated with body composition and meat quality traits, genome-wide association studies were conducted using the Illumina 60 K SNP Beadchip to genotype 724 Beijing-You chickens. For each bird, a total of 16 traits were measured, including carcass weight (CW), eviscerated weight (EW), dressing percentage, breast muscle weight (BrW) and percentage (BrP), thigh muscle weight and percentage, abdominal fat weight and percentage, dry matter and intramuscular fat contents of breast and thigh muscle, ultimate pH, and shear force of the pectoralis major muscle at 100 d of age. The SNPs that were significantly associated with the phenotypic traits were identified using both simple (GLM) and compressed mixed linear (MLM) models. For nine of ten body composition traits studied, SNPs showing genome wide significance (P<2.59E−6) have been identified. A consistent region on chicken (Gallus gallus) chromosome 4 (GGA4), including seven significant SNPs and four candidate genes (LCORL, LAP3, LDB2, TAPT1), were found to be associated with CW and EW. Another 0.65 Mb region on GGA3 for BrW and BrP was identified. After measuring the mRNA content in beast muscle for five genes located in this region, the changes in GJA1 expression were found to be consistent with that of breast muscle weight across development. It is highly possible that GJA1 is a functional gene for breast muscle development in chickens. For meat quality traits, several SNPs reaching suggestive association were identified and possible candidate genes with their functions were discussed.

The identification of 14 new genes for meat quality traits in chicken using a genome-wide association study.

BackgroundMeat quality is an important economic trait in chickens. To identify loci and genes associated with meat quality traits, we conducted a genome-wide association study (GWAS) of F2 populations derived from a local Chinese breed (Beijing-You chickens) and a commercial fast-growing broiler line (Cobb-Vantress).ResultsIn the present study, 33 association signals were detected from the compressed mixed linear model (MLM) for 10 meat quality traits: dry matter in breast muscle (DMBr), dry matter in thigh muscle (DMTh), intramuscular fat content in breast muscle (IMFBr), meat color lightness (L*) and yellowness (b*) values, skin color L*, a* (redness) and b* values, abdominal fat weight (AbFW) and AbFW as a percentage of eviscerated weight (AbFP). Relative expressions of candidate genes identified near significant signals were compared using samples of chickens with High and Low phenotypic values. A total of 14 genes associated with IMFBr, meat color L*, AbFW, and AbFP, were differentially expressed between the High and Low phenotypic groups. These genes are, therefore, prospective candidate genes for meat quality traits: protein tyrosine kinase (TYRO3) and microsomal glutathione S-transferase 1 (MGST1) for IMFBr; collagen, type I, alpha 2 (COL1A2) for meat color L*; and RET proto-oncogene (RET), natriuretic peptide B (NPPB) and sterol regulatory element binding transcription factor 1 (SREBF1) for the abdominal fat (AbF) traits.ConclusionsBased on the association signals and differential expression of nearby genes, 14 candidate loci and genes for IMFBr, meat L* and b* values, and AbF are identified. The results provide new insight into the molecular mechanisms underlying meat quality traits in chickens.

Epigenetic modification of TLRs in leukocytes is associated with increased susceptibility to Salmonella enteritidis in chickens.

Toll-like receptors (TLRs) signaling pathways are the first lines in defense against Salmonella enteritidis (S. enteritidis) infection but the molecular mechanism underlying susceptibility to S. enteritidis infection in chicken remains unclear. SPF chickens injected with S. enteritidis were partitioned into two groups, one consisted of those from Salmonella-susceptible chickens (died within 5 d after injection, n = 6), the other consisted of six Salmonella-resistant chickens that survived for 15 d after injection. The present study shows that the bacterial load in susceptible chickens was significantly higher than that in resistant chickens and TLR4, TLR2-1 and TLR21 expression was strongly diminished in the leukocytes of susceptible chickens compared with those of resistant chickens. The induction of expression of pro-inflammatory cytokine genes, IL-6 and IFN-β, was greatly enhanced in the resistant but not in susceptible chickens. Contrasting with the reduced expression of TLR genes, those of the zinc finger protein 493 (ZNF493) gene and Toll-interacting protein (TOLLIP) gene were enhanced in the susceptible chickens. Finally, the expression of TLR4 in peripheral blood mononuclear cells (PBMCs) infected in vitro with S. enteritidis increased significantly as a result of treatment with 5-Aza-2-deoxycytidine (5-Aza-dc) while either 5-Aza-dc or trichostatin A was effective in up-regulating the expression of TLR21 and TLR2-1. DNA methylation, in the predicted promoter region of TLR4 and TLR21 genes, and an exonic CpG island of the TLR2-1 gene was significantly higher in the susceptible chickens than in resistant chickens. Taken together, the results demonstrate that ZNF493-related epigenetic modification in leukocytes probably accounts for increased susceptibility to S. enteritidis in chickens by diminishing the expression and response of TLR4, TLR21 and TLR2-1.

FSH stimulates lipid biosynthesis in chicken adipose tissue by upregulating the expression of its receptor FSHR.

Transcripts and protein for follicle-stimulating hormone receptor (FSHR) were demonstrated in abdominal adipose tissue of female chickens. There was no expression of the Fsh gene, but FSH and FSHR colocalized, suggesting that FSH was receptor bound. Partial correlations indicted that changes in abdominal fat (AF) content were most directly correlated with Fshr mRNA expression, and the latter was directly correlated with tissue FSH content. These relationships were consistent with FSH inducing Fshr mRNA expression and with the finding that FSH influenced the accumulation of AF in chickens, a novel role for the hormone. Chicken preadipocytes responded linearly to doubling concentrations of FSH in Fshr mRNA expression and quantities of FSHR and lipid, without discernable effect on proliferation. Cells exposed to FSH more rapidly acquired adipocyte morphology. Treatment of young chickens with chicken FSH (4 mIU/day, subcutaneous, days 7–13) did not significantly decrease live weight but increased AF weight by 54.61%, AF as a percentage of live weight by 55.45%, and FSHR transcripts in AF by 222.15% (2 h after injection). In cells stimulated by FSH, genes related to lipid metabolism, including Rdh10, Dci, RarB, Lpl, Acsl3, and Dgat2, were expressed differentially, compared with no FSH. Several pathways of retinal and fatty acid metabolism, and peroxisome proliferator-activated receptor (PPAR) signaling changed. In conclusion, FSH stimulates lipid biosynthesis by upregulating Fshr mRNA expression in abdominal adipose tissue of chickens. Several genes involved in fatty acid and retinal metabolism and the PPAR signaling pathway mediate this novel function of FSH.

Comparison of breast muscle meat quality in 2 broiler breeds

On the basis of meat quality traits, muscle fiber characteristics, and nutrient components and contents in chickens at market age, 120-d-old Beijing-you (BJY) chickens (the Chinese local breed) had distinct breast muscle features when compared with 42-d-old Arbor Acres (AA) chickens (the genetically improved broiler line). The phospholipid (P < 0.05) and essential fatty acid (P < 0.05) contents in BJY chickens were significantly higher than those in AA chickens. No differences (P > 0.05) were found between the breeds in the contents of polyunsaturated fatty acids, unsaturated fatty acids, protein, or amino acids. Breast muscle fiber diameter was significantly smaller (~55.76%) and fiber density was higher (~174.86%) in BJY chickens than in AA chickens (P < 0.05). In this study, breast muscle from 120-d-old BJY chickens was judged to have better quality of phospholipids and essential fatty acid contents and muscle fiber characteristics than breast muscle from 42-d-old AA chickens.

Integrated analysis of microRNA and mRNA expression profiles in abdominal adipose tissues in chickens.

Excessive fat accretion is a crucial problem during broiler production. Abdominal fat weight (AbFW) and abdominal fat percentage (AbFP) are major phenotypic indices of fat traits. The present study used F2 females derived from a cross between Beijing-You and Cobb-Vantress chickens. Cohorts with extreme AbFP and AbFW phenotypes were chosen to construct high- and low-abdominal fat libraries (HAbF and LAbF, respectively) to investigate the expression profiles by RNA-sequencing and microRNA (miRNA)-sequencing. Compared with the LAbF library, 62 differentially expressed miRNAs (DEMs) and 303 differentially expressed genes (DEGs) were identified in the HAbF birds. Integrated analysis of DEMs and DEGs showed that a total of 106 DEGs were identified as target genes for the 62 DEMs. These genes were designated as intersection genes, and 11 of these genes are involved in lipid metabolism pathways. The miRNA gga-miR-19b-3p accelerated the proliferation of preadipocytes, as well as adipocyte differentiation, by down- regulating ACSL1. These findings suggest that some strong candidate miRNAs and genes, important in relation to abdominal adipose deposition, were identified by the integrated analysis of DEMs and DEGs. These findings add to our current understanding of the molecular genetic controls underlying abdominal adipose accumulation in chickens.

Genome-Wide Linkage Analysis and Association Study Identifies Loci for Polydactyly in Chickens

Polydactyly occurs in some chicken breeds, but the molecular mechanism remains incompletely understood. Combined genome-wide linkage analysis and association study (GWAS) for chicken polydactyly helps identify loci or candidate genes for the trait and potentially provides further mechanistic understanding of this phenotype in chickens and perhaps other species. The linkage analysis and GWAS for polydactyly was conducted using an F2 population derived from Beijing-You chickens and commercial broilers. The results identified two QTLs through linkage analysis and seven single-nucleotide polymorphisms (SNPs) through GWAS, associated with the polydactyly trait. One QTL located at 35 cM on the GGA2 was significant at the 1% genome-wise level and another QTL at the 1% chromosome-wide significance level was detected at 39 cM on GGA19. A total of seven SNPs, four of 5% genome-wide significance (P < 2.98 × 10−6) and three of suggestive significance (5.96 × 10−5) were identified, including two SNPs (GGaluGA132178 and Gga_rs14135036) in the QTL on GGA2. Of the identified SNPs, the eight nearest genes were sonic hedgehog (SHH), limb region 1 homolog (mouse) (LMBR1), dipeptidyl-peptidase 6, transcript variant 3 (DPP6), thyroid-stimulating hormone, beta (TSHB), sal-like 4 (Drosophila) (SALL4), par-6 partitioning defective 6 homolog beta (Caenorhabditis elegans) (PARD6B), coenzyme Q5 (COQ5), and tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein, etapolypeptide (YWHAH). The GWAS supports earlier reports of the importance of SHH and LMBR1 as regulating genes for polydactyly in chickens and other species, and identified others, most of which have not previously been associated with limb development. The genes and associated SNPs revealed here provide detailed information for further exploring the molecular and developmental mechanisms underlying polydactyly.

Effects of diet-induced differences in growth rate on metabolic, histological, and meat-quality properties of 2 muscles in male chickens of 2 distinct broiler breeds

This paper, the second from a comprehensive study, describes the effects of varying growth rate by feeding at different planes of nutrition with a constant ME:CP ratio on muscle characteristics and meat quality in 2 distinct broiler breeds of male chickens (Arbor Acres, a commercial line; and Beijing-You, a Chinese nonimproved line). Experimental diets, differing on average by 2% CP, were formulated with high-, medium-, or low-nutrient densities for 3 growing phases. Male hatchlings (216 of each breed) were randomly assigned to 6 pens of 12 birds in each treatment. Altered histological characteristics of muscle fibers, early postmortem muscle metabolism, and meat quality were investigated in the pectoralis major and biceps femoris. At their market age, Arbor Acres broilers had significantly higher concentrations of plasma protein and lipid metabolites, ratios of white to red and intermediate fibers, pH, L* and b* values, and lower concentrations of plasma glucose metabolites, muscle-fiber diameter, muscle contents of energy stores, a* value, drip loss, and shear force than the values found for the Beijing-You chickens (P < 0.01). Higher nutrient density increased the size of the muscle fibers, decreased glycogen reserve, and reduced the rate and extent of acidification in the Arbor Acres chickens, while accelerating transformation of red and intermediate-to-white fibers, enhancing energy stores, and hastening the decrease in pH postmortem in the Beijing-You chickens (P < 0.05). In each breed, most meat quality variables (e.g., shear force, drip loss, and color) were consistent with the histological and biochemical changes caused by the feeding strategy. Together, dietary nutrient density can influence meat quality as a result of altered histological and initial energy and metabolic characteristics of the muscle. Many of the responses to diet are breed and tissue dependent in broiler chickens.

Up-regulation of the MyD88-dependent pathway of TLR signaling in spleen and caecum of young chickens infected with Salmonella serovar Pullorum.

TLR signaling plays a role in Salmonella infection, but less information is available in chickens infected with Salmonella serovar Pullorum. The present study with young chickens, experimentally infected with S. Pullorum, has used real-time quantitative RT-PCR to investigate the relative expression of genes of the TLR4 signaling pathway (TLR4, MyD88, TRAF6 and NF-kappaB) in the spleen and caecum at 1, 3, 7 and 14 days post-infection (dpi). Three-day-old specific-pathogen-free chickens were orally infected with S. Pullorum or saline (controls). In addition to gene expression, the bacterial burden of spleen and caecum was assessed and serum concentrations and total IgG were measured. Significantly more bacteria were found in the caecum than in the spleen. Concentrations of IgG were significantly higher in infected birds at 1 dpi, reached a peak at 3 dpi (P<0.01), and then gradually decreased with the time. Infected young chickens had significant up-regulation of the expression of TLR4, MyD88, TRAF6 and NF-kappaB in the spleen at 3 dpi compared to controls (P<0.05). In the caecum, only increased expression of NF-kappaB mRNA (P<0.05) was observed at 1 dpi. The pattern of immune response to infection with S. Pullorum differed between spleen and caecum and did not directly correspond with pathogen burden. The observed changes indicate that the MyD88-dependent pathway of TLR4 signaling plays a role in young chickens infected with S. Pullorum.