Transcriptional analysis of abdominal fat in genetically fat and lean chickens reveals adipokines, lipogenic genes and a link between hemostasis and leanness

This descriptive study of the abdominal fat transcriptome takes advantage of two experimental lines of meat-type chickens (Gallus domesticus), which were selected over seven generations for a large difference in abdominal (visceral) fatness. At the age of selection (9 wk), the fat line (FL) and lean line (LL) chickens exhibit a 2.5-fold difference in abdominal fat weight, while their feed intake and body weight are similar. These unique avian models were originally created to unravel genetic and endocrine regulation of adiposity and lipogenesis in meat-type chickens. The Del-Mar 14K Chicken Integrated Systems microarray was used for a time-course analysis of gene expression in abdominal fat of FL and LL chickens during juvenile development (1–11 weeks of age). Microarray analysis of abdominal fat in FL and LL chickens revealed 131 differentially expressed (DE) genes (FDR≤0.05) as the main effect of genotype, 254 DE genes as an interaction of age and genotype and 3,195 DE genes (FDR≤0.01) as the main effect of age. The most notable discoveries in the abdominal fat transcriptome were higher expression of many genes involved in blood coagulation in the LL and up-regulation of numerous adipogenic and lipogenic genes in FL chickens. Many of these DE genes belong to pathways controlling the synthesis, metabolism and transport of lipids or endocrine signaling pathways activated by adipokines, retinoid and thyroid hormones. The present study provides a dynamic view of differential gene transcription in abdominal fat of chickens genetically selected for fatness (FL) or leanness (LL). Remarkably, the LL chickens over-express a large number of hemostatic genes that could be involved in proteolytic processing of adipokines and endocrine factors, which contribute to their higher lipolysis and export of stored lipids. Some of these changes are already present at 1 week of age before the divergence in fatness. In contrast, the FL chickens have enhanced expression of numerous lipogenic genes mainly after onset of divergence, presumably directed by multiple transcription factors. This transcriptional analysis shows that abdominal fat of the chicken serves a dual function as both an endocrine organ and an active metabolic tissue, which could play a more significant role in lipogenesis than previously thought.https://doi.org/10.1186/1471-2164-14-55

Use of high-density SNP data to identify patterns of diversity and signatures of selection in broiler chickens

The development of broiler chickens over the last 70 years has been accompanied by large phenotypic changes, so that the resulting genomic signatures of selection should be detectable by current statistical techniques with sufficiently dense genetic markers. Using two approaches, this study analysed high‐density SNP data from a broiler chicken line to detect low‐diversity genomic regions characteristic of past selection. Seven regions with zero diversity were identified across the genome. Most of these were very small and did not contain many genes. In addition, fifteen regions were identified with diversity increasing asymptotically from a low level. These regions were larger and thus generally included more genes. Several candidate genes for broiler traits were found within these ‘regression regions’, including IGF1,GPD2 and MTNR1AI. The results suggest that the identification of zero‐diversity regions is too restrictive for characterizing regions under selection, but that regions showing patterns of diversity along the chromosome that are consistent with selective sweeps contain a number of genes that are functional candidates for involvement in broiler development. Many regions identified in this study overlap or are close to regions identified in layer chicken populations, possibly due to their shared precommercialization history or to shared selection pressures between broilers and layers

Combination of novel and public RNA-seq datasets to generate an mRNA expression atlas for the domestic chicken

Background: The domestic chicken (Gallus gallus) is widely used as a model in developmental biology and is also an important livestock species. We describe a novel approach to data integration to generate an mRNA expression atlas for the chicken spanning major tissue types and developmental stages, using a diverse range of publicly-archived RNA-seq datasets and new data derived from immune cells and tissues. Results: Randomly down-sampling RNA-seq datasets to a common depth and quantifying expression against a reference transcriptome using the mRNA quantitation tool Kallisto ensured that disparate datasets explored comparable transcriptomic space. The network analysis tool Graphia was used to extract clusters of co-expressed genes from the resulting expression atlas, many of which were tissue or cell-type restricted, contained transcription factors that have previously been implicated in their regulation, or were otherwise associated with biological processes, such as the cell cycle. The atlas provides a resource for the functional annotation of genes that currently have only a locus ID. We cross-referenced the RNA-seq atlas to a publicly available embryonic Cap Analysis of Gene Expression (CAGE) dataset to infer the developmental time course of organ systems, and to identify a signature of the expansion of tissue macrophage populations during development. Conclusion: Expression profiles obtained from public RNA-seq datasets - despite being generated by different laboratories using different methodologies - can be made comparable to each other. This meta-analytic approach to RNA-seq can be extended with new datasets from novel tissues, and is applicable to any species

Transcriptional analysis of abdominal fat in chickens divergently selected on bodyweight at two ages reveals novel mechanisms controlling adiposity: validating visceral adipose tissue as a dynamic endocrine and metabolic organ

Decades of intensive genetic selection in the domestic chicken (Gallus gallus domesticus) have enabled the remarkable rapid growth of today’s broiler (meat-type) chickens. However, this enhanced growth rate was accompanied by several unfavorable traits (i.e., increased visceral fatness, leg weakness, and disorders of metabolism and reproduction). The present descriptive analysis of the abdominal fat transcriptome aimed to identify functional genes and biological pathways that likely contribute to an extreme difference in visceral fatness of divergently selected broiler chickens. We used the Del-Mar 14 K Chicken Integrated Systems microarray to take time-course snapshots of global gene transcription in abdominal fat of juvenile [1-11 weeks of age (wk)] chickens divergently selected on bodyweight at two ages (8 and 36 wk). Further, a RNA sequencing analysis was completed on the same abdominal fat samples taken from high-growth (HG) and low-growth (LG) cockerels at 7 wk, the age with the greatest divergence in body weight (3.2-fold) and visceral fatness (19.6-fold). Time-course microarray analysis revealed 312 differentially expressed genes (FDR ≤ 0.05) as the main effect of genotype (HG versus LG), 718 genes in the interaction of age and genotype, and 2918 genes as the main effect of age. The RNA sequencing analysis identified 2410 differentially expressed genes in abdominal fat of HG versus LG chickens at 7 wk. The HG chickens are fatter and over-express numerous genes that support higher rates of visceral adipogenesis and lipogenesis. In abdominal fat of LG chickens, we found higher expression of many genes involved in hemostasis, energy catabolism and endocrine signaling, which likely contribute to their leaner phenotype and slower growth. Many transcription factors and their direct target genes identified in HG and LG chickens could be involved in their divergence in adiposity and growth rate. The present analyses of the visceral fat transcriptome in chickens divergently selected for a large difference in growth rate and abdominal fatness clearly demonstrate that abdominal fat is a very dynamic metabolic and endocrine organ in the chicken. The HG chickens overexpress many transcription factors and their direct target genes, which should enhance in situ lipogenesis and ultimately adiposity. Our observation of enhanced expression of hemostasis and endocrine-signaling genes in diminished abdominal fat of LG cockerels provides insight into genetic mechanisms involved in divergence of abdominal fatness and somatic growth in avian and perhaps mammalian species, including humans.https://doi.org/10.1186/s12864-017-4035-

The abdominal fat contribution to adiposity in chickens divergently selected for fatness or growth: Cross-model elucidation and validation of gene expression

Cogburn, LarryThe domestic chicken (Gallus domesticus) is an important global source of high-quality dietary protein and a widely used biological model. Decades of intensive genetic selection have established the remarkable growth rate of the commercial broiler today; however, increased growth rate has been accompanied by the magnification of several unfavorable traits. This thesis addresses abdominal fatness, one of the traits that are incidentally amplified by selection for increased growth. Excessive fatness, coupled with several unique avian features of metabolism (i.e., fasting hyperglycemia and insulin insensitivity), parallels conditions observed in humans with metabolic diseases. Thus, understanding the genetic influence on excessive fatness in chickens will not only serve to improve the quality of production from an agricultural standpoint, but will also advance the knowledge of metabolic disorders in humans. Four experimental lines of meat-type chickens that were divergently selected for either a large difference in abdominal (visceral) fatness or in growth rate were used to characterize the role of adipose tissue (classically thought to have a minimal lipogenic contribution) in regulating adiposity. At the age of selection (9 weeks), the fat line (FL) and lean line (LL) chickens exhibit a 2.5-fold difference in abdominal adipose weight, while their body weight and feed intake are similar. The high growth (HG) and low growth (LG) chickens were divergently selected for either high (HG) or low (LG) body weight at 8 and 32 weeks of age resulting in a 2.7-fold increase in bodyweight and an 8-fold increase in abdominal fatness (as a percentage of BW) in HG chickens on average from 1 through 11 weeks. The adipose transcriptomes of these four genotypes (FL compared to LL and HG compared to LG) were analyzed at 1 through 11 weeks of age using the Del-Mar 14K Chicken Integrated Systems microarray, and at a single age (7 weeks) by RNA sequencing. Microarray analysis of abdominal fat in FL and LL chickens revealed 131 differentially expressed (DE) genes (FDR???0.05) as the main effect of genotype, 254 DE genes as an interaction of age and genotype and 3,194 DE genes (FDR???0.01) as the main effect of age. The most notable discoveries in the abdominal fat transcriptome during juvenile development were higher expression of many genes involved in hemostasis in the LL and up-regulation of numerous adipogenic and lipogenic genes in FL chickens. Many of these DE genes belong to pathways controlling the synthesis, metabolism and transport of lipids or endocrine signaling pathways activated by adipokines, retinoids and thyroid hormones. The importance of these processes in regulating adiposity in abdominal fat of FL and LL chickens was reinforced by the deep RNA sequencing analysis at 7 weeks. Remarkably, the highest expressed genes at this age included those involved in the metabolism of lipid and carbohydrates which are functionally associated with endocrine system and metabolic disorders. There were 1,687 DE genes between fat and lean chickens at 7 weeks including transcription factors and metabolic enzymes which have direct influences on lipogenesis and adipogenesis. The findings of the microarray analysis were further verified by the abundance of DE hemostatic factors uncovered by RNA sequencing analysis. This deep sequencing analysis also revealed a number of ectopically expressed genes suggesting that visceral fat functions autonomously as well as an endocrine organ in the regulation of lipid metabolism and perhaps feed intake. Microarray analysis of HG and LG chickens at 1 through 11 weeks of age revealed DE genes (FDR???0.05) as the main effect of genotype (321 genes), the interaction of age and genotype (718 genes), and the main effect of age (2,918 genes). RNA sequencing at 7 weeks uncovered 280 DE genes (FDR???0.1). Similar to the FL, HG chickens over-express many genes involved in adipogenesis and lipogenesis (including biosynthesis of fatty acids, cholesterol and triglycerides) which could at least partially account for their increase in abdominal fatness. Conversely, LG chickens up-regulate several energy producing processes (i.e., peroxisomal ???-oxidation, mitochondrial ???-oxidation, ketogenesis and oxidative phosphorylation) early on in juvenile development which are likely responsible for their extreme leanness. Hemostasis also appears to have a critical role in the maintenance of the lean phenotype at the age of maximal difference in adiposity in these chickens (7 weeks). These findings validate abdominal fat as a major contributor to adiposity in response to either divergent selection on abdominal fatness in the FL and LL or body weight in the HG and LG chickens.This work was supported by a grant from the United States Department of Agriculture, Initiative for Future Agricultural and Food Systems, Animal Genome Program (USDA-IFAFS; Award # 00-52100-9614) and a grant from the USDA Cooperative State Research, Education, and Extension Service (CSREES; Award # 2009-34562-20008) to the Avian Biosciences Center, University of Delaware.University of Delaware, Department of Animal ScienceM.S