Wilfrid Carre

Functional genomics in chickens: development of integrated-systems microarrays for transcriptional profiling and discovery of regulatory pathways.

The genetic networks that govern the differentiation and growth of major tissues of economic importance in the chicken are largely unknown. Under a functional genomics project, our consortium has generated 30 609 expressed sequence tags (ESTs) and developed several chicken DNA microarrays, which represent the Chicken Metabolic/Somatic (10 K) and Neuroendocrine/Reproductive (8 K) Systems (http://udgenome.ags.udel.edu/cogburn/). One of the major challenges facing functional genomics is the development of mathematical models to reconstruct functional gene networks and regulatory pathways from vast volumes of microarray data. In initial studies with liver-specific microarrays (3.1 K), we have examined gene expression profiles in liver during the peri-hatch transition and during a strong metabolic perturbation—fasting and re-feeding—in divergently selected broiler chickens (fast vs. slow-growth lines). The expression of many genes controlling metabolic pathways is dramatically altered by these perturbations. Our analysis has revealed a large number of clusters of functionally related genes (mainly metabolic enzymes and transcription factors) that control major metabolic pathways. Currently, we are conducting transcriptional profiling studies of multiple tissues during development of two sets of divergently selected broiler chickens (fast vs. slow growing and fat vs. lean lines). Transcriptional profiling across multiple tissues should permit construction of a detailed genetic blueprint that illustrates the developmental events and hierarchy of genes that govern growth and development of chickens. This review will briefly describe the recent acquisition of chicken genomic resources (ESTs and microarrays) and our consortiums efforts to help launch the new era of functional genomics in the chicken.

Mapping quantitative trait loci affecting fatness and breast muscle weight in meat-type chicken lines divergently selected on abdominal fatness

Quantitative trait loci (QTL) for abdominal fatness and breast muscle weight were investigated in a three-generation design performed by inter-crossing two experimental meat-type chicken lines that were divergently selected on abdominal fatness. A total of 585 F2 male offspring from 5 F1 sires and 38 F1 dams were recorded at 8 weeks of age for live body, abdominal fat and breast muscle weights. One hundred-twenty nine microsatellite markers, evenly located throughout the genome and heterozygous for most of the F1 sires, were used for genotyping the F2 birds. In each sire family, those offspring exhibiting the most extreme values for each trait were genotyped. Multipoint QTL analyses using maximum likelihood methods were performed for abdominal fat and breast muscle weights, which were corrected for the effects of 8-week body weight, dam and hatching group. Isolated markers were assessed by analyses of variance. Two significant QTL were identified on chromosomes 1 and 5 with effects of about one within-family residual standard deviation. One breast muscle QTL was identified on GGA1 with an effect of 2.0 within-family residual standard deviation.

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

BackgroundThis 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).ResultsMicroarray 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.ConclusionsThe 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.

Mapping QTL for growth and shank traits in chickens divergently selected for high or low body weight

An F(2) population (695 individuals) was established from broiler chickens divergently selected for either high (HG) or low (LG) growth, and used to localize QTL for developmental changes in body weight (BW), shank length (SL9) and shank diameter (SD9) at 9 weeks. QTL mapping revealed three genome-wide QTL on chromosomes (GGA) 2, 4 and 26 and three suggestive QTL on GGA 1, 3 and 5. Most of the BW QTL individually explained 2-5% of the phenotypic variance. The BW QTL on GGA2 explained about 7% of BW from 3 to 7 weeks of age, while that on GGA4 explained 15% of BW from 5 to 9 weeks. The BW QTL on GGA2 and GGA4 could be associated with early and late growth respectively. The GGA4 QTL also had the largest effect on SL9 and SD9 and explained 7% and 10% of their phenotypic variances respectively. However, when SL9 and SD9 were corrected with BW9, a shank length percent QTL was identified on GGA2. We identified novel QTL and also confirmed previously identified loci in other chicken populations. As the foundation population was established from commercial broiler strains, it is possible that QTL identified in this study could still be segregating in commercial strains.

Manipulation of thyroid status and/or GH injection alters hepatic gene expression in the juvenile chicken.

Both thyroid hormone (T3) and growth hormone (GH) are important regulators of somatic growth in birds and mammals. Although T3-mediated gene transcription is well known, the molecular basis of T3 interaction with GH on growth and development of birds remains unknown. In earlier studies, we discovered that exogenous GH alone increased accumulation of visceral fat in young chickens, while the combination of GH injections and dietary T3 worked synergistically to deplete body fat. In the present study, cDNA microarray and quantitative RT-PCR analyses enabled us to examine hepatic gene expression in young chickens after chronic manipulation of thyroid status and GH injection alone or in combination with T3. Thyroid status modulates expression of common and unique sets of genes involved in a wide range of molecular functions (i.e., energy metabolism, storage and transport, signal transduction, protein turnover and drug detoxification). Hepatic expression of 35 genes was altered by hypothyroidism (e.g., ADFP, ANGPTL3, GSTΑ, CAT, PPARG, HMGCL, GHR, IGF1, STAT3, THRSPα), whereas hyperthyroidism affected expression of another cluster of 13 genes (e.g., IGFBP1, KHK, LDHB, BAIA2L1, SULT1B, TRIAD3). Several genes were identified which have not been previously ascribed as T3 responsive (e.g., DEFB9, EPS8L2, ARHGAP1, LASS2, INHBC). Exogenous GH altered expression of 17 genes (e.g., CCAR1, CYP2C45, GYS2, ENOB, HK1, FABP1, SQLE, SOCS2, UPG2). The T3+GH treatment depleted the greatest amount of body fat, where 34 differentially expressed genes were unique to this group (e.g., C/EBP, CDC42EP1, SYDE2, PCK2, PIK4CA, TH1L, GPT2, BHMT). The marked reduction in body fat brought about by the T3+GH synergism could involve modulation of hormone signaling via altered activity of the Ras superfamily of molecular switches, which control diverse biological processes. In conclusion, this study provides the first global analysis of endocrine (T3 and GH) regulation of hepatic gene transcription in the chicken.

Transcriptional profiling of liver in riboflavin-deficient chicken embryos explains impaired lipid utilization, energy depletion, massive hemorrhaging, and delayed feathering

BackgroundA strain of Leghorn chickens (rd/rd), unable to produce a functional riboflavin-binding protein, lays riboflavin-deficient eggs, in which all embryos suddenly die at mid-incubation (days 13-15). This malady, caused by riboflavin deficiency, leads to excessive lipid accumulation in liver, impaired β-oxidation of lipid, and severe hypoglycemia prior to death. We have used high-density chicken microarrays for time-course transcriptional scans of liver in chicken embryos between days 9-15 during this riboflavin-deficiency-induced metabolic catastrophe. For comparison, half of rd/rd embryos (n = 16) were rescued from this calamity by injection of riboflavin just prior to incubation of fertile eggs from rd/rd hens.ResultsNo significant differences were found between hepatic transcriptomes of riboflavin-deficient and riboflavin-rescued embryos at the first two ages (days 9 and 11). Overall, we found a 3.2-fold increase in the number of differentially expressed hepatic genes between day 13 (231 genes) and day 15 (734 genes). Higher expression of genes encoding the chicken flavoproteome was more evident in rescued- (15 genes) than in deficient-embryos (4 genes) at day 15. Diminished activity of flavin-dependent enzymes in riboflavin-deficient embryos blocks catabolism of yolk lipids, which normally serves as the predominant source of energy required for embryonic development.ConclusionsRiboflavin deficiency in mid-stage embryos leads to reduced expression of numerous genes controlling critical functions, including β-oxidation of lipids, blood coagulation and feathering. Surprisingly, reduced expression of feather keratin 1 was found in liver of riboflavin-deficient embryos at e15, which could be related to their delayed feathering and sparse clubbed down. A large number of genes are expressed at higher levels in liver of riboflavin-deficient embryos; these up-regulated genes control lipid storage/transport, gluconeogenesis, ketogenesis, protein catabolism/ubiquitination and cell death.