RESUMO
In mammals, leptin and tumor-necrosis factor (TNF) are prominent interacting adipokines mediating appetite control and insulin sensitivity. While TNF pleiotropically functions in immune defense and cell survival, leptin is largely confined to signaling energy stores in adipocytes. Knowledge about the function of avian leptin and TNF is limited and they are absent or lowly expressed in adipose, respectively. Employing radiation-hybrid mapping and FISH-TSA, we mapped TNF and its syntenic genes to chicken chromosome 16 within the major histocompatibility complex (MHC) region. This mapping position suggests that avian TNF has a role in regulating immune response. To test its possible interaction with leptin within the immune system and beyond, we compared the transcription patterns of TNF, leptin and their cognate receptors obtained by meta-analysis of GenBank RNA-seq data. While expression of leptin and its receptor (LEPR) were detected in the brain and digestive tract, TNF and its receptor mRNAs were primarily found in viral-infected and LPS-treated leukocytes. We confirmed leptin expression in the duodenum by immunohistochemistry staining. Altogether, we suggest that whereas leptin and TNF interact as adipokines in mammals, in birds, they have distinct roles. Thus, the interaction between leptin and TNF may be unique to mammals.
Assuntos
Galinhas/genética , Mapeamento Cromossômico , Digestão , Regulação da Expressão Gênica , Leptina/genética , Mamíferos/genética , Transdução de Sinais , Fator de Necrose Tumoral alfa/metabolismo , Animais , Linhagem Celular , Galinhas/metabolismo , Duodeno/metabolismo , Feminino , Leptina/metabolismo , Metáfase/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Mapeamento de Híbridos Radioativos , Receptores para Leptina/metabolismo , Receptores do Fator de Necrose Tumoral/genética , Receptores do Fator de Necrose Tumoral/metabolismo , Sintenia/genética , Fator de Necrose Tumoral alfa/genéticaRESUMO
RNA editing results in a post-transcriptional nucleotide change in the RNA sequence that creates an alternative nucleotide not present in the DNA sequence. This leads to a diversification of transcription products with potential functional consequences. Two nucleotide substitutions are mainly described in animals, from adenosine to inosine (A-to-I) and from cytidine to uridine (C-to-U). This phenomenon is described in more details in mammals, notably since the availability of next generation sequencing technologies allowing whole genome screening of RNA-DNA differences. The number of studies recording RNA editing in other vertebrates like chicken is still limited. We chose to use high throughput sequencing technologies to search for RNA editing in chicken, and to extend the knowledge of its conservation among vertebrates. We performed sequencing of RNA and DNA from 8 embryos. Being aware of common pitfalls inherent to sequence analyses that lead to false positive discovery, we stringently filtered our datasets and found fewer than 40 reliable candidates. Conservation of particular sites of RNA editing was attested by the presence of 3 edited sites previously detected in mammals. We then characterized editing levels for selected candidates in several tissues and at different time points, from 4.5 days of embryonic development to adults, and observed a clear tissue-specificity and a gradual increase of editing level with time. By characterizing the RNA editing landscape in chicken, our results highlight the extent of evolutionary conservation of this phenomenon within vertebrates, attest to its tissue and stage specificity and provide support of the absence of non A-to-I events from the chicken transcriptome.
Assuntos
Galinhas/genética , Genoma , Edição de RNA , Animais , Embrião de Galinha , Biologia Computacional , DNA/química , Evolução Molecular , Sequenciamento de Nucleotídeos em Larga Escala , RNA/química , Análise de Sequência de DNA , Análise de Sequência de RNARESUMO
Very few causal genes have been identified by quantitative trait loci (QTL) mapping because of the large size of QTL, and most of them were identified thanks to functional links already known with the targeted phenotype. Here, we propose to combine selection signature detection, coding SNP annotation, and cis-expression QTL analyses to identify potential causal genes underlying QTL identified in divergent line designs. As a model, we chose experimental chicken lines divergently selected for only one trait, the abdominal fat weight, in which several QTL were previously mapped. Using new haplotype-based statistics exploiting the very high SNP density generated through whole-genome resequencing, we found 129 significant selective sweeps. Most of the QTL colocalized with at least one sweep, which markedly narrowed candidate region size. Some of those sweeps contained only one gene, therefore making them strong positional causal candidates with no presupposed function. We then focused on two of these QTL/sweeps. The absence of nonsynonymous SNPs in their coding regions strongly suggests the existence of causal mutations acting in cis on their expression, confirmed by cis-eQTL identification using either allele-specific expression or genetic mapping analyses. Additional expression analyses of those two genes in the chicken and mice contrasted for adiposity reinforces their link with this phenotype. This study shows for the first time the interest of combining selective sweeps mapping, coding SNP annotation and cis-eQTL analyses for identifying causative genes for a complex trait, in the context of divergent lines selected for this specific trait. Moreover, it highlights two genes, JAG2 and PARK2, as new potential negative and positive key regulators of adiposity in chicken and mice.
Assuntos
Adiposidade/genética , Proteínas de Membrana/genética , Locos de Características Quantitativas , Ubiquitina-Proteína Ligases/genética , Tecido Adiposo Branco/metabolismo , Alelos , Animais , Linhagem Celular , Galinhas , Mapeamento Cromossômico , Genoma , Sequenciamento de Nucleotídeos em Larga Escala , Proteína Jagged-2 , Proteínas de Membrana/metabolismo , Camundongos , Anotação de Sequência Molecular , Miosinas/genética , Miosinas/metabolismo , Polimorfismo de Nucleotídeo Único , Análise de Sequência de DNARESUMO
Classical quantitative trait loci (QTL) analysis and gene expression QTL (eQTL) were combined to identify the causal gene (or QTG) underlying a highly significant QTL controlling the variation of breast meat color in a F2 cross between divergent high-growth (HG) and low-growth (LG) chicken lines. Within this meat quality QTL, BCMO1 (Accession number GenBank: AJ271386), encoding the ß-carotene 15, 15'-monooxygenase, a key enzyme in the conversion of ß-carotene into colorless retinal, was a good functional candidate. Analysis of the abundance of BCMO1 mRNA in breast muscle of the HG x LG F2 population allowed for the identification of a strong cis eQTL. Moreover, reevaluation of the color QTL taking BCMO1 mRNA levels as a covariate indicated that BCMO1 mRNA levels entirely explained the variations in meat color. Two fully-linked single nucleotide polymorphisms (SNP) located within the proximal promoter of BCMO1 gene were identified. Haplotype substitution resulted in a marked difference in BCMO1 promoter activity in vitro. The association study in the F2 population revealed a three-fold difference in BCMO1 expression leading to a difference of 1 standard deviation in yellow color between the homozygous birds at this haplotype. This difference in meat yellow color was fully consistent with the difference in carotenoid content (i.e. lutein and zeaxanthin) evidenced between the two alternative haplotypes. A significant association between the haplotype, the level of BCMO1 expression and the yellow color of the meat was also recovered in an unrelated commercial broiler population. The mutation could be of economic importance for poultry production by making possible a gene-assisted selection for color, a determining aspect of meat quality. Moreover, this natural genetic diversity constitutes a new model for the study of ß-carotene metabolism which may act upon diverse biological processes as precursor of the vitamin A.
Assuntos
Galinhas/genética , Carne , Locos de Características Quantitativas/genética , beta-Caroteno 15,15'-Mono-Oxigenase/genética , Animais , Cruzamento , Carotenoides/metabolismo , Linhagem Celular Tumoral , Mapeamento Cromossômico , Feminino , Regulação Enzimológica da Expressão Gênica , Genótipo , Haplótipos , Humanos , Luciferases/genética , Luciferases/metabolismo , Masculino , Músculos/metabolismo , Mutação , Pigmentação/genética , Polimorfismo de Nucleotídeo Único , Regiões Promotoras Genéticas/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Análise de Sequência de DNARESUMO
We present a gene-based RH map of the chicken microchromosome GGA14, known to have synteny conservations with human chromosomal regions HSA16p13.3 and HSA17p11.2. Microsatellite markers from the genetic map were used to check the validity of the RH map and additional markers were developed from chicken EST data to yield comparative mapping data. A high rate of intra-chromosomal rearrangements was detected by comparison to the assembled human sequence. Finally, the alignment of the RH map to the assembled chicken sequence showed a small number of discordances, most of which involved the same region of the chromosome spanning between 40.5 and 75.9 cR(6000) on the RH map.
Assuntos
Galinhas/genética , Cromossomos/genética , Mapeamento de Híbridos Radioativos , Animais , Sequência de Bases , Biologia Computacional , Primers do DNA , Etiquetas de Sequências Expressas , Ordem dos Genes , Marcadores Genéticos/genética , Humanos , Repetições de Microssatélites/genética , Dados de Sequência Molecular , Alinhamento de Sequência , Análise de Sequência de DNARESUMO
BACKGROUND: The ChickRH6 whole chicken genome radiation hybrid (RH) panel recently produced has already been used to build radiation hybrid maps for several chromosomes, generating comparative maps with the human and mouse genomes and suggesting improvements to the chicken draft sequence assembly. Here we present the construction of a RH map of chicken chromosome 2. Markers from the genetic map were used for alignment to the existing GGA2 (Gallus gallus chromosome 2) linkage group and EST were used to provide valuable comparative mapping information. Finally, all markers from the RH map were localised on the chicken draft sequence assembly to check for eventual discordances. RESULTS: Eighty eight microsatellite markers, 10 genes and 219 EST were selected from the genetic map or on the basis of available comparative mapping information. Out of these 317 markers, 270 gave reliable amplifications on the radiation hybrid panel and 198 were effectively assigned to GGA2. The final RH map is 2794 cR6000 long and is composed of 86 framework markers distributed in 5 groups. Conservation of synteny was found between GGA2 and eight human chromosomes, with segments of conserved gene order of varying lengths. CONCLUSION: We obtained a radiation hybrid map of chicken chromosome 2. Comparison to the human genome indicated that most of the 8 groups of conserved synteny studied underwent internal rearrangements. The alignment of our RH map to the first draft of the chicken genome sequence assembly revealed a good agreement between both sets of data, indicative of a low error rate.
Assuntos
Mapeamento de Híbridos Radioativos/métodos , Animais , Galinhas , Mapeamento Cromossômico , Biologia Computacional/métodos , Sequência Conservada , Primers do DNA/química , Etiquetas de Sequências Expressas , Ligação Genética , Marcadores Genéticos , Técnicas Genéticas , Genoma , Humanos , Repetições de Microssatélites , Modelos Genéticos , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Reprodutibilidade dos Testes , Especificidade da EspécieRESUMO
BACKGROUND: The resolution of radiation hybrid (RH) maps is intermediate between that of the genetic and BAC (Bacterial Artificial Chromosome) contig maps. Moreover, once framework RH maps of a genome have been constructed, a quick location of markers by simple PCR on the RH panel is possible. The chicken ChickRH6 panel recently produced was used here to construct a high resolution RH map of chicken GGA5. To confirm the validity of the map and to provide valuable comparative mapping information, both markers from the genetic map and a high number of ESTs (Expressed Sequence Tags) were used. Finally, this RH map was used for testing the accuracy of the chicken genome assembly for chromosome 5. RESULTS: A total of 169 markers (21 microsatellites and 148 ESTs) were typed on the ChickRH6 RH panel, of which 134 were assigned to GGA5. The final map is composed of 73 framework markers extending over a 1315.6 cR distance. The remaining 61 markers were placed alongside the framework markers within confidence intervals. CONCLUSION: The high resolution framework map obtained in this study has markers covering the entire chicken chromosome 5 and reveals the existence of a high number of rearrangements when compared to the human genome. Only two discrepancies were observed in relation to the sequence assembly recently reported for this chromosome.
Assuntos
Galinhas/genética , Cromossomos Humanos/genética , Cromossomos/genética , Mapeamento de Híbridos Radioativos/métodos , Animais , DNA/genética , Etiquetas de Sequências Expressas , Marcadores Genéticos/genética , Humanos , Camundongos , Alinhamento de Sequência/métodosRESUMO
To validate the ChickRH6 whole-genome radiation hybrid (WGRH) panel, we constructed a map of chicken Chromosome 7 based on 19 microsatellite markers from the genetic map and 76 ESTs (expressed sequence tags), whose efficient targeted development was made possible by using the ICCARE software. This high-density radiation hybrid (RH) map of a chicken macrochromosome gives us indications on characteristics of ChickRH6. The potential resolution of the panel is 325 kb and the practical resolution of our framework map is 1.3 Mb. Based on these results, a complete framework map of the chicken genome would comprise 1000 markers. The marker order is in good agreement with the genetic map and comparison with the human and mouse sequence maps revealed a number of internal rearrangements.