Birds and mammals shared a common ancestor 
310 million years ago (Mya) (Hedges 2002
). Sequence comparisons between these groups are characterized with a high signal-to-noise ratio for the detection of functional elements. Taken together with the ready access to chicken embryos and as a major food source, chicken genomics is likely to have major applications and benefits in comparative genomics, evolutionary biology and systematics, models of development and human disease, and agriculture.
Comparative genomics
A major reason for sequencing the chicken genome was to increase our understanding of the human genome through comparative genomics, for example, to define regions under selection such as coding and regulatory elements (Hillier et al. 2004). Comparisons with known functional sequences suggested that 75% of coding regions and 30%–40% of regulatory elements are conserved. Only 2.5% of the chicken sequence could be aligned with that of the human (44% coding, 25% intronic, and 31% intergenic) and, given that 5% of the mammalian genome is under selection, almost all of this is likely to be of functional significance.
Comparative genomics has identified ~400 ultra-conserved regions (UCR) greater than 200 bp sharing at least 95% sequence identity between human and chicken (Sandelin et al. 2004). Surprisingly, highly conserved, noncoding regions like the UCR often exist far from any predicted gene within so-called "gene deserts" that are apparently free of any known protein-coding genes and are often clustered (Ovcharenko et al. 2005). Genes with a role in transcriptional regulation and development flank many of these UCR and gene deserts. These regions are often far from genes and may represent distant regulatory signals.
Parent-specific gene expression by genomic imprinting is only found in mammals and not birds or lower vertebrates. Therefore, comparison of imprinted genes in mammals with orthologs in the chicken may uncover features about the origins of imprinting. Comparative mapping suggests these genes cluster on macrochromosomes in regions that preferentially undergo asynchronous DNA replication (Dunzinger et al. 2005). Analysis of the chicken region orthologous to the imprinted mammalian ASCL2–H19 region (Yokomine et al. 2005) revealed extensive conservation of gene organization, except H19, a critical noncoding imprinted gene. This gene and its regulatory elements were absent from the chicken genome. These studies suggest that imprinted genes were clustered before the evolution of imprinting, an event that occurred after the divergence of birds and mammals ~310 Mya. Subsequently, imprinting control elements, such as the H19 gene region, must have evolved by duplication and/or transposition into these gene clusters.
A long-standing question in genome evolution has been the question of genome size. The chicken genome is 35% the size of the human and 45% of mouse. In part, this can be explained in terms of the low frequency of repeats, pseudogenes, segmental duplication, and gene duplications (Hillier et al. 2004). However, these factors only account for 20%–25% of the variation in genome size, so other factors are at work, possibly a dearth of ancient repeats (that are no longer detectably repetitive) or reduction in cell size and energy conservation (Hughes and Piontkivska 2005).
Developmental biology
Applications in developmental biology are likely to be another major beneficiary of the genome sequence (Burt 2004b; Stern 2005). The chicken has always been a favorite among developmental biologists (Brown et al. 2003; Stern 2005) because of easy access to the chick embryo and ease of manipulation. These features, when combined with the new tools of genomics, are ideal for testing gene function and predicted regulatory sequences in vivo. For example, studies on the conservation of the avian SOX2 genes have identified neural specific enhancers, confirmed in vivo by electroporation of chick embryo neural tubes (Uchikawa et al. 2004).
In the mouse and other model systems, whole-mount in situ hybridization screens have been useful in identifying patterns of expression that may suggest developmental functions of novel genes (EMAP). A similar effort has started in the chicken using the large collection of sequenced chicken ESTs (Boardman et al. 2002; ARK-Genomics; ChickEST). Data can be accessed at GEISHA and standard three-dimensional embryo reconstructions are under development (EMAP).
Genetic variation and complex trait analysis
In parallel with the chicken genome sequencing project, a consortium (Wong et al. 2004; Wang et al. 2005b; ChickVD) generated 2.8 million SNPs from a comparison of the Red Jungle Fowl reference sequence and partial genome scans of Silkie, Broiler, and Layer lines. Nucleotide diversity (5 x 10-3 per nucleotide) was six times the rate found in humans (Ellegren 2005). Resequencing confirmed 94% of the total and 83% of the nonsynonymous SNPs. An initial surprise was that ~70% of SNPs were common to all breeds, suggesting an origin prior to domestication 5,000–10,000 years ago. Another possibility is that their ancestry has been lost because of extensive cross breeding between Asian and western poultry populations. The next steps are to verify a larger sample of SNPs and create high-resolution genetic and linkage disequilibrium maps of chicken populations. These assays will be used to map and identify genes controlling traits of economic and biological interest at quantitative trait loci (QTL). Currently, more than 600 QTL have been mapped using microsatellites (Andersson and Georges 2004; Hocking 2005; Wang et al. 2005b). The availability of a standard set of 10,000 or more SNPs combined with the ease of building structured large resource populations hold much promise towards the identification of genes controlling these traits.
Animal health and the avian immune system
One area that has benefited most from genomic approaches has been the characterization of the genes and proteins in the avian immune system. The MHC was the first major chicken genome sequence to be assembled (Kaufman et al. 1999) and was a surprise, being relatively compact and simpler than those of mammals. Since then, there has been slow progress in the isolation of avian cytokines and other signaling molecules. The main problem has been their high rate of evolution, limiting their detection using homology to mammalian sequences (Staeheli et al. 2001). Even now, one must be careful in concluding that avian homologs to mammalian immune genes do not exist, as several examples known from ESTs or directed sequencing were not found in the genome assembly. This started to change when analysis of large EST data sets identified 185 immune-related sequences (Lynn et al. 2003; Smith et al. 2004). This compared with the 80 genes identified by Tirunagaru et al. (2000) and the 28 genes listed in the review by Staeheli et al. (2001). Sequences included interleukins, transcription factors, chemokines, differentiation antigens, receptors, genes involved in the Toll pathway, and MHC-associated genes. The discovery of IL4 and other cytokines involved in the Th2 response (Smith et al. 2004) was a surprise, since it had previously been speculated that the chicken does not elicit a typical Th2 response (Staeheli et al. 2001). The receptors for IL10 and IL13 were also identified, indicating that the chicken probably also contained these genes, which are typical Tr1 and Th2 cytokines. This was confirmed by sequencing specific BAC clones identified assuming conservation of synteny between chicken and mammalian genomes (Avery et al. 2004; Rothwell et al. 2004).
A comprehensive analysis of the chicken genome sequence has identified many cytokines, chemokines, and their receptors (Hillier et al. 2004; Kaiser et al. 2004, 2005; Wang et al. 2005a). Even genes once thought to be mammalian-specific, including IL3, IL7, IL9, IL26, CSMF, LIF, and Cathelicidin, were found (Hillier et al. 2004). These are proteins that evolve rapidly and require more effort to detect. A number of orthologs to human chemokines are absent from the chicken genome, including CCL2, 7, 8, 11, 15, 18, 23, 24, and 26; CXCL1–7, 9, 10, and 11, possibly products of independent gene duplications in mammals. Similarly, missing chemokine receptors included CCR1, CCR3, CCR10, CXCR3, and CXCR6. The lack of functional eosinophils correlates with the absence of the eotaxin genes (CCL22, CCL24, CCL26) and their receptor (CCR3). Chickens lack lymph nodes and also the genes for the lymphotoxins (LT-α and -β) and their receptors. TNF is also absent, but its receptor, TNFRSF1A (ENSGALG00000014890) is present, suggesting that further sequencing will reveal this gene in the chicken. Similar analyses have been performed on the leukocyte receptor complex (Nikolaidis et al. 2005) that regulates the activity of T- and B-lymphocytes and NK cells. A model of evolution by repeated birth and death of these Ig-like receptors' genes was proposed.
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