β-Keratins in crocodiles reveal amino acid homology with avian keratins

The DNA sequences encoding β-keratin have been obtained from Marsh Mugger (Crocodylus palustris) and Orinoco Crocodiles (Crocodylus intermedius). Through the deduced amino acid sequence, these proteins are rich in glycine, proline and serine. The central region of the proteins are composed of two beta-folded regions and show a high degree of identity with β-keratins of aves and squamates. This central part is thought to be the site of polymerization to build the framework of β-keratin filaments. It is believed that the β-keratins in reptiles and birds share a common ancestry. Near the C-terminal, these β-keratins contain a peptide rich in glycine-X and glycine-X-X, and the distinctive feature of the region is some 12-amino acid repeats, which are similar to the 13-amino acid repeats in chick scale keratin but absent from avian feather keratin. From our phylogenetic analysis, the β-keratins in crocodile have a closer relationship with avian keratins than the other keratins in reptiles.     

Linking the molecular evolution of avian beta (β) keratins to the evolution of feathers

Feathers of today's birds are constructed of beta (β)-keratins, structural proteins of the epidermis that are found solely in reptiles and birds. Discoveries of "feathered dinosaurs" continue to stimulate interest in the evolutionary origin of feathers, but few studies have attempted to link the molecular evolution of their major structural proteins (β-keratins) to the appearance of feathers in the fossil record. Using molecular dating methods, we show that before the appearance of Anchiornis (～155 Million years ago (Ma)) the basal β-keratins of birds began diverging from their archosaurian ancestor ～216?Ma. However, the subfamily of feather β-keratins, as found in living birds, did not begin diverging until ～143?Ma. Thus, the pennaceous feathers on Anchiornis, while being constructed of avian β-keratins, most likely did not contain the feather β-keratins found in the feathers of modern birds. Our results demonstrate that the evolutionary origin of feathers does not coincide with the molecular evolution of the feather β-keratins found in modern birds. More likely, during the Late Jurassic, the epidermal structures that appeared on organisms in the lineage leading to birds, including early forms of feathers, were constructed of avian β-keratins other than those found in the feathers of modern birds. Recent biophysical studies of the β-keratins in feathers support the view that the appearance of the subfamily of feather β-keratins altered the biophysical nature of the feather establishing its role in powered flight.     

Genomic organization and molecular phylogenies of the beta (β) keratin multigene family in the chicken (<Emphasis Type="Italic">Gallus gallus</Emphasis>) and zebra finch (<Emphasis Type="Italic">Taeniopygia guttata</Emphasis>): implications for feather evolution

Background  

The epidermal appendages of reptiles and birds are constructed of beta (β) keratins. The molecular phylogeny of these keratins is important to understanding the evolutionary origin of these appendages, especially feathers. Knowing that the crocodilian β-keratin genes are closely related to those of birds, the published genomes of the chicken and zebra finch provide an opportunity not only to compare the genomic organization of their β-keratins, but to study their molecular evolution in archosaurians.     

The Expression of Beta ({beta}) Keratins in the Epidermal Appendages of Reptiles and Birds

The integuments of extant vertebrates display a variety of epidermalappendages whose patterns, morphology and terminal differentiation(epidermal keratins) depend upon interactions between ectodermal(epidermis) and mesodermal (dermis) tissues. In reptiles andbirds, appendage morphogenesis precedes terminal differentiation.Studies have demonstrated that appendage morphogenesis influencesthe expression of the appendage specific keratin genes. However,little is known about the nature of the structural genes expressedby the epidermal appendages of reptiles. How pattern formationand/or appendage morphogenesis influence terminal differentiationof reptilian appendages is not known. The epidermal appendages of reptiles and birds are characterizedby the presence of both alpha () and beta (ß) typekeratin proteins. Studies have focused on the genes of avianß keratins because they are the major structural proteinsof feathers. The occurrence of ß keratin proteinsin the scales and claws of both birds and reptiles and theirimmunological cross-reactivity suggest that the genes for reptilianß keratins may be homologous with those of birds.In bird appendages, the ß keratins are the productsof a large family of homologous genes. Specific members of thisgene family are expressed during the development of each appendage.Recent sequence analyses of feather ß keratins, fromdifferent orders of birds, demonstrate that there is more diversityat the DNA level than was implied by earlier protein sequencingstudies. Immunological techniques show that the same antibodies thatreact with the epidermal ß keratins of the chicken(Gallus domesticus) react with the epidermal ß keratinsof American alligators (Alligator mississippiensis). Furthermore,a peptide sequence (20 amino acids) from an alligator claw ßkeratin is similar to a highly conserved region of avian claw,scale, feather, and feather-like ß keratins. Theseobservations suggest that the ß keratin genes of avianepidermal appendages have homologues in the American alligator.Understanding the origin and evolution of the ß keratingene families in reptiles and birds will undoubtedly add toour understanding of the evolution of skin appendages such asscales and feathers.     

Greater prairie chickens have a compact MHC-B with a single class IA locus

The major histocompatibility complex (MHC) plays a central role in innate and adaptive immunity, but relatively little is known about the evolution of the number and arrangement of MHC genes in birds. Insights into the evolution of the MHC in birds can be gained by comparing the genetic architecture of the MHC between closely related species. We used a fosmid DNA library to sequence a 60.9-kb region of the MHC of the greater prairie chicken (Tympanuchus cupido), one of five species of Galliformes with a physically mapped MHC. Greater prairie chickens have the smallest core MHC yet observed in any bird species, and major changes are observed in the number and arrangement of MHC loci. In particular, the greater prairie chicken differs from other Galliformes in the deletion of an important class I antigen binding gene. Analysis of the remaining class IA gene in a population of greater prairie chickens in Wisconsin, USA revealed little evidence for selection at the region responsible for antigen binding.     

On the origin of feathers

The appearance of feathers defines the appearance of birds. A number of changes defined, preceded or accompanied the event. The changes were hierarchical in nature and included revolutions in genomic organization (i.e., HOX and the feather keratin genes), protein sequence and shape, the large scale organization of proteins into filaments, and in the geometry of the cells and their roles in the follicle. Changes at each of these levels differ or produced different products than found in its analog in reptiles. They are essentially unique to birds and produced an evolutionary novelty. I used analysis of extant structure and information on development to reconstruct key events in the evolution of feathers. The ancestral reptilian epidermal structure, while probably a scale or tubercles, is still unidentified. The structural genes of feather proteins (φ-keratin) are tandem repeats probably assembled from pre-existing exons. They are unlike the alpha-keratin of vertebrate soft epidermis. Amino-acid composition, shape, and behavior of feather keratins are unique among vertebrates. The 3-dimensional organization of the follicle and the developmental processes are also unique. Although we lack a complete understanding of the appearance and early role of feathers, they are clearly the results of novel events.     

Comparative embryonic development of the skeleton of the domestic turkey (Meleagris gallopavo) and other galliform birds

Ossification sequences are poorly known for birds in general, even for common domestic and experimental species. Such sequences constitute a rich source of data on character evolution, and may even provide phylogenetic information. It is not clear, however, what factors influence ossification sequences and what the relative importance of phylogeny is to the sequences. Galliformes constitute a good group to examine these variables. These birds are osteologically conservative, have precocial young, but have a broad spectrum of body sizes and incubation periods. Here, I describe the embryonic ossification of the skeleton in the domestic turkey (Meleagris gallopavo), and compare it to the domestic chicken (Gallus gallus) and the Japanese quail (Coturnix coturnix). Ossification sequences in this group are not affected by egg size or incubation period. They also appear to be independent of both the spatial location and the embryonic tissue from which the osteogenic cells originated. Accumulation of a wider sample of ossification sequences from more morphologically variable avian taxa will be necessary in order to test functional and phylogenetic effects.     

黑水鸡与家鸡染色体带型同源性比较研究

从二倍体细胞大染色体组(Macrochromosomes)的G带特征比较分析,鹤形目的黑水鸡与鸡形目的家鸡带型相似程度很大,标志着这两种鸟类的亲缘关系较近,同时也充分说明了不同目鸟类在进化上染色体变异的保守性。其带型差异仅在于两次臂间倒位、两次臂内倒位和一次着丝点融合。此外,我们采用QM荧光——银染的相继染色法发现黑水鸡的W染色体上有NOR存在,这种核仁组织者区域与性染色质发生连锁的现象在鸟类是首次发现。     

Colour-producing β-keratin nanofibres in blue penguin (Eudyptula minor) feathers

The colours of living organisms are produced by the differential absorption of light by pigments (e.g. carotenoids, melanins) and/or by the physical interactions of light with biological nanostructures, referred to as structural colours. Only two fundamental morphologies of non-iridescent nanostructures are known in feathers, and recent work has proposed that they self-assemble by intracellular phase separation processes. Here, we report a new biophotonic nanostructure in the non-iridescent blue feather barbs of blue penguins (Eudyptula minor) composed of parallel β-keratin nanofibres organized into densely packed bundles. Synchrotron small angle X-ray scattering and two-dimensional Fourier analysis of electron micrographs of the barb nanostructure revealed short-range order in the organization of fibres at the appropriate size scale needed to produce the observed colour by coherent scattering. These two-dimensional quasi-ordered penguin nanostructures are convergent with similar arrays of parallel collagen fibres in avian and mammalian skin, but constitute a novel morphology for feathers. The identification of a new class of β-keratin nanostructures adds significantly to the known mechanisms of colour production in birds and suggests additional complexity in their self-assembly.     

Comparative evidence for costs of secondary sexual characters: adaptive vane emargination of ornamented feathers in birds

We used a comparative approach, by comparing bird species with tail ornamentation with sister taxa without ornamentation, to deduce the aerodynamic function of extravagant feather ornaments and the costs of such ornaments in birds. First, the aerodynamic function of tail feather ornaments in birds can be deduced from asymmetry in the width of tail feather vanes, since flightless birds have symmetrical vanes while flying birds without feather exaggeration by sexual selection have asymmetrical vanes. Distal inner vanes at the tip of tail feathers were more narrow in ornamented as compared to nonornamented birds, and vane asymmetry at the tip of the feather was therefore reduced in ornamented species, suggesting marginal aerodynamic function of the distal part of extravagant feather ornaments. Second, the cost of feather ornaments due to parasite drag is proportional to the area of feathers extending beyond the maximum continuous width of the tail, and aerodynamic costs of long tails could therefore be diminished by a reduction in feather width. Consistent with this prediction, the outermost tip of feather ornaments was narrower than the homologous character in nonornamented sister taxa, while the base of the feather had similar width in the two groups of birds. These results suggest that the costs of extravagant ornamentation have been diminished by a reduction in feather width, leading to a reduction in drag. Costs of feather ornaments, as demonstrated by their fine morphology, thus appear to have been extensive during the evolution of these characters.     

Comparative genomics analyses of alpha-keratins reveal insights into evolutionary adaptation of marine mammals

Background

Diversity of hair in marine mammals was suggested as an evolutionary innovation to adapt aquatic environment, yet its genetic basis remained poorly explored. We scanned α-keratin genes, one major structural components of hair, in 16 genomes of mammalian species, including seven cetaceans, two pinnipeds, polar bear, manatee and five terrestrial species.

Results

Extensive gene loss and high pseudogenization rate of α-keratin genes were identified in cetaceans when compared to terrestrial artiodactylans (average number of α-keratins 37.29 vs. 58.33; pseudogenization rate 29.89% vs. 8.00%), especially of hair follicle-specific keratin genes (average pseudogenization rate in cetaceans of 43.88% relative to 3.80% artiodactylian average). Compared to toothed whale, the much more number of intact functional α-keratin genes was examined in the baleen whale that had specific keratinized baleen. In contrast, the number of keratin genes in pinnipeds, polar bear and manatee were comparable to those of their respective terrestrial relatives. Additionally, four keratin genes (K39, K9, K42, and K74) were found to be pseudogenes or lost uniquely in cetaceans and manatees.

Conclusions

Species-specific evolution of α-keratin gene family identified in the marine mammals might be responsible for their different hair characteristics. Increased gene loss and pseudogenization rate identified in cetacean lineages was likely to contribute to hair-less phenotype to adaptation for complete aquatic environment. However, the fully aquatic manatee still remained the comparable number of intact genes to its terrestrial relative, probably due to its perioral bristles and bristle-like hairs on the oral disk. By contrast, similar evolution pattern of α-keratin gene repertoire in the pinnipeds, polar bear and their terrestrial relatives was likely due to abundant hair to keep warm when they went ashore. Interestingly, some keratin genes were exclusively lost in cetaceans and manatees, likely as a result of convergent hair-loss phenotype to inhabit completely aquatic environment in both groups.

     

Complete nucleotide sequence of the Coturnix chinensis (blue-breasted quail) mitochondrial genome and a phylogenetic analysis with related species

Coturnix chinensis (blue-breasted quail) has been classically grouped in Galliformes Phasianidae Coturnix, based on morphologic features and biochemical evidence. Since the blue-breasted quail has the smallest body size among the species of Galliformes, in addition to a short generation time and an excellent reproductive performance, it is a possible model fowl for breeding and physiological studies of the Coturnix japonica (Japanese quail) and Gallus gallus domesticus (chicken), which are classified in the same family as blue-breasted quail. However, since its phylogenetic position in the family Phasianidae has not been determined conclusively, the sequence of the entire blue-breasted quail mitochondria (mt) genome was obtained to provide genetic information for phylogenetic analysis in the present study. The blue-breasted quail mtDNA was found to be a circular DNA of 16,687 base pairs (bp) with the same genomic structure as the mtDNAs of Japanese quail and chicken, though it is smaller than Japanese quail and chicken mtDNAs by 10 bp and 88 bp, respectively. The sequence identity of all mitochondrial genes, including those for 12S and 16S ribosomal RNAs, between blue-breasted quail and Japanese quail ranged from 84.5% to 93.5%; between blue-breasted quail and chicken, sequence identity ranged from 78.0% to 89.6%. In order to obtain information on the phylogenetic position of blue-breasted quail in Galliformes Phasianidae, the 2,184 bp sequence comprising NADH dehydrogenase subunit 2 and cytochrome b genes available for eight species in Galliformes [Japanese quail, chicken, Gallus varius (green junglefowl), Bambusicola thoracica (Chinese bamboo partridge), Pavo cristatus (Indian peafowl), Perdix perdix (gray partridge), Phasianus colchicus (ring-neck pheasant), and Tympanchus phasianellus (sharp-tailed grouse)] together with that of Aythya americana (redhead) were examined using a maximum likelihood (ML) method. The ML analyses on the first/second codon positions, the third codon positions, and amino acid sequence consistently demonstrated that blue-breasted quail and Japanese quail are in the same phylogenetic cluster.     

In vitro reconstitution of wool intermediate filaments

Although the hard α-keratins of wool are recognized as members of the intermediate filaments by sequence comparison thus for all attempts on reconstitution of wool α-keratin in filaments in vitro have failed. Here we show the oxidative sulphitolysis rather than the previously used S-carboxymethylation is the method of choice to prepare α-keratin derivatives suitable for assembly experiments. Once the protecting S-sulpho group is removed by 2-mercaptoethanol in vitro filaments formation can be induced. Electron micrographs show filaments with a diameter of 7–11 nm as in all other intermediate filaments. Thus, filament formation of α-keratins does not require the presence of matrix proteins.     

The structure and expression of a gene encoding chick claw keratin.

A cDNA library was constructed from embryonic chick claw mRNA and a claw keratin (cKer)-encoding clone was isolated and sequenced. Subsequently, a genomic clone, containing four cKer-encoding genes (cKer) was isolated and one of the genes (cKer1) was completely sequenced. The cKerl gene appears to be differentially expressed in the keratinizing tissue appendages of the embryonic chick, being abundantly expressed in the claw and at a low level in feather tissue. Comparison of the deduced amino acid (aa) sequence of the cKer to those of feather (fKer) and scale keratins (sKer) showed that the regions conserved between fKer and sKer are also found in the cKer. The glycine-rich as repeat region characteristic of sKer is also present in a shortened form in the cKer sequence. Like the fKer genes (fKer) and the feather histidine-rich protein-encoding gene (HRP), the cKer1 gene also contains one intron which interrupts the 5'-noncoding region at an equivalent position to that found in the fKer and HRP genes. Genomic Southern analysis using the cKer cDNA as a probe indicated the presence of several related genes in the chick genome.     

There and back again: switching between host orders by avian body lice (Ischnocera: Goniodidae)

Studies of major switches by parasites between highly divergent host lineages are important for understanding new opportunities for parasite diversification. One such major host switch is inferred for avian feather lice (Ischnocera) in the family Goniodidae, which parasitize two distantly‐related groups of birds: Galliformes (pheasants, quail, partridges, etc.) and Columbiformes (pigeons and doves). Although there have been several cophylogenetic studies of lice at the species level, few studies have focused on such broad evolutionary patterns and major host‐switching events. Using a phylogeny based on DNA sequences for goniodid feather lice, we investigated the direction of this major host switch. Unexpectedly, we found that goniodid feather lice have switched host orders, not just once, but twice. A primary host switch occurred from Galliformes to Columbiformes, leading to a large radiation of columbiform body lice. Subsequently, there was also a host switch from Columbiformes back to Galliformes, specifically to megapodes in the Papua–Australasian region. The results of the present study further reveal that, although morphologically diagnosable lineages are supported by molecular data, many of the existing genera are not monophyletic and a revision of generic limits is needed. © 2011 The Linnean Society of London, Biological Journal of the Linnean Society, 2011, 102 , 614–625.     

Molecular cloning and characterization of novel centromeric repetitive DNA sequences in the blue-breasted quail (Coturnix chinensis,Galliformes)

A new family of centromeric highly repetitive DNA sequences was isolated from EcoRI-digested genomic DNA of the blue-breasted quail (Coturnix chinensis, Galliformes), and characterized by filter hybridization and chromosome in situ hybridization. The repeated elements were divided into two types by nucleotide length and chromosomal distribution; the 578-bp element predominantly localized to microchromosomes and the 1,524-bp element localized to chromosomes 1 and 2. The 578-bp element represented tandem arrays and did not hybridize to genomic DNAs of other Galliformes species, chicken (Gallus gallus), Japanese quail (Coturnix japonica) and guinea fowl (Numida meleagris). On the other hand, the 1,524-bp element was not organized in tandem arrays, and did hybridize to the genomic DNAs of three other Galliformes species, suggesting that the 1,524-bp element is highly conserved in the Galliformes. The 578-bp element was composed of basic 20-bp internal repeats, and the consensus nucleotide sequence of the internal repeats had homologies to the 41-42 bp CNM repeat and the XHOI family repeat of chicken. Our data suggest that the microchromosome-specific highly repetitive sequences of the blue-breasted quail and chicken were derived from a common ancestral sequence, and that they are one of the major and essential components of chromosomal heterochromatin in Galliformes species.     

Molecular evolutionary genomics of birds

Insight into the molecular evolution of birds has been offered by the steady accumulation of avian DNA sequence data, recently culminating in the first draft sequence of an avian genome, that of chicken. By studying avian molecular evolution we can learn about adaptations and phenotypic evolution in birds, and also gain an understanding of the similarities and differences between mammalian and avian genomes. In both these lineages, there is pronounced isochore structure with highly variable GC content. However, while mammalian isochores are decaying, they are maintained in the chicken lineage, which is consistent with a biased gene conversion model where the high and variable recombination rate of birds reinforces heterogeneity in GC. In Galliformes, GC is positively correlated with the rate of nucleotide substitution; the mean neutral mutation rate is 0.12-0.15% at each site per million years but this estimate comes with significant local variation in the rate of mutation. Comparative genomics reveals lower d(N)/d(S) ratios on micro- compared to macrochromosomes, possibly due to population genetic effects or a non-random distribution of genes with respect to chromosome size. A non-random genomic distribution is shown by genes with sex-biased expression, with male-biased genes over-represented and female-biased genes under-represented on the Z chromosome. A strong effect of selection is evident on the non-recombining W chromosome with high d(N)/d(S) ratios and limited polymorphism. Nucleotide diversity in chicken is estimated at 4-5 x 10(-3) which might be seen as surprisingly high given presumed bottlenecks during domestication, but is lower than that recently observed in several natural populations of other species. Several important aspects of the molecular evolutionary process of birds remain to be understood and it can be anticipated that the upcoming genome sequence of a second bird species, the zebra finch, as well as the integration of data on gene expression, shall further advance our knowledge of avian evolution.