A DNA Replication Mechanism Can Explain Structural Variation at the Pigeon Recessive Red Locus.

For species to adapt to their environment, evolution must act upon genetic variation that is present in the population. Elucidating the molecular mechanisms that give rise to this variation is thus of crucial importance for understanding how organisms evolve. In addition to variation caused by point mutations, structural variation (deletions, duplications, inversions, translocations) is also an important source of variety. Mechanisms involving recombination, transposition and retrotransposition, and replication have been proposed for generating structural variation, and each are capable of explaining certain rearrangements. In this study, we conduct a detailed analysis of two partially overlapping rearrangements (e1 and e2 allele) in domestic rock pigeon (Columba livia) which are both associated with the recessive red phenotype. We find that a replicative mechanism is best able to explain the complex architecture of the e1 allele, and is also compatible with the simpler architecture of the e2 allele as well.

SOX10 regulates multiple genes to direct eumelanin versus pheomelanin production in domestic rock pigeon.

The domesticated rock pigeon (Columba livia) has been bred for hundreds of years to display an immense variety of ornamental attributes such as feather color and color patterns. Color is influenced by multiple loci that impact the type and amount of melanin deposited on the feathers. Pigeons homozygous for the "recessive red" mutation, which causes downregulation of Sox10, display brilliant red feathers instead of blue/black feathers. Sox10 encodes a transcription factor important for melanocyte differentiation and function, but the genes that mediate its promotion of black versus red pigment are unknown. Here, we present a transcriptomic comparison of regenerating feathers from wild-type and recessive red pigeons to identify candidate SOX10 targets. Our results identify both known and novel targets, including many genes not previously implicated in pigmentation. These data highlight the value of using novel, emerging model organisms to gain insight into the genetic basis of pigment variation.

Genetic tests reveal extra-pair paternity among Gentoo penguins (Pyogoscelis papua ellsworthii) at Loveland Living Planet Aquarium: Implications for ex situ colony management.

Maintenance of ex situ populations for species conservation is a collaborative effort involving multiple agencies, institutions, and individuals around the world. Gentoo penguins (Pyogoscelis papua) are one species involved in such a conservation effort, and a Species Survival Plan (SSP) has been put in place by the Association of Zoos and Aquariums (AZA) to foster their long-term sustainability. As a part of their SSP, a Breeding and Transfer Plan has been created to support interagency exchanges of specimens. These transfers are vital to the demographic health and stability of ex situ populations, as well as the maintenance of genetic diversity. In populations such as the Gentoo, where social monogamy exists, paternal inferences of offspring are usually made through observation of birds' social behavior. However, social monogamy does not guarantee reproductive monogamy. In this study, we utilize Illumina high-throughput DNA sequencing to genetically test the postulated paternity of Gentoo penguins born at Loveland Living Planet Aquarium (LLPA) in Draper, UT. While our data support the majority of the postulated relationships, we did identify two offspring that were the result of extra-pair paternity (EPP). The results of this research highlight the importance of genetic tests to validate pedigrees used in SSPs, to provide more-accurate data for the support of species conservation.

Pigeonetics takes flight: Evolution, development, and genetics of intraspecific variation.

Intensive artificial selection over thousands of years has produced hundreds of varieties of domestic pigeon. As Charles Darwin observed, the morphological differences among breeds can rise to the magnitude of variation typically observed among different species. Nevertheless, different pigeon varieties are interfertile, thereby enabling forward genetic and genomic approaches to identify genes that underlie derived traits. Building on classical genetic studies of pigeon variation, recent molecular investigations find a spectrum of coding and regulatory alleles controlling derived traits, including plumage color, feather growth polarity, and limb identity. Developmental and genetic analyses of pigeons are revealing the molecular basis of variation in a classic example of extreme intraspecific diversity, and have the potential to nominate genes that control variation among other birds and vertebrates in general.

Molecular shifts in limb identity underlie development of feathered feet in two domestic avian species.

Birds display remarkable diversity in the distribution and morphology of scales and feathers on their feet, yet the genetic and developmental mechanisms governing this diversity remain unknown. Domestic pigeons have striking variation in foot feathering within a single species, providing a tractable model to investigate the molecular basis of skin appendage differences. We found that feathered feet in pigeons result from a partial transformation from hindlimb to forelimb identity mediated by cis-regulatory changes in the genes encoding the hindlimb-specific transcription factor Pitx1 and forelimb-specific transcription factor Tbx5. We also found that ectopic expression of Tbx5 is associated with foot feathers in chickens, suggesting similar molecular pathways underlie phenotypic convergence between these two species. These results show how changes in expression of regional patterning genes can generate localized changes in organ fate and morphology, and provide viable molecular mechanisms for diversity in hindlimb scale and feather distribution.

Syndactyly in a novel Fras1(rdf) mutant results from interruption of signals for interdigital apoptosis.

BACKGROUND: Fras1 encodes an extracellular matrix protein that is critical for the establishment of the epidermal basement membrane during gestation. In humans, mutations in FRAS1 cause Fraser Syndrome (FS), a pleiotropic condition with many clinical presentations such as limb, eye, kidney, and craniofacial deformations. Many of these defects are mimicked by loss of Fras1 in mice, and are preceded by the formation of epidermal blisters in utero. RESULTS: In this study, we identified a novel ENU-derived rounded foot (rdf) mouse mutant with highly penetrant hindlimb soft-tissue syndactyly, among other structural defects. Mapping and sequencing revealed that rdf is a novel loss-of-function nonsense allele of Fras1 (Fras1(rdf)). Focusing on the limb, we found that the Fras1(rdf) syndactyly phenotype originates from loss of interdigital cell death (ICD). Despite normal expression of bone morphogenetic protein (BMP) ligands and their receptors, the BMP downstream target gene Msx2, which is also necessary and sufficient to promote ICD, was down-regulated in the interdigital regions of Fras1(rdf) hindlimb buds. CONCLUSIONS: The close correlation between limb bud epidermal blistering, decreased Msx2 expression, and reduced ICD in the Fras1(rdf) hindlimb buds suggests that epithelium detachment from the mesenchyme may create a physical gap that interrupts the transmission of BMP, among other signals, resulting in soft tissue syndactyly.

Wham: Identifying Structural Variants of Biological Consequence.

Existing methods for identifying structural variants (SVs) from short read datasets are inaccurate. This complicates disease-gene identification and efforts to understand the consequences of genetic variation. In response, we have created Wham (Whole-genome Alignment Metrics) to provide a single, integrated framework for both structural variant calling and association testing, thereby bypassing many of the difficulties that currently frustrate attempts to employ SVs in association testing. Here we describe Wham, benchmark it against three other widely used SV identification tools-Lumpy, Delly and SoftSearch-and demonstrate Wham's ability to identify and associate SVs with phenotypes using data from humans, domestic pigeons, and vaccinia virus. Wham and all associated software are covered under the MIT License and can be freely downloaded from github (https://github.com/zeeev/wham), with documentation on a wiki (http://zeeev.github.io/wham/). For community support please post questions to https://www.biostars.org/.

Convergent Evolution of Head Crests in Two Domesticated Columbids Is Associated with Different Missense Mutations in EphB2.

Head crests are important display structures in wild bird species and are also common in domesticated lineages. Many breeds of domestic rock pigeon (Columba livia) have crests of reversed occipital feathers, and this recessive trait is associated with a nonsynonymous coding mutation in the intracellular kinase domain of EphB2 (Ephrin receptor B2). The domestic ringneck dove (Streptopelia risoria) also has a recessive crested morph with reversed occipital feathers, and interspecific crosses between crested doves and pigeons produce crested offspring, suggesting a similar genetic basis for this trait in both species. We therefore investigated EphB2 as a candidate for the head crest phenotype of ringneck doves and identified a nonsynonymous coding mutation in the intracellular kinase domain that is significantly associated with the crested morph. This mutation is over 100 amino acid positions away from the crest mutation found in rock pigeons, yet both mutations are predicted to negatively affect the function of ATP-binding pocket. Furthermore, bacterial toxicity assays suggest that "crest" mutations in both species severely impact kinase activity. We conclude that head crests are associated with different mutations in the same functional domain of the same gene in two different columbid species, thereby representing striking evolutionary convergence in morphology and molecules.

Epistatic and combinatorial effects of pigmentary gene mutations in the domestic pigeon.

Understanding the molecular basis of phenotypic diversity is a critical challenge in biology, yet we know little about the mechanistic effects of different mutations and epistatic relationships among loci that contribute to complex traits. Pigmentation genetics offers a powerful model for identifying mutations underlying diversity and for determining how additional complexity emerges from interactions among loci. Centuries of artificial selection in domestic rock pigeons (Columba livia) have cultivated tremendous variation in plumage pigmentation through the combined effects of dozens of loci. The dominance and epistatic hierarchies of key loci governing this diversity are known through classical genetic studies, but their molecular identities and the mechanisms of their genetic interactions remain unknown. Here we identify protein-coding and cis-regulatory mutations in Tyrp1, Sox10, and Slc45a2 that underlie classical color phenotypes of pigeons and present a mechanistic explanation of their dominance and epistatic relationships. We also find unanticipated allelic heterogeneity at Tyrp1 and Sox10, indicating that color variants evolved repeatedly though mutations in the same genes. These results demonstrate how a spectrum of coding and regulatory mutations in a small number of genes can interact to generate substantial phenotypic diversity in a classic Darwinian model of evolution.

Genomic diversity and evolution of the head crest in the rock pigeon.

The geographic origins of breeds and the genetic basis of variation within the widely distributed and phenotypically diverse domestic rock pigeon (Columba livia) remain largely unknown. We generated a rock pigeon reference genome and additional genome sequences representing domestic and feral populations. We found evidence for the origins of major breed groups in the Middle East and contributions from a racing breed to North American feral populations. We identified the gene EphB2 as a strong candidate for the derived head crest phenotype shared by numerous breeds, an important trait in mate selection in many avian species. We also found evidence that this trait evolved just once and spread throughout the species, and that the crest originates early in development by the localized molecular reversal of feather bud polarity.

Signaling through BMP receptors promotes respiratory identity in the foregut via repression of Sox2.

The mammalian foregut gives rise to the dorsally located esophagus and stomach and the ventrally located trachea and lung. Proper patterning and morphogenesis of the common foregut tube and its derived organs is essential for viability of the organism at birth. Here, we show that conditional inactivation of BMP type I receptor genes Bmpr1a and Bmpr1b (Bmpr1a;b) in the ventral endoderm leads to tracheal agenesis and ectopic primary bronchi. Molecular analyses of these mutants reveal a reduction of ventral endoderm marker NKX2-1 and an expansion of dorsal markers SOX2 and P63 into the prospective trachea and primary bronchi. Subsequent genetic experiments show that activation of canonical WNT signaling, previously shown to induce ectopic respiratory fate in otherwise wild-type mice, is incapable of promoting respiratory fate in the absence of Bmpr1a;b. Furthermore, we find that inactivation of Sox2 in Bmpr1a;b mutants does not suppress ectopic lung budding but does rescue trachea formation and NKX2-1 expression. Together, our data suggest that signaling through BMPR1A;B performs at least two roles in early respiratory development: first, it promotes tracheal formation through repression of Sox2; and second, it restricts the site of lung bud initiation.

Patterning and plasticity in development of the respiratory lineage.

The mammalian respiratory lineage, consisting of the trachea and lung, originates from the ventral foregut in an early embryo. Reciprocal signaling interactions between the foregut epithelium and its associated mesenchyme guide development of the respiratory endoderm, from a naive sheet of cells to multiple cell types that line a functional organ. This review synthesizes current understanding of the early events in respiratory system development, focusing on three main topics: (1) specification of the respiratory system as a distinct organ of the endoderm, (2) patterning and differentiation of the nascent respiratory epithelium along its proximal-distal axis, and (3) plasticity of the respiratory cells during the process of development. This review also highlights areas in need of further study, including determining how early endoderm cells rapidly switch their responses to the same signaling cues during development, and how the general proximal-distal pattern of the lung is converted to fine-scale organization of multiple cell types along this axis.

beta-Catenin promotes respiratory progenitor identity in mouse foregut.

The mammalian respiratory system, consisting of both trachea and lung, initiates from the foregut endoderm. The molecular program that instructs endodermal cells to adopt the respiratory fate is not fully understood. Here we show that conditional inactivation of beta-Catenin (also termed Ctnnb1) in foregut endoderm leads to absence of both the trachea and lung due to a failure in maintaining the respiratory fate. In converse, conditional expression of an activated form of beta-Catenin leads to expansion of Nkx2.1, an early marker for the trachea and lung, into adjacent endoderm including the stomach epithelium. Analyses of these mutants show that the loss or gain of trachea/lung progenitor identity is accompanied by an expansion or contraction of esophagus/stomach progenitor identity, respectively. Our findings reveal an early role for beta-Catenin in the establishment of respiratory progenitors in mouse foregut endoderm.

Fibroblast growth factor 9 signaling inhibits airway smooth muscle differentiation in mouse lung.

In mammalian lungs, airway smooth muscle cells (airway SMCs) are present in the proximal lung adjacent to bronchi and bronchioles, but are absent in the distal lung adjacent to terminal sacs that expand during gas exchange. Evidence suggests that this distribution is essential for the formation of a functional respiratory tree, but the underlying genetic mechanism has not been elucidated. In this study, we test the hypothesis that fibroblast growth factor 9 (Fgf9) signaling is essential to restrict SMC differentiation to the proximal lung. We show that loss of Fgf9 or conditional inactivation of Fgf receptors (Fgfr) 1 and 2 in mouse lung mesenchyme results in ectopic SMCs. Our data support a model where FGF9 maintains a SMC progenitor population by suppressing differentiation and promoting growth. This model also represents our findings on the genetic relationship between FGF9 and sonic hedgehog (SHH) in the establishment of airway SMC pattern.